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Title: The Works of Francis Maitland Balfour, Volume 1
       Separate Memoirs

Author: Francis Maitland Balfour

Editor: Michael Foster
        Adam Sedgwick

Release Date: November 12, 2012 [EBook #41357]

Language: English

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THE WORKS

OF

FRANCIS MAITLAND BALFOUR.

VOL. I.

Memorial Edition.

Cambridge:

PRINTED BY C. J. CLAY, M.A. AND SON,
AT THE UNIVERSITY PRESS.

Memorial Edition.

THE WORKS

OF

FRANCIS MAITLAND BALFOUR.

M.A., LL.D., F.R.S.,

FELLOW OF TRINITY COLLEGE,
AND PROFESSOR OF ANIMAL MORPHOLOGY IN THE UNIVERSITY OF
CAMBRIDGE.

EDITED BY

M. FOSTER, F.R.S.,

PROFESSOR OF PHYSIOLOGY IN THE UNIVERSITY OF CAMBRIDGE;

AND

ADAM SEDGWICK, M.A.,

FELLOW AND LECTURER OF TRINITY COLLEGE, CAMBRIDGE.

VOL. I.

SEPARATE MEMOIRS.

London:

MACMILLAN AND CO.
1885

[The Right of Translation is reserved.]

PREFACE.

Upon the death of Francis Maitland Balfour, a desire very naturally arose among his friends and admirers to provide some memorial of him. And, at a public meeting held at Cambridge in October 1882, the Vice-Chancellor presiding, and many distinguished men of science being present, it was decided to establish a 'Balfour Fund' the proceeds of which should be applied: firstly to maintain a studentship, the holder of which should devote himself to original research in Biology, especially in Animal Morphology, and secondly, 'by occasional grants of money, to further in other ways original research in the same subject'. The sum of £8446 was subsequently raised; this was, under certain conditions, entrusted to and accepted by the University of Cambridge; and the first 'Balfour student' was appointed in October 1883.

The publication of Balfour's works in a collected form was not proposed as an object on which part of the fund should be expended, since his family had expressed their wish to take upon themselves the charge of arranging for a memorial edition of their brother's scientific writings. [Pg ii] That edition, with no more delay than circumstances have rendered necessary, is now laid before the public. It comprises four volumes.

The first volume contains, in chronological order, all Balfour's scattered original papers, including those published by him in conjunction with his pupils, as well as the Monograph on the Elasmobranch Fishes. The last memoir in the volume, that on the Anatomy and Development of Peripatus Capensis, was published after his death, from his notes and drawings, with additions by Prof. Moseley and Mr Adam Sedgwick, who prepared the manuscript for publication. To the volume is prefixed an introductory biographical notice.

The second and third volumes are the two volumes of the Comparative Embryology reprinted from the original edition without alteration, save the correction of obvious misprints and omissions.

The fourth volume contains the plates illustrating the memoirs contained in Vol. 1. We believe that we are consulting the convenience of readers in adopting this plan, rather than in distributing the plates among the memoirs to which they belong. To assist the reader the explanations of these plates have been given twice: at the end of the memoir to which they belong (in the case of the Monograph on Elasmobranch Fishes at the end of each separate chapter), and in the volume of plates.

All the figures of these plates had to be redrawn on the stone, and our best thanks are due to the Cambridge Scientific Instrument Company for the pains which they have taken in executing this work. We are also indebted to the Committee of Publication of the Zoological Society for the gift of electrotypes of the woodcuts illustrating memoir no. XX. of Vol. 1. [Pg iii]

Several photographs of Balfour, taken at different times of his life, the last shortly before his death, are in the possession of his relatives and friends; but these, in the opinion of many, leave much to be desired.

There is also a portrait of him in oils painted since his death by Mr John Collier, A.R.A., and Herr Hildebrand of Florence has executed a posthumous bust in bronze[1]. The portrait which forms the frontispiece of Vol. 1. has been drawn on stone by Mr E. Wilson of the Cambridge Scientific Instrument Company, after the latest photograph. Should it fail, in the eyes of those who knew Balfour well, to have reproduced with complete success his features and expression, we would venture to ask them to bear in mind the acknowledged difficulties of posthumous portraiture.

[1] In possession of the family. Copies also exist in the Library of Trinity College, and in the Morphological Laboratory, at Cambridge.

[Pg iv]
[Pg v]

Francis Maitland Balfour, the sixth child and third son of James Maitland Balfour of Whittinghame, East Lothian, and Lady Blanche, daughter of the second Marquis of Salisbury, was born at Edinburgh, during a temporary stay of his parents there, on the 10th November, 1851. He can hardly be said to have known his father, who died of consumption in 1856, at the early age of thirty-six, and who spent the greater part of the last two years of his life at Madeira, separated from the younger children who remained at home. He fancied at one time that he had inherited his father's constitution; and this idea seems to have spurred him on to achieve early what he had to do. But, though there was a period soon after he went to College, during which he seemed delicate, and the state of his health caused considerable anxiety to his friends, he eventually became fairly robust, and that in spite of labours which greatly taxed his strength.

The early years of his life were spent chiefly at Whittinghame under the loving care of his mother. She made it a point to attempt to cultivate in all her children some taste for natural science, especially for natural history, and in this she was greatly helped by the boys' tutor, Mr J. W. Kitto. They were encouraged to make collections and to form a museum, and the fossils found in the gravel spread in front of the house served as the nucleus of a geological series. Frank soon became greatly interested in these things, and indeed they may be said to have formed the beginnings of his scientific career. At all events there was thus awakened in him a love for geology, which science continued to be his favorite study all through his [Pg 2] boyhood, and interested him to the last. He was most assiduous in searching for fossils in the gravel and elsewhere, and so great was his love for his collections that while as yet quite a little boy the most delightful birthday present he could think of was a box with trays and divisions to hold his fossils and specimens. His mother, thinking that his fondness for fossils was a passing fancy and that he might soon regret the purchase of the box, purposely delayed the present. But he remained constant to his wish and in time received his box. He must at this time have been about seven or eight years old. In the children's museum, which has been preserved, there are specimens labelled with his childish round-hand, such as a piece of stone with the label marks of some shels; and his sister Alice, who was at that time his chief companion, remembers discussing with him one day after the nursery dinner, when he was about nine years old, whether it were better to be a geologist or a naturalist, he deciding for the former on the ground that it was better to do one thing thoroughly than to attempt many branches of science and do them imperfectly.

Besides fossils, he collected not only butterflies, as do most boys at some time or other, but also birds; and he with his sister Alice, being instructed in the art of preparing and preserving skins, succeeded in making a very considerable collection. He thus acquired before long not only a very large but a very exact knowledge of British birds.

In the more ordinary work of the school-room he was somewhat backward. This may have been partly due to the great difficulty he had in learning to write, for he was not only left-handed but, in his early years, singularly inapt in acquiring particular muscular movements, learning to dance being a great trouble to him. Probably however the chief reason was that he failed to find any interest in the ordinary school studies. He fancied that the family thought him stupid, but this does not appear to have been the case.

In character he was at this time quick tempered, sometimes even violent, and the energy which he shewed in after life even thus early manifested itself as perseverance, which, when he was crossed, often took on the form of obstinacy, causing at times no little trouble to his nurses and tutors. But he was at the [Pg 3] same time warm-hearted and affectionate; full of strong impulses, he disliked heartily and loved much, and in his affections was wonderfully unselfish, wholly forgetting himself in his thought for others, and ready to do things which he disliked to please those whom he loved. Though, as we have said, somewhat clumsy, he was nevertheless active and courageous; in learning to ride he shewed no signs of fear, and boldly put his pony to every jump which was practicable.

In 1861 he was sent to the Rev. C. G. Chittenden's preparatory school at Hoddesden in Hertfordshire, and here the qualities which had been already visible at home became still more obvious. He found difficulty not only in writing but also in spelling, and in the ordinary school-work he took but little interest and made but little progress.

In 1865 he was moved to Harrow and placed in the house of the Rev. F. Rendall. Here, as at Hoddesden, he did not show any great ability in the ordinary school studies, though as he grew older his progress became more marked. But happily he found at Harrow an opportunity for cultivating that love of scientific studies which was yearly growing stronger in him. Under the care of one of the Masters, Mr G. Griffith, the boys at Harrow were even then taught the elements of natural science. The lessons were at that time, so to speak, extra-academical, carried on out of school hours; nevertheless, many of the boys worked at them with diligence and even enthusiasm, and among these Balfour became conspicuous, not only by his zeal but by his ability. Griffith was soon able to recognize the power of his new pupil, and thus early began to see that the pale, earnest, somewhat clumsy-handed lad, though he gave no promise of being a scholar in the narrower sense of the word, had in him the makings of a man of science. Griffith chiefly confined his teaching to elementary physics and chemistry with some little geology, but he also encouraged natural history studies and began the formation of a museum of comparative anatomy. Balfour soon began to be very zealous in dissecting animals, and was especially delighted when the Rev. A. C. Eaton, the well-known entomologist, on a visit to Harrow, initiated Griffith's pupils in the art of dissecting under water. The dissection of a caterpillar in this way was probably an [Pg 4] epoch in Balfour's life. Up to that time his rough examination of such bodies had revealed to him nothing more than what in school-boy language he spoke of as squash; but when under Eaton's deft hands the intricate organs of the larval Arthropod floated out under water and displayed themselves as a labyrinth of threads and sheets of silvery whiteness a new world of observation opened itself up to Balfour, and we may probably date from this the beginning of his exact morphological knowledge.

While thus learning the art of observing, he was at the same time developing his power of thinking. He was by nature fond of argument, and defended with earnestness any opinions which he had been led to adopt. He was very active in the Harrow Scientific Society, reading papers, taking part in the discussions, and exhibiting specimens. He gained in 1867 a prize for an essay on coal, and when, in 1868, Mr Leaf offered a prize (a microscope) for the best account of some locality visited by the writer during the Easter Holidays, two essays sent in, one by Balfour, the other by his close friend, Mr Arthur Evans, since well known for his researches in Illyria, were found to be of such unusual merit that Prof. Huxley was specially requested to adjudicate between them. He judged them to be of equal merit, and a prize was given to each. The subject of Balfour's essay was The Geology and Natural History of East Lothian. When biological subjects were discussed at the Scientific Society, Balfour appears to have spoken as a most uncompromising opponent of the views of Mr Charles Darwin, little thinking that in after life his chief work would be to develop and illustrate the doctrine of evolution.

The years at Harrow passed quickly away, Balfour making fair, but perhaps not more than fair, progress in the ordinary school learning. In due course however he reached the upper sixth form, and in his last year, became a monitor. At the same time his exact scientific knowledge was rapidly increasing. Geology still continued to be his favorite study, and in this he made no mean progress. During his last years at Harrow he and his brother Gerald worked out together some views concerning the geology of their native county. These views they ultimately embodied in a paper, which was published in their joint names in the Geological Magazine for 1872, under the title [Pg 5] of Some Points in the Geology of the East Lothian Coast, and which was in itself a work of considerable promise. Geology however was beginning to find a rival in natural history. Much of his holiday time was now spent in dredging for marine animals along the coast off Dunbar. Each specimen thus obtained was carefully determined and exact records were kept of the various 'finds,' so that the dredgings (which were zealously continued after he had left Harrow and gone to Cambridge) really constituted a serious study of the fauna of this part of the coast. They also enabled him to make a not inconsiderable collection of shells, in the arrangement of which he was assisted by his sister Evelyn, of crustacea and of other animals.

Both to the masters and to his schoolfellows he became known as a boy of great force of character. Among the latter his scrupulous and unwavering conscientiousness made him less popular perhaps than might have been expected from his bright kindly manner and his unselfish warmheartedness. In the incidents of school life a too strict conscience is often an inconvenience, and the sternness and energy with which Balfour denounced acts of meanness and falsehood were thought by some to be unnecessarily great. He thus came to be feared rather than liked by many, and comparatively few grew to be sufficiently intimate with him to appreciate the warmth of his affections and the charm of his playful moments.

At the Easter of 1870 he passed the entrance examination at Trinity College, Cambridge, and entered into residence in the following October. His college tutor was Mr J. Prior, but he was from the first assisted and guided in his studies by his friend, Mr Marlborough Pryor, an old Harrow boy, who in the same October had been, on account of his distinction in Natural Science, elected a Fellow of the College, in accordance with certain new regulations which then came into action for the first time, and which provided that every three years one of the College Fellowships should be awarded for excellence in some branch or branches of Natural Science, as distinguished from mathematics, pure or mixed. During the whole of that year and part of the next Mr Marlborough Pryor remained in residence, and his influence in wisely directing Balfour's studies had a most beneficial effect on the latter's progress.

[Pg 6] During his first term Balfour was occupied in preparation for the Previous Examination; and this he successfully passed at Christmas. After that he devoted himself entirely to Natural Science, attending lectures on several branches. During the Lent term he was a very diligent hearer of the lectures on Physiology which I was then giving as Trinity Prælector, having been appointed to that post in the same October that Balfour came into residence. At this time he was not very strong, and I remember very well noticing among my scanty audience, a pale retiring student, whose mind seemed at times divided between a desire to hear the lecture and a feeling that his frequent coughing was growing an annoyance to myself and the class. This delicate-looking student, I soon learnt, was named Balfour, and when the Rev. Coutts Trotter, Mr Pryor and myself came to examine the candidates for the Natural Science Scholarships which were awarded at Easter, we had no difficulty in giving the first place to him. In point of knowledge, and especially in the thoughtfulness and exactitude displayed in his papers and work, he was very clearly ahead of his competitors.

During the succeeding Easter term and the following winter he appears to have studied physics, chemistry, geology and comparative anatomy, both under Mr Marlborough Pryor and by means of lectures. He also continued to attend my lectures, but though I gradually got to know him more and more we did not become intimate until the Lent term of 1872. He had been very much interested in some lectures on embryology which I had given, and, since Marlborough Pryor had left or was about to leave Cambridge, he soon began to consult me a good deal about his studies. He commenced practical histological and embryological work under me, and I remember very vividly that one day when we were making a little excursion in search of nests and eggs of the stickleback in order that he might study the embryology of fishes, he definitely asked my opinion as to whether he might take up a scientific career with a fair chance of success. I had by this time formed a very high opinion of his abilities, and learning then for the first time that he had an income independent of his own exertions, my answer was very decidedly a positive one. Soon after, feeling more and [Pg 7] more impressed with his power and increasingly satisfied both with his progress in biological studies and his sound general knowledge of other sciences, anxious also, it may be, at the same time that as much original inquiry as possible should be carried on at Cambridge in my department, I either suggested to him or acquiesced in his own suggestion that he should at once set to work on some distinct research; and as far as I remember the task which I first proposed to him was an investigation of the layers of the blastoderm in the chick. It must have been about the same time that I proposed to him to join me in preparing for publication a small work on Embryology, the materials for this I had ready to hand in a rough form as lectures which I had previously given. To this proposal he enthusiastically assented, and while the lighter task of writing what was to be written fell to me, he undertook to work over as far as was possible the many undetermined points and unsatisfactory statements across which we were continually coming.

During his two years at College his health had improved; though still hardly robust and always in danger of overworking himself, he obviously grew stronger. He rejoiced exceedingly in his work, never tiring of it, and was also making his worth felt among his fellow students, and especially perhaps among those of his own college whose studies did not lie in the same direction as his own. At this time he must have been altogether happy, but a sorrow now came upon him. His mother, to whom he was passionately attached, and to whose judicious care in his early days not only the right development of his strong character but even his scientific leanings were due, had for some time past been failing in health, though her condition caused no immediate alarm. In May 1872, however, she died quite suddenly from unsuspected heart disease. Her loss was a great blow to him, and for some time afterward I feared his health would give way; but he bore his grief quietly and manfully and threw himself with even increased vigour into his work.

During the academic session of 1872-3, he continued steadily at work at his investigations, and soon began to make rapid progress. At the beginning he had complained to me about what he considered his natural clumsiness, and expressed a fear [Pg 8] that he should never be able to make satisfactory microscopic sections; as to his being able to make drawings of his dissections and microscopical preparations, he looked upon that at first as wholly impossible. I need hardly say that in time he acquired great skill in the details of microscopical technique, and that his drawings, if wanting in so-called artistic finish, were always singularly true and instructive. While thus struggling with the details which I could teach him, he soon began to manifest qualities which no teacher could give him. I remember calling his attention to Dursy's paper on the Primitive Streak, and suggesting that he should work the matter over, since if such a structure really existed, it must, most probably, have great morphological significance. I am free to confess that I myself rather doubted the matter, and a weaker student might have been influenced by my preconceptions. Balfour, however, thus early had the power of seeing what existed and of refusing to see what did not exist. He was soon able to convince me that Dursy's streak was a reality, and the complete working out of its significance occupied his thoughts to the end of his days.

The results of these early studies were made known in three papers which appeared in the Quarterly Journal of Microscopical Science for July 1873, and will be found in the beginning of this volume. The summer and autumn of that year were spent partly in a visit to Finland, in company with his friend and old school-fellow Mr Arthur Evans, and partly in formal preparation for the approaching Tripos examination. Into this preparation Balfour threw himself with characteristic energy, and fully justified my having encouraged his spending so much of the preceding time in original research, not only by the rapidity with which he accumulated the stock of knowledge of various kinds necessary for the examination but also by the manner in which he acquitted himself at the trial itself. At that time the position of the candidates in the Natural Sciences Tripos was determined by the total number of marks, and Balfour was placed second, the first place being gained by H. Newell Martin of Christ's College, now Professor at Baltimore, U.S.A. In the examination, in which I took part, Balfour did not write much, and he had not yet learnt the art of putting his statements in the best [Pg 9] possible form; he won his position chiefly by the firm thought and clear insight which was present in almost all his answers.

The examination was over in the early days of Dec. 1873 and Balfour was now free to devote himself wholly to his original work. Happily, the University had not long before secured the use of two of the tables at the then recently founded Stazione Zoologica at Naples. And upon the nomination of the University, Balfour, about Christmas, started for Naples in company with his friend Mr A. G. Dew-Smith, also of Trinity College. The latter was about to carry on some physiological observations; Balfour had set himself to work out as completely as he could the embryology of Elasmobranch fishes, about which little was at that time known, but which, from the striking characters of the adult animals could not help proving of interest and importance.

From his arrival there at Christmas 1873 until he left in June 1874, he worked assiduously, and with such success, that as the result of the half-year's work he had made a whole series of observations of the greatest importance. Of these perhaps the most striking were those on the development of the urogenital organs, on the neurenteric canal, on the development of the spinal nerves, on the formation of the layers and on the phenomena of segmentation, including a history of the behaviour of nuclei in cell division. He returned home laden with facts and views both novel and destined to influence largely the progress of embryology.

In August of the same year he attended the meeting of the British Association for the Advancement of Science at Belfast; and the account he then gave of his researches formed one of the most important incidents at the Biological Section on that occasion.

In the September of that year the triennial fellowship for Natural Science was to be awarded at Trinity College, and Balfour naturally was a candidate. The election was, according to the regulations, to be determined partly by the result of an examination in various branches of science, and partly by such evidence of ability and promise as might be afforded by original work, published or in manuscript. He spent the remainder of the autumn in preparation for this examination. But when the [Pg 10] examination was concluded it was found that in his written answers he had not been very successful; he had not even acquitted himself so well as in the Tripos of the year before, and had the election been determined by the results of the examination alone, the examiners would have been led to choose the gentleman who was Balfour's only competitor. The original work however which Balfour sent in, including a preliminary account of the discoveries made at Naples, was obviously of so high a merit and was spoken of in such enthusiastic terms by the External Referee Prof. Huxley, that the examiners did not hesitate for a moment to neglect altogether the formal written answers (and indeed the papers of questions were only introduced as a safeguard, or as a resource in case evidence of original power should be wanted) and unanimously recommended him for election. Accordingly he was elected Fellow in the early days of October.

Almost immediately after, the little book on Embryology appeared, on which he and I had been at work, he doing his share even while his hands and mind were full of the Elasmobranch inquiry. The title-page was kept back some little time in order that his name might appear on it with the addition of Fellow of Trinity, a title of which he was then, and indeed always continued to be, proud. He also published in the October number of the Quarterly Journal of Microscopical Science a preliminary account of his Elasmobranch researches.

He and his friends thought that after these almost incessant labours, and the excitement necessarily contingent upon the fellowship election, he needed rest and change. Accordingly on the 17th of October he started with his friend Marlborough Pryor on a voyage to the west coast of South America. They travelled thither by the Isthmus of Panama, visited Peru and Chili, and returned home along the usual route by the Horn; reaching England some time in Feb. 1875.

Refreshed by this holiday, he now felt anxious to complete as far as possible his Elasmobranch work, and very soon after his return home, in fact in March, made his way again to Naples, where he remained till the hot weather set in in May. [Pg 11] On his return to Cambridge, he still continued working on the Elasmobranchii, receiving material partly from Naples, partly from the Brighton Aquarium, the then director of which, Mr Henry Lee, spared no pains to provide him both with embryo and adult fishes. While at Naples, he communicated to the Philosophical Society at Cambridge a remarkable paper on The Early Stages of Vertebrates, which was published in full in the Quarterly Journal of Microscopical Science, July, 1875; he also sent me a paper on The Development of the Spinal Nerves, which I communicated to the Royal Society, and which was subsequently published in the Philosophical Transactions of 1876. He further wrote in the course of the summer and published in the Journal of Anatomy and Physiology in October, 1875, a detailed account of his Observations and Views on the Development of the Urogenital Organs.

Some time in August of the same year he started in company with Mr Arthur Evans and Mr J. F. Bullar for a second trip to Finland, the travellers on this occasion making their way into regions very seldom visited, and having to subsist largely on the preserved provisions which they carried with them, and on the produce of their rods and guns. From a rough diary which Balfour kept during this trip it would appear that while enjoying heartily the fun of the rough travelling, he occupied himself continually with observations on the geology and physical phenomena of the country, as well as on the manners, antiquities, and even language of the people. It was one of his characteristic traits, a mark of the truly scientific bent of his mind, of his having, as Dohrn soon after Balfour's first arrival at Naples said, 'a real scientific head,' that every thing around him wherever he was, incited him to careful exact observation, and stimulated him to thought.

In the early part of the Long Vacation of the same year he had made his first essay in lecturing, having given a short course on Embryology in a room at the New Museums, which I then occupied as a laboratory. Though he afterwards learnt to lecture with great clearness he was not by nature a fluent speaker, and on this occasion he was exceedingly [Pg 12] nervous. But those who listened to him soon forgot these small defects as they began to perceive the knowledge and power which lay in their new teacher.

Encouraged by the result of this experiment, he threw himself, in spite of the heavy work which the Elasmobranch investigation was entailing, with great zeal into an arrangement which Prof. Newton, Mr J. W. Clark and myself had in course of the summer brought about, that he and Mr A. Milnes Marshall, since Professor at Owens College, Manchester, should between them give a course on Animal Morphology, with practical instruction, Prof. Newton giving up a room in the New Museums for the purpose.

In the following October (1875) upon Balfour's return from Finland, these lectures were accordingly begun and carried on by the two lecturers during the Michaelmas and Lent Terms. The number of students attending this first course, conducted on a novel plan, was, as might be expected, small, but the Lent Term did not come to an end before an enthusiasm for morphological studies had been kindled in the members of the class.

The ensuing Easter term (1876) was spent by Balfour at Naples, in order that he might carry on towards completion his Elasmobranch work. He had by this time determined to write as complete a monograph as he could of the development of these fishes, proposing to publish it in instalments in the Journal of Anatomy and Physiology, and subsequently to gather together the several papers into one volume. The first of these papers, dealing with the ovum, appeared in Jan. 1876; most of the numbers of the Journal during that and the succeeding year contained further portions; but the complete monograph did not leave the publisher's hands until 1878.

He returned to England with his pupil and friend Mr J. F. Bullar some time in the summer; on their way home they passed through Switzerland, and it was during the few days which he then spent in sight of the snow-clad hills that the beginnings of a desire for that Alpine climbing, which was destined to be so disastrous, seem to have been kindled in him.

In October, 1876, he resumed the lectures on Morphology, taking the whole course himself, his colleague, Mr Marshall, [Pg 13] having meanwhile left Cambridge. Indeed, from this time onward, he may be said to have made these lectures, in a certain sense, the chief business of his life. He lectured all three terms, devoting the Michaelmas and Lent terms to a systematic course of Animal Morphology, and the Easter term to a more elementary course of Embryology. These lectures were given under the auspices of Prof. Newton; but Balfour's position was before long confirmed by his being made a Lecturer of Trinity College, the lectures which he gave at the New Museums, and which were open to all students of the University, being accepted in a liberal spirit by the College as equivalent to College Lectures. He very soon found it desirable to divide the morphological course into an elementary and an advanced course, and to increase the number of his lectures from three to four a week. Each lecture was followed by practical work, the students dissecting and examining microscopically, an animal or some animals chosen as types to illustrate the subject-matter of the lecture; and although Balfour had the assistance at first of one[2], and ultimately of several demonstrators, he himself put his hand to the plough, and after the lecture always spent some time in the laboratory among his pupils. Had Balfour been only an ordinary man, the zeal and energy which he threw into his lectures, and into the supervision of the practical work, added to the almost brotherly interest which he took in the individual development of every one of the pupils who shewed any love whatever for the subject, would have made him a most successful teacher. But his talents and powers were such as could not be hid even from beginners. His extensive and exact knowledge, the clearness with which in spite of, or shall I not rather say, by help of a certain want of fluency, he explained difficult and abstruse matters, the trenchant way in which he lay bare specious fallacies, and the presence in almost his every word of that power which belongs only to the man who has thought out for himself everything which he says, these things aroused and indeed could hardly fail to arouse in his hearers feelings which, except in the case of the very dullest, grew to be those of [Pg 14] enthusiasm. His class, at first slowly, but afterwards more rapidly, increased in numbers, and, what is of more importance, grew in quality. The room allotted to him soon became far too small and steps were taken to provide for him, for myself, whose wants were also urgent, and for the biological studies generally, adequate accommodation; but it was not until Oct. 1877 that we were able to take possession of the new quarters.

Even this new accommodation soon became insufficient, and in the spring of 1882 a new morphological laboratory was commenced in accordance with plans suggested by himself. He was to have occupied them in the October term, 1883, but did not live to see them finished.

As might have been expected from his own career, he regarded the mere teaching of what is known as a very small part of his duties as Lecturer; and as soon as any of his pupils became sufficiently advanced, he urged or rather led them to undertake original investigations; and he had the satisfaction before his death of seeing the researches of his pupils (such as those by Messrs. Bullar, Sedgwick, Mitzikuri, Haddon, Scott, Osborne, Caldwell, Heape, Weldon, Parker, Deighton and others) carried to a successful end. In each of these inquiries he himself took part, sometimes a large part, generally suggesting the problem to be solved, indicating the methods, and keeping a close watch over the whole progress of the study. Hence in many cases the published account bears his name as well as that of the pupil.

In the year 1878 his Monograph on Elasmobranch Fishes was published as a complete volume, and in the same year he received the honour of being elected a Fellow of the Royal Society, a distinction which now-a-days does not often fall to one so young. No sooner was the Monograph completed than in spite of the labours which his lectures entailed, he set himself to the great task of writing a complete treatise on Comparative Embryology. This not only laid upon him the heavy burden of gathering together the observations of others, enormous in number and continually increasing, scattered through many journals and books, and recorded in many different languages, as well as of putting them in orderly array, and of winnowing [Pg 15] out the grain from the chaff (though his critical spirit found some relief in the latter task), but also caused him much labour, inasmuch as at almost every turn new problems suggested themselves, and demanded inquiry before he could bring his mind to writing about them. This desire to see his way straight before him, pursued him from page to page, and while it has resulted in giving the book an almost priceless value, made the writing of it a work of vast labour. Many of the ideas thus originated served as the bases of inquiries worked out by himself or his pupils, and published in the form of separate papers, but still more perhaps never appeared either in the book or elsewhere and were carried with him undeveloped and unrecorded to the grave.

The preparation of this work occupied the best part of his time for the next three years, the first volume appearing in 1880, the second in 1881.

In the autumn of 1880, he attended the Meeting at Swansea of the British Association for the Advancement of Science, having been appointed Vice-President of the Biological Section with charge of the Department of Anatomy and Physiology. At the Meetings of the Association, especially of late years, much, perhaps too much, is expected in the direction of explaining the new results of science in a manner interesting to the unlearned. Popular expositions were never very congenial to Balfour, his mind was too much occupied with the anxiety of problems yet to be solved; he was therefore not wholly at his ease, in his position on this occasion. Yet his introductory address, though not of a nature to interest a large mixed audience, was a luminous, brief exposition of the modern development and aims of embryological investigation.

During these years of travail with the Comparative Embryology the amount of work which he got through was a marvel to his friends, for besides his lectures, and the researches, and the writing of the book, new labours were demanded of him by the University for which he was already doing so much. Men at Cambridge, and indeed elsewhere as well, soon began to find out that the same clear insight which was solving biological problems could be used to settle knotty [Pg 16] questions of policy and business. Moreover he united in a remarkable manner, the power of boldly and firmly asserting and maintaining his own views, with a frank courteousness which went far to disarm opponents. Accordingly he found himself before long a member of various Syndicates, and indeed a very great deal of his time was thus occupied, especially with the Museums and Library Syndicates, in both of which he took the liveliest interest. Besides these University duties his time and energy were also at the service of his College. In the preparation of the New Statutes, with which about this time the College was much occupied, the Junior Fellows of the College took a conspicuous share, and among these Junior Fellows Balfour was perhaps the most active; indeed he was their leader, and he threw himself into the investigation of the bearings and probable results of this and that proposed new statute with as much zeal as if he were attacking some morphological problem.

While he was in the midst of these various labours, his friends often feared for his strength, for though gradually improving in health after his first year at Cambridge, he was not robust, and from time to time he seemed on the point of breaking down. Still, hard as he was working, he was in reality wisely careful of himself, and as he grew older, paid more and more attention to his health, daily taking exercise in the form either of bicycle rides or of lawn-tennis. Moreover he continued to spend some part of his vacations in travel. Combining business with pleasure, he made frequent visits to Germany and France, and especially to Naples. The Christmas of 1876-7 he spent in Greece, that of 1878-9 at Ragusa, where his old school-fellow and friend Mr Arthur Evans was at that time residing, and the appointment of his friend Kleinenberg to a Professorship at Messina led to a journey there. Early in the long vacation of 1880, he went with his sister, Mrs H. Sidgwick, and her husband to Switzerland, and was joined there for a short time by his friend and pupil Adam Sedgwick. During this visit he took his first lessons in Alpine climbing, making several excursions, some of them difficult and dangerous; and the love of mountaineering laid so firm a hold upon him, that he returned to Switzerland later on in the autumn of the same year, in company with his [Pg 17] brother Gerald, and spent some weeks near Zermatt in systematic climbing, ascending, among other mountains, the Matterhorn and the Weisshorn. In the following summer, 1881, he and his brother Gerald again visited the Alps, dividing their time between the Chamonix district and the Bernese Oberland. On this occasion some of the excursions which they made were of extreme difficulty, and such as needed not only great presence of mind and bodily endurance, but also skilful and ready use of the limbs. As a climber indeed Balfour soon shewed himself fearless, indefatigable, and expert in all necessary movements as well as full of resources and expedients in the face of difficulties, so much so that he almost at once took rank among the foremost of distinguished mountaineers. In spite of his apparent clumsiness in some matters, he had even as a lad proved himself to be a bold and surefooted climber. Moreover he had been perhaps in a measure prepared for the difficulties of Alpine climbing by his experience in deer-stalking. This sport he had keenly and successfully pursued for many years at his brother's place in Rosshire. When however about the year 1877, the question of physiological experiments on animals became largely discussed in public, he felt that to continue the pursuit of this or any other sport involving, for the sake of mere pleasure, the pain and death of animals, was inconsistent with the position which he had warmly taken up, as an advocate of the right to experiment on animals; and he accordingly from that time onward wholly gave it up.

His fame as an investigator and teacher, and as a man of brilliant and powerful parts, was now being widely spread. Pupils came to him, not only from various parts of England, but from America, Australia and Japan. At the York Meeting of the British Association for the Advancement of Science, in August, 1881, he was chosen as one of the General Secretaries. In April, 1881, the honorary degree of LL.D. was conferred upon him by the University of Glasgow, and in November of the same year the Royal Society gave him one of the Royal Medals in recognition of his embryological discoveries, and at the same time placed him on its Council.

At Cambridge he was chosen, in the autumn of 1880, President of the Philosophical Society, and in the December of that [Pg 18] year a brilliant company were gathered together at the Annual Dinner to do honour to their new young President. Otherwise nothing as yet had been done for him in his own University in the way of recognition of his abilities and services; and he still remained a Lecturer of Trinity College, giving lectures in a University building. An effort had been made by some of his friends to urge the University to take some step in this direction; but it was thought at that time impossible to do anything. In 1881 a great loss fell upon the sister University of Oxford in the death of Prof. George Rolleston; and soon after very vigorous efforts were made to induce Balfour to become a candidate for the vacant chair. The prospect was in many ways a tempting one, and Balfour seeing no very clear way in the future for him at his own University, was at times inclined to offer himself, but eventually he decided to remain at Cambridge. Hardly had this temptation if we may so call it been overcome when a still greater one presented itself. Through the lamented death of Sir Wyville Thomson in the winter of 1881-2, the chair of Natural History at Edinburgh, perhaps the richest and most conspicuous biological chair in the United Kingdom, became vacant. The post was in many ways one which Balfour would have liked to hold. The teaching duties were it is true laborious, but they had in the past been compressed into a short time, occupying only the summer session and leaving the rest of the year free, and it seemed probable that this arrangement might be continued with him. The large emolument would also have been grateful to him inasmuch as he would have felt able to devote the whole of it to scientific ends; and the nearness to Whittinghame, his native place and brother's home, added to the attractions; but what tempted him most was the position which it would have given him, and the opportunities it would have afforded, with the rich marine Fauna of the north-eastern coast close at hand, to develop a large school of Animal Morphology. The existing Professors at Edinburgh were most desirous that he should join them, and made every effort to induce him to come. On the part of the Crown, in whose hands the appointment lay, not only were distinct offers made to him, but he was repeatedly pressed to accept the post. Nor was it until after a considerable [Pg 19] struggle that he finally refused, his love for his own University in the end overcoming the many inducements to leave; he elected to stay where he was, trusting to the future opening up for him some suitable position. In this decision he was undoubtedly influenced by the consideration that Cambridge, besides being the centre of his old friendships, had become as it were a second home for his own family. By the appointment of Lord Rayleigh to the chair of Experimental Physics his sister Lady Rayleigh had become a resident, his sister Mrs Sidgwick had lived there now for some years, and his brother Gerald generally spent the summer there; their presence made Cambridge doubly dear to him.

At the close of the Michaelmas term, with feelings of relief at having completed his Comparative Embryology, the preparation of the second volume of which had led to almost incessant labour during the preceding year, he started to spend the Christmas vacation with his friend Kleinenberg at Messina. Stopping at Naples on his way thither he found his pupil Caldwell, who had been sent to occupy the University table at the Stazione Zoologica, lying ill at Capri, with what proved to be typhoid fever. The patient was alone, without any friend to tend him, and his mother who had been sent for had not yet arrived. Accordingly Balfour (with the kindness all forgetful of himself which was his mark all his life through) stayed on his journey to nurse the sick man until the mother came. He then went on to Messina, and there seemed to be in good health, amusing himself with the ascent of Etna. Yet in January, soon after his return home, he complained of being unwell, and in due time distinct symptoms of typhoid fever made their appearance. The attack at first promised to be severe, but happily the crisis was soon safely passed and the convalescence was satisfactory.

While yet on his sick bed, a last attempt was made to induce him to accept the Edinburgh offer, and for the last time he refused. These repeated offers, and the fact that the dangers of his grave illness had led the University vividly to realize how much they would lose if Balfour were taken away from them, encouraged his friends to make a renewed effort to gain for him some adequate position in the University. This time [Pg 20] the attempt was successful, and the authorities took a step, unusual but approved of by the whole body of resident members of the University; they instituted a new Professorship of Animal Morphology, to be held by Balfour during his life or as long as he should desire, but to terminate at his death or resignation unless it should be otherwise desirable. Accordingly in May, 1882, he was admitted into the Professoriate as Professor of Animal Morphology.

During his illness his lectures had been carried on by his Demonstrator, Mr Adam Sedgwick, who continued to take his place during the remainder of that Lent Term and during the ensuing Easter Term. The spring Balfour spent partly in the Channel Islands with his sister Alice, partly in London with his eldest brother, but in the course of the Easter Term returned to Cambridge and resumed his work though not his lectures. His recovery to health was steady and satisfactory, the only drawback being a swelling over the shin-bone of one leg, due to a blow on the rocks at Sark; otherwise he was rapidly becoming strong. He himself felt convinced that a visit to the Alps, with some mountaineering of not too difficult a kind, would complete his restoration to health. In this view many of his friends coincided; for the experience of former years had shewn them what a wonderfully beneficial effect the Alpine air and exercise had upon his health. He used to go away pale, thin and haggard, to return bronzed, clear, firm and almost stout; nor was there anything in his condition which seemed to forbid his climbing, provided that he was cautious at the outset. Accordingly, early in June he left Cambridge for Switzerland, having long ago, during his illness in fact, engaged his old guide, Johann Petrus, whom he had first met in 1880, and who had always accompanied him in his expeditions since.

His first walking was in the Chamonix district; and here he very soon found his strength and elasticity come back to him. Crossing over from Montanvert to Courmayeur, by the Col du Géant, he was attracted by the peak called the Aiguille Blanche de Peuteret, a virgin peak, the ascent of which had been before attempted but not accomplished. Consulting with Petrus he determined to try it, feeling that the fortnight, which by this [Pg 21] time he had spent in climbing, had brought back to him his old vigour, and that his illness was already a thing of the past.

There is no reason to believe that he regarded the expedition as one of unusual peril; and an incident which at the time of his death was thought by some to indicate this was in reality nothing more than a proof of his kindly foresight. The guide Petrus was burdened by a debt on his land amounting to about £150. In the previous year Balfour and his brother had come to know of this debt; and, seeing that no Alpine ascent is free from danger, that on any expedition some accident might carry them off, had conceived the idea of making some provision for Petrus' family in case he might meet with sudden death in their service. This suggestion of the previous year Balfour carried out on this occasion, and sent home to his brother Gerald a cheque of £150 for this purpose. But the cheque was sent from Montanvert before he had even conceived the idea of ascending the Aiguille Blanche. It was not a provision for any specially dangerous ascent, and must be regarded as a measure prompted not by a sense of coming peril but rather by the donor's generous care for his servant.

On Tuesday afternoon, July 18, he and Petrus, with a porter to carry provisions and firing to their sleeping-place on the rocks, set out from Courmayeur, the porter returning the same night. They expected to get back to Courmayeur some time on the Thursday, but the day passed without their appearing. This did not cause any great anxiety because it was supposed that they might have found it more convenient to pass over to the Chamonix side than to return to Courmayeur. When on Friday however telegrams dispatched to Chamonix and Montanvert brought answers that nothing had been seen of them, it became evident that some accident had happened, and an exploring party set out for the hills. It was not until early on the Sunday morning that this search party found the bodies, both partly covered with snow, lying on the Glacier de Fresney, below the impassable icefall which separates the upper basin of the glacier from the lower portion, and at the foot of a couloir which descends by the side of the icefall. Their tracks were visible on the snow at the top of the couloir. Balfour's neck was broken, and his skull fractured [Pg 22] in three places; Petrus' body was also fractured in many places. The exact manner of their death will never be known, but there can be no doubt that, in Balfour's case at all events, it was instantaneous, and those competent to form a judgment are of opinion that they were killed by a sudden fall through a comparatively small height, slipping on the rocks as they were descending by the side of the ice-fall, and not precipitated from the top of the couloir. There is moreover indirect evidence which renders it probable that in the fatal fall Petrus slipped first and carried Balfour with him. Whether they had reached the summit of the Aiguille and were returning home after a successful ascent or whether they were making their way back disheartened and wearied with failure, is not and perhaps never will be known. Since the provisions at the sleeping-place were untouched, the deaths probably took place on Wednesday the 19th. The bringing down the bodies proved to be a task of extreme difficulty, and it was not till Wednesday the 26th that the remains reached Courmayeur, where M. Bertolini, the master of the hotel, and indeed everyone, not least the officers of a small body of Italian troops stationed there, shewed the greatest kindness and sympathy to Balfour's brothers, Gerald and Eustace, who hastened to the spot as soon as the news of the terrible disaster was telegraphed home. Mr Walter Leaf also and Mr Conway, friends of Balfour, the former a very old one, who had made their way to Courmayeur from other parts of Switzerland as soon as they heard of the accident, rendered great assistance. The body was embalmed, brought to England, and buried at Whittinghame on Saturday, Aug. 5, the Fellows of Trinity College holding a service in the College Chapel at the same time.

In person he was tall, being fully six feet in height, well built though somewhat spare. A broad forehead overhanging deeply set dark brown eyes whose light shining from beneath strongly marked eye-brows told all the changes of his moods, slightly prominent cheek-bones, a pale skin, at times inclined to be even sallow, dark brown hair, allowed to grow on the face only as a small moustache, and slight whiskers, made up a countenance which bespoke at once strength of character and delicacy of constitution. It was an open countenance, hiding [Pg 23] nothing, giving sign at once, both when his body was weary or weak, and when his mind was gladdened, angered or annoyed.

The record of some of his thoughts and work, all that he had given to the world will be found in the following pages. But who can tell the ideas which had passed into his quick brain, but which as yet were known only to himself, of which he had given no sign up to that sad day on which he made the fatal climb? And who can say whither he might not have reached had he lived, and his bright young life ripened as years went on? This is not the place to attempt any judgment of his work: that may be left to other times, and to other hands; but it may be fitting to place here on record a letter which shews how much the greatest naturalist of this age appreciated his younger brother. Among Balfour's papers was found a letter from Charles Darwin, acknowledging the receipt of Vol. II. of the Comparative Embryology in the following words:

"July 6, 1881.

Down, Beckenham, Kent.

My Dear Balfour,

I thank you heartily for the present of your grand book, and I congratulate you on its completion. Although I read almost all of Vol. I, I do not feel that I am worthy of your present, unless indeed the fullest conviction that it is a memorable work makes me worthy to receive it.

* * * * *

Once again accept my thanks, for I am proud to receive a book from you, who, I know, will some day be the chief of the English Biologists.

Believe me,

Yours sincerely,

Charles Darwin."

The loss of him was a manifold loss. He is mourned, and will long be mourned, for many reasons. Some miss only the brilliant investigator; others feel that their powerful and sympathetic teacher is gone; some look back on his memory [Pg 24] and grieve for the charming companion whose kindly courtesy and bright wit made the hours fly swiftly and pleasantly along; and to yet others is left an aching void when they remember that they can never again lean on the friend whose judgment seemed never to fail and whose warm-hearted affection was a constant help. And to some he was all of these. At the news of his death the same lines came to the lips of all of us, so fittingly did Milton's words seem to speak our loss and grief—

"For Lycidas is dead, dead ere his prime,
  Young Lycidas, and hath not left his peer."

M. FOSTER.

[2] His first Demonstrator up to Christmas 1877, was Mr J. F. Bullar. In Jan. 1878, Mr Adam Sedgwick took the post of Senior Demonstrator, and held it until Balfour's death.

[Pg 25]

I. On some Points in the Geology of the East Lothian Coast[3].

By G. W. and F. M. Balfour, Trinity College, Cambridge.

The interesting relation between the Porphyrite of Whitberry Point, at the mouth of the Tyne, near Dunbar, and the adjacent sedimentary rocks, was first noticed, we believe, by Professor Geikie, who speaks of it in the Memoirs of the Geological Survey of East Lothian, pages 40 and 31, and again in the new edition of Jukes's Geology, p. 269. The volcanic mass which forms the point consists of a dark felspathic base with numerous crystals of augite: it is circular in form, and is exposed for two-thirds of its circumference in a vertical precipice facing the sea, about twenty feet in height.

The rock is traversed by numerous joints running both in a horizontal and in a vertical direction. The latter are by far the most conspicuous, and give the face of the cliff, when seen from a distance, a well-marked columnar appearance, though the columns themselves are not very distinct or regular. They are quadrangular in form, and are evidently produced by the intersection at right-angles of the two series of vertical joints.

It is clear that the face of the precipice has been gradually receding in proportion as it yielded to the action of the waves; and that at a former period the volcanic rock extended considerably further than at present over the beds which are seen to dip beneath it. These latter consist of hard fine-grained calcareous sandstones belonging to the Lower Carboniferous formation. Their colour varies from red to white, and their prevailing dip is in a N.W. direction, with an average inclination of 12-20°. If the volcanic mass is a true intrusive rock, we should naturally expect the strata which surround it to dip away in all directions, the amount of their inclination diminishing in [Pg 26] proportion to their distance from it. We find, however, that the case is precisely the reverse: as the beds approach the base of the cliff, they dip towards it from every side at perpetually increasing angles, until at the point of junction the inclination amounts in places to as much as 55 degrees. The exact amount of dip in the various positions will be seen on referring to the accompanying map.

Fig. 1. Map of Strata at Whitberry Point. Scale, 6 in. to the mile.

A. Lava sheet. B. Sandstone Beds, dipping from every side towards the lava. CC. Line of Section along which Fig. 2 is supposed to be drawn.

We conceive that the phenomenon is to be explained by supposing the orifice through which the lava rose and overflowed the surface of the sedimentary strata to have been very much smaller in area than the extent of igneous rock at present visible; and that the pressure of the erupted mass on the soft beds beneath, aided perhaps by the abstraction of matter from below, caused them to incline towards the central point at a gradually increasing angle. The diagram, fig. 2, will serve further to illustrate this hypothesis. A is the neck or orifice by which the melted matter is supposed to ascend. C shews the sheet of lava after it has overspread the surface of the sandstone beds B, so as to cause them to assume their present inclination. The dotted [Pg 27] lines represent the hypothetical extension of the igneous mass and sandstones previous to the denudation which they have suffered from the action of the waves.

Professor Geikie, in his admirable treatise on the Geology of the county[4], adopts a view on this subject which is somewhat different from that which is suggested in this paper. He considers that the whole mass is an intrusive neck of rock with perpendicular sides; and that it once filled up an orifice through the surrounding sedimentary strata, of which it is now the only remnant.

Fig. 2. Vertical Section through CC. Diagram (Fig. 1).

A. Orifice by which the lava ascended. B. Sandstone Beds. B´. Hypothetical extension of ditto. C. Sheet of lava spread over the sandstones B. C´. Hypothetical extension of ditto.

He admits that the inclination of the sandstone beds towards the igneous mass in the centre is a phenomenon that is somewhat difficult to explain, and suggests that a subsequent contraction of the column may have tended to produce such a result. To use his own words: In the case of a solid column of felstone or basalt, the contraction of the melted mass on cooling may have had some effect in dragging down the sides of the orifice[5].

But, apart from other objections, it is scarcely conceivable that this result should have been produced by the contraction of the column.

In his recent edition of Jukes's Manual of Geology (p. 269), in which he also refers to this instance, he states that in other cases of necks it is found to be an almost invariable rule, that [Pg 28] strata are bent down so as to dip into the neck all round its margin. We are not aware to what other instances Prof. Geikie may allude; but on referring to his Memoir on the Geology of East Lothian, we find that he states in the cases of 'North Berwick Law' and 'Traprain' (which he compares with the igneous mass at Whitberry Point), that the beds at the base of these two necks, where exposed, dip away from them, and that at a high angle.

In support of the hypothesis which we have put forward, the following arguments may be urged:

(1) That in one place at least the sedimentary strata are seen to be actually dipping beneath the superincumbent basalt; and that the impression produced by the general relation of the two rocks is, that they do so everywhere.

(2) Since the columns into which the lava is split are vertical, the cooling surface must have been horizontal: the mass must, therefore, have formed a sheet, and not a dyke; for, in the latter case, the cooling surfaces would have been vertical.

(3) It is difficult to conceive, on the supposition that the volcanic rock is a neck with perpendicular sides, that the marine denudation should have uniformly proceeded only so far as to lay bare the junction between the two formations. We should have expected that in many places the igneous rock itself would have been cut down to the general level, whereas the only signs of such an effect are shown in a few narrow inlets where the rock was manifestly softer than in the surrounding parts.

The last objection is greatly confirmed by the overhanging cliffs and numerous blocks of porphyrite which lie scattered on the beach, as if to attest the former extension of that ancient sheet of which these blocks now form but a small remnant. Indeed, the existence of such remains appears sufficient of itself to condemn any hypothesis which presumes the present face of the cliff to have formed the original boundary of the mass.

It may be fairly objected to our theory, as Prof. Geikie himself has suggested, that the high angle at which the strata dip is difficult to account for. But, in fact, this steep inclination constitutes the very difficulty which any hypothesis on the subject must be framed to explain; and it is a difficulty which is not more easily solved by Prof. Geikie's theory than by our own.

[3] From the Geological Magazine, Vol. IX. No. 4. April, 1872.

[4] Memoirs of Geological Survey of Scotland, sheet 33, pp. 40, 41.

[5] Note on p. 41 of Mem. Geol. Survey of East Lothian.

[Pg 29]

II. The Development and Growth of the Layers of the Blastoderm[6].

With Plate 1, figs. 1-5 and 9-12.

The following paper deals with the changes which take place in the cells of the blastoderm of the hen's egg during the first thirty or forty hours of incubation. The subject is one which has, as a general rule, not been much followed up by embryologists, but is nevertheless of the greatest interest, both in reference to embryology itself, and to the growth and changes of protoplasm exhibited in simple embryonic cells. I am far from having exhausted the subject in this paper, and in some cases I shall be able merely to state facts, without being able to give any explanation of their meaning.

My method of investigation has been the examination of sections and surface views. For hardening the blastoderm I have employed, as usual, chromic acid, and also gold chloride. It is, however, difficult to make sections of blastoderms hardened by this latter reagent, and the sections when made are not in all cases satisfactory. For surface views I have chiefly used silver nitrate, which brings out the outlines of the cells in a manner which leaves nothing to be desired as to clearness. If the outlines only of the cells are to be examined, a very short immersion (half a minute) of the blastoderm in a half per cent. solution of silver nitrate is sufficient, but if the immersion lasts for a longer period the nuclei will be brought out also. For studying the latter, however, I have found it better to employ gold chloride or carmine in conjunction with the silver nitrate.

My observations begin with the blastoderm of a freshly laid egg. The appearances presented by sections of this have been accurately described by Peremeschko, Ueber die Bildung der [Pg 30] Keimblätter im Hühnerei, Sitzungsberichte der K. Akademie der Wissenschaften in Wien, 1868. Oellacher, Untersuchung über die Furchung und Blatterbildung im Hühnerei, Studien aus dem Institut für Experim. Pathologie in Wien, 1870 (pp. 54-74), and Dr Klein, lxiii. Bande der Sitz. der K. Acadamie der Wiss. in Wien, 1871.

The unincubated blastoderm (Pl. 1, fig. 1) consists of two layers. The upper layer is composed of a single row of columnar cells. Occasionally, however, the layer may be two cells thick. The cells are filled with highly refracting spherules of a very small size, and similar in appearance to the finest white yolk spherules, and each cell also contains a distinct oval nucleus. This membrane rests with its extreme edge on the white yolk, its central portion covering in the segmentation cavity. From the very first it is a distinct coherent membrane, and exhibits with silver nitrate a beautiful hexagonal mosaic of the outlines (Pl. 1, fig. 6) of the cells. The diameter of the cells when viewed from above is from 1/2000 - 1/3000 of an inch. The under layer is very different from this: it is composed of cells which are slightly, if at all, united, and which vary in size and appearance, and in which a nucleus can rarely be seen. The cells of which it is composed fill up irregularly the segmentation cavity, though a distinct space is even at this time occasionally to be found at the bottom of it. Later, when the blastoderm has spread and the white yolk floor has been used as food, a considerable space filled with fluid may generally be found.

The shape of the floor of the cavity varies considerably, but it is usually raised in the middle and depressed near the circumference. In this case the under layer is perhaps only two cells deep at the centre and three or four cells deep near the circumference.

The cells of which this layer is composed vary a good deal in size; the larger cells being, however, more numerous in the lower layers. In addition, there are usually a few very large cells quite at the bottom of the cavity, occasionally separated from the other cells by fluid. They were called formative cells (Bildungselemente) by Peremeschko (loc. cit.); and, according to Oellacher's observations (loc. cit.), some of them, at any rate, fall to the bottom of the segmentation cavity during the later [Pg 31] stages of segmentation. They do not differ from the general lower layer cells except in size, and even pass into them by insensible gradations. All the cells of the lower layer are granular, and are filled with highly refracting spherules precisely similar to the smaller white yolk spherules which line the bottom of the segmentation cavity.

The size of the ordinary cells of the lower layer varies from 1/2000 - 1/1000 of an inch. The largest of the formative cells come up to 1/300 of an inch. It will be seen from this description that, morphologically speaking, we cannot attach much importance to the formative cells. The fact that they broke off from the blastoderm, towards the end of the segmentation—even if we accept it as a normal occurrence, rather than the result of manipulation—is not of much importance, and, except in size, it is impossible to distinguish these cells from other cells of the lower layer of the blastoderm.

Physiologically, however, as will be afterwards shewn, they are of considerable importance.

The changes which the blastoderm undergoes during the first three or four hours of incubation are not very noticeable. At about the sixth or eighth hour, or in some cases considerably earlier, changes begin to take place very rapidly. These changes result in the formation of a hypoblast and mesoblast, the upper layer of cells remaining comparatively unaltered as the epiblast.

To form the hypoblast a certain number of the cells of the lower layer begin to undergo remarkable changes. From being spherical and, as far as can be seen, non-nucleated, they become (vide fig. 2, h) flattened and nucleated, still remaining granular, but with fewer spherules.

Here, then, is a direct change, of which all the stages can be followed, of a cell of one kind into a cell of a totally different character. The new cell is not formed by a destruction of the old one, but directly from it by a process of metamorphosis. These hypoblast cells are formed first at the centre and later at the circumference, so that from the first the cells at the circumference are less flattened and more granular than the cells at the centre. A number of cells of the original lower layer are enclosed between this layer and the epiblast; and, [Pg 32] in addition to these, the formative cells (as has been shewn by Peremeschko, Oellacher, and Klein, whose observations I can confirm) begin to travel towards the circumference, and to pass in between the epiblast and hypoblast.

Both the formative cells, and the lower layer cells enclosed between the hypoblast and epiblast, contribute towards the mesoblast, but the mode in which the mesoblast is formed is very different from that in which the hypoblast originates.

It is in this difference of formation that the true distinction between the mesoblast and hypoblast is to be looked for, rather than in the original difference of the cells from which they are derived.

The cells of the mesoblast are formed by a process which seems to be a kind of free cell formation. The whole of the interior of each of the formative cells, and of the other cells which are enclosed between the epiblast and the hypoblast, become converted into new cells. These are the cells of the mesoblast. I have not been able perfectly to satisfy myself as to the exact manner in which this takes place, but I am inclined to think that some or all of the spherules which are contained in the original cells develop into nuclei for the new cells, the protoplasm of the new cells being formed from that of the original cells.

The stages of formation of the mesoblast cells are shewn in the section (Pl. 1, fig. 2), taken from the periphery of a blastoderm of eight hours.

The first formation of the mesoblast cells takes place in the centre of the blastoderm, and the mass of cells so formed produces the opaque line known as the primitive streak. This is shown in Pl. 1, fig. 9.

One statement I have made in the above description in reference to the origin of the mesoblast cells, viz. that they are only partly derived from the formative cells at the bottom of the segmentation cavity, is to a certain extent opposed to the statements of the three investigators above mentioned. They state that the mesoblast is entirely derived from the formative cells. It is not a point to which I attach much importance, considering that I can detect no difference between these cells and any other cells of the original lower layer except that of size; and even this difference is probably to be explained [Pg 33] by their proximity to the white yolk, whose spherules they absorb. But my reason for thinking it probable that these cells alone do not form the mesoblast are: 1st. That the mesoblast and hypoblast are formed nearly synchronously, and except at the centre a fairly even sprinkling of lower layer cells is from the first to be distinguished between the epiblast and hypoblast. 2nd. That if some of the lower layer cells are not converted into mesoblast, it is difficult to see what becomes of them, since they appear to be too numerous to be converted into the hypoblast alone. 3rd. That the chief formation of mesoblast at first takes place in the centre, while if the formative cells alone took part in its formation, it would be natural to expect that it would begin to be formed at the periphery.

Oellacher himself has shewn (Zeitschrift für wissenschaftliche Zoologie, 1873, Beiträge zur Entwick. Gesch. der Knochenfische) that in osseous fishes the cells which break away from the blastoderm take no share in the formation of the mesoblast, so that we can derive no argument from the formation of the mesoblast in these animals, for believing that in the chick it is derived only from the formative cells.

In the later stages, however, from the twelfth to the twenty-fifth hour, the growth of the mesoblast depends almost entirely on these cells, and Peremeschko's discovery of the fact is of great value.

Waldeyer (Henle und v. Pfeufer's Zeitschrift, xxxiv. Band, für 1869) has given a different account of the origin of the layers. There is no doubt, however, in opposition to his statements and drawings, that from the very first the hypoblast is distinct from the mesoblast, which is, indeed, most conspicuously shewn in good sections; and his drawings of the derivation of the mesoblast from the epiblast are not very correct.

The changes which have been described are also clearly shewn by means of silver nitrate. Whereas, at first this reagent brought out no outline markings of cells in the lower layer, by the eighth to the twelfth hour the markings (Pl. 1, fig. 3) are very plain, and shew that the hypoblast is a distinct coherent membrane.

In section, the cells of the hypoblast appear generally very thin and spindle shaped, but the outlines brought out by the [Pg 34] silver nitrate shew that they are much expanded horizontally, but very irregular as to size, varying even within a small area from 1/4000 - 1/400 of an inch in the longest diameter.

At about the twelfth hour they are uniformly smaller a short way from each extremity of its longer axis than over the rest of the blastoderm.

It is, perhaps, fair to conclude from this that growth is most rapid at these parts.

At this time the hypoblast, both in sections and from a surface view after treatment with silver nitrate, appears to end abruptly against the white yolk. The surface view also shews that its cells are still filled with highly refractive globules, making it difficult to see the nucleus. In some cases I thought that I could (fig. 3, a) make out that it was hour-glass shaped, and some cells certainly contain two nuclei. Some of the cells (fig. 3, b) shew re-entrant curves, which prove that they have undergone division.

The cells of the epiblast, up to the thirteenth hour, have chiefly undergone change in becoming smaller.

In surface views they are about 1/4000 of an inch in diameter over the centre of the pellucid area, and increase to 1/2000 of an inch over the opaque area.

In the centre of the pellucid area the form of the epiblast cells is more elongated vertically and over the opaque area more flattened than was the case with the original upper layer cells. In the centre the epiblast is two or three cells deep.

Before going on to the further changes of the blastodermic cells it will be well to say a few words in reference to the origin of the mesoblast.

From the description given above it will be clear that in the chick the mesoblast has an independent origin; it can be said neither to originate from the epiblast nor from the hypoblast. It is formed coincidently with the latter out of apparently similar segmentation cells. The hypoblast, as has been long known, shews in the chick no trace of its primitive method of formation by involution, neither does the mesoblast shew any signs of its primitive mode of formation. In so excessively highly differentiated a type as birds we could hardly expect to find, and certainly do not find, any traces of the [Pg 35] primitive origin of the mesoblast, either from the epiblast or hypoblast, or from both. In the chick the mesoblast cells are formed directly from the ultimate products of segmentation. From having a secondary origin in most invertebrates the mesoblast comes to have, in the chick, a primary origin from the segmentation spheres, precisely as we find to be the case with the nervous layer in osseous fishes. It is true we cannot tell which segmentation-cells will form the mesoblast, and which the hypoblast; but the mesoblast and hypoblast are formed at the same time, and both of them directly from segmentation spheres.

The process of formation of the mesoblast in Loligo, as observed by Mr Ray Lankester (Annals and Magazine of Natural History, February, 1873), is still more modified. Here the mesoblast arises independently of the blastoderm, and by a process of free cell-formation in the yolk round the edge of the blastoderm. If Oellacher's observations in reference to the origin of formative cells are correct, then the modes of origin of the mesoblast in Loligo and the chick would have nothing in common; but if the formative cells are in reality derived from the white yolk, and also are alone concerned in the formation of the mesoblast, then the modes of formation of the mesoblast in the chick would be substantially the same as that observed by Mr Ray Lankester in Loligo.

No very important changes take place in the actual forms of the cells during the next few hours. A kind of fusion takes place between the epiblast and the mesoblast along the line of the primitive streak forming the axis-string of His; but the line of junction between the layers is almost always more or less visible in sections. In any case it does not appear that there is any derivation of mesoblast cells from the epiblast; and since the fusion only takes place in the region of the primitive groove, and not in front, where the medullary groove arises (see succeeding paper), it cannot be considered of any importance in reference to the possible origin of the Wolffian duct, &c., from the epiblast (as mooted by Waldeyer, Eierstock und Ei, Leipzig, 1870). The primitive groove, as can be seen in sections, begins to appear very early, generally before the twelfth hour. The epiblast spreads rapidly over the white yolk, and the area pellucida also increases in size.

[Pg 36] From the mesoblast forming at first only a small mass of cells, which lies below the primitive streak, it soon comes to be the most important layer of the blastoderm. Its growth is effected by means of the formative cells. These cells are generally not very numerous in an unincubated blastoderm, but rapidly increase in numbers, probably by division; at the same time they travel round the edge of, and in some cases through, the hypoblast, and then become converted in the manner described into mesoblast cells. They act as carriers of food from the white yolk to the mesoblast till, after the formation of the vascular area, they are no longer necessary. The numerous cases in which two nucleoli and even two nuclei can be seen in one cell prove that the mesoblast cells also increase by division.

The growth of the hypoblast takes place in a very different way. It occurs by a direct conversion, cell for cell, of the white yolk spheres into hypoblast cells. This interpretation of the appearances, which I will describe presently, was first suggested to me by Dr Foster, from an examination of some of my specimens of about thirty-six hours, prepared with silver nitrate. Where there is no folding at the junction between the pellucid and opaque areas, there seems to be a perfect continuity in the silver markings and a gradual transition in the cells, from what would be undoubtedly called white yolk spheres, to as undoubted hypoblast cells (vide Pl. 1, fig. 5). In passing from the opaque to the pellucid areas the number of white yolk spherules in each cell becomes less, but it is not till some way into the pellucid area that they quite cease to be present. I at first thought that this was merely due to the hypoblast cells feeding on the white yolk sphericles, but the perfect continuity of the cells, and the perfect gradation in passing from the white yolk cells to the hypoblast, proves that the other interpretation is the correct one, viz. that the white yolk spheres become directly converted into the hypoblast cells. This is well shewn in sections (vide Pl. 1, fig. 4) taken from embryos of all ages from the fifteenth to the thirty-sixth hour and onwards. But it is, perhaps, most easily seen in embryos of about twenty hours. In such an embryo there is a most perfect gradation: the cells of the hypoblast become, as they approach the edge [Pg 37] of the pellucid area, broader, and are more and more filled with white yolk sphericles, till at the line of junction it is quite impossible to say whether a particular cell is a white-yolk cell (sphere) or a hypoblast cell. The white-yolk cells near the line of junction can frequently be seen to possess nuclei. At first the hypoblast appears to end abruptly against the white yolk; this state of things, however, soon ends, and there supervenes a complete and unbroken continuity between the hypoblast and the white yolk.

Of the mode of increase of the epiblast I have but little to say. The cells undoubtedly increase entirely by division, and the new material is most probably derived directly from the white yolk.

Up to the sixth hour the cells of the upper layer retain their early regular hexagonal pattern, but by the twelfth hour they have generally entirely lost this, and are irregularly shaped and very angular. The cells over the centre of the pellucid area remain the smallest up to the twenty-fifth hour or later, while those over the rest of the pellucid area are uniformly larger.

In the hypoblast the cells under the primitive groove, and on each side as far as the fold which marks off the exterior limit of the protovertebræ are at the eighteenth hour considerably smaller than any other cells of this layer.

In all the embryos between the eighteenth and twenty-third hour which I have examined for the purpose, I have found that at about two-thirds of the distance from the anterior end of the pellucid area, and just external to the side fold, there is a small space on each side in which the cells are considerably larger than anywhere else in the hypoblast. These larger cells, moreover, contain a greater number of highly refractive spherules than any other cells. It is not easy to understand why growth should have been less rapid here than elsewhere, as the position does not seem to correspond to any feature in the embryo. In some specimens the hypoblast cells at the extreme edge of the pellucid area are smaller than the cells immediately internal to them. At about the twenty-third hour these cells begin rapidly to lose the refractive spherules they contained in the earlier stages of incubation, and come [Pg 38] to consist of a nucleus surrounded simply by granular protoplasm.

At about this period of incubation the formative cells are especially numerous at the periphery of the blastoderm, and, no doubt, become converted into the mass of mesoblast which is found at about the twenty-fifth hour in the region of the vascular area. Some of them are lobate, and appear as if they were undergoing division. At this time also the greatest number of formative cells are to be found at the bottom of the now large segmentation cavity.

In embryos of from thirty to forty hours the cells of the hypoblast have, over the central portion of the pellucid area, entirely lost their highly refractive spherules, and in the fresh state are composed of the most transparent protoplasm. When treated with reagents they are found to contain an oval nucleus with one or sometimes two nucleoli, imbedded in a considerable mass of protoplasm. The protoplasm appears slightly granular and generally contains one or two small vacuoles. I have already spoken of the gradation of the hypoblast at the edge of the blastoderm into white yolk. I have, therefore, only to mention the variations in the size of its cells in different parts of the pellucid area. The points where the cells are smallest seem generally to coincide with the points of maximum growth. Over the embryo the cells are more regular than elsewhere. They are elongated and arranged transversely to the long axis of the embryo. They are somewhat hexagonal in shape, and not unlike the longer pieces in the dental plate of a Myliobatis (Pl. 1, fig. 10). This regularity, however, is much more marked in some specimens than in others. These cells are about 1/4000th of an inch in breadth, and 1/1000th in length. On each side of the embryo immediately external to the protovertebræ the cells are frequently about the same size as those over the embryo itself. In the neck, however, and near the end of the sinus rhomboidalis, they are considerably smaller, about 1/4000th inch each way. The reason of this small size is not very clear, but probably shews that the greatest growth is taking place at these two points. The cells, again, are very small at the head fold, but are very much larger in front of this—larger, in fact, than any other cells of the hypoblast. Outside the embryo they gradually increase [Pg 39] in size towards the edge of the pellucid area. Here they are about 1/1000th of an inch in diameter, irregular in shape and rather angular.

The outlines of the cells of the epiblast at this time are easily distinguished from the cells of the hypoblast by being more elongated and angular; they are further distinguished by the presence of numerous small oval cells, frequently at the meeting point of several cells, at other times at points along the lines of junction of two cells (Pl. 1, fig. 12). These small cells look very like the smaller stomata of endothelial membranes, but are shewn to be cells by possessing a nucleus. There is considerable variation in size in the cells in different parts of the epiblast. Between the front lobes of the brain the cells are very small, 1/4000th inch, rising to 1/2000th on each side. They are about the latter size over the greater part of the embryo. But over the sinus rhomboidalis they fall again to from 1/3000th to 1/4000th inch. This is probably to be explained by the growth of the medullary fold at this point, which pushes back the primitive groove. At the sides of the head the cells are larger than anywhere else in the epiblast, being here about 1/1000th inch in diameter. I at present see no explanation of this fact. At the periphery of the pellucid area and over the vascular area the cells are 1/1500th to 1/2000th inch in diameter, but at the periphery of the opaque area they are smaller again, being about the 1/3000th of an inch. This smaller size at the periphery of the area opaca is remarkable, since in the earlier stages the most peripheral epiblast cells were the largest. It, perhaps, implies that more rapid growth is at this time taking place in that part of the epiblast which is spreading over the yolk sac.

[Pg 40] EXPLANATION OF PLATE 1, Figs. 1-5 and 9-12.

Fig. 1. Section through an unincubated blastoderm, shewing the upper layer, composed of a single row of columnar cells, and the lower layer, composed of several rows of rounded cells in which no nucleus is visible. Some of the formative cells, at the bottom of the segmentation cavity, are seen at (b).

Fig. 2. Section through the periphery of an eight hours' blastoderm, shewing the epiblast (p), the hypoblast (h), and the mesoblast commencing to be formed (c), partly by lower-layer cells enclosed between the epiblast and hypoblast, and partly by formative cells. Formative cells at the bottom of the segmentation cavity are seen at b. At s is one of the side folds parallel to the primitive groove.

Fig. 3. Portion of the hypoblast of a thirteen hours' blastoderm, treated with silver nitrate, shewing the great variation in the size of the cells at this period. An hour-glass shaped nucleus is seen at a.

Fig. 4. Periphery of a twenty-three hours' blastoderm, shewing cell for cell the junction between the hypoblast (h) and white-yolk spheres (w).

Fig. 5. Junction between the white-yolk spheres and the hypoblast cells at the passage from the area pellucida to the area opaca. The specimen was treated with silver nitrate to bring out the shape of the cells. The line of junction between the opaque and pellucid areas passes diagonally.

Fig. 9. Section through the primitive streak of an eight hours' blastoderm. The specimen shews the mesoblast very much thickened in the immediate neighbourhood of the primitive streak, but hardly formed at all on each side of the streak. It also shews the primitive groove just beginning to be formed (pr), and the fusion between the epiblast and the mesoblast under the primitive groove. The hypoblast is completely formed in the central part of the blastoderm. At f is seen one of the side folds parallel to the primitive groove. Its depth has been increased by the action of the chromic acid.

Fig. 10. Hypoblast cells from the hinder end of a thirty-six hours' embryo, treated with silver nitrate, shewing the regularity and elongated shape of the cells over the embryo and the smaller cells on each side.

Fig. 11. Epiblast cells from an unincubated blastoderm, treated with silver nitrate, shewing the regular hexagonal shape of the cells and the small spherules they contain.

Fig. 12. Portion of the epiblast of a thirty-six hours' embryo, treated with silver nitrate, shewing the small rounded cells frequently found at the meeting-points of several larger cells which are characteristic of the upper layer.

[6] From the Quarterly Journal of Microscopical Science, Vol. XIII., 1873.

[Pg 41]

III. On the Disappearance of the Primitive Groove in the Embryo Chick[7].

With Plate 1, Figs. 6-8 and 13-19.

The investigations of Dursy (Der Primitivstreif des Hühnchens, von Dr E. Dursy. Lahr, 1866) on the primitive groove, shewing that it is a temporary structure, and not connected with the development of the neural canal, have in this country either been ignored or rejected. They are, nevertheless, perfectly accurate; and had Dursy made use of sections to support his statements I do not think they would so long have been denied. In Germany, it is true, Waldeyer has accepted them with a few modifications, but I have never seen them even alluded to in any English work. The observations which I have made corroborating Dr Dursy may, perhaps, under these circumstances be worth recording.

After about twelve hours of incubation the pellucid area of a hen's egg has become somewhat oval, with its longer axis at right angles to the long axis of the egg. Rather towards the hinder (narrower) end of this an opaque streak has appeared, with a somewhat lighter line in the centre. A section made at the time shews that the opaque streak is due partly to a thickening of the epiblast, but more especially to a large collection of the rounded mesoblast cells, which along this opaque line form a thick mass between the epiblast and the hypoblast. The mesoblast cells are in contact with both hypoblast and epiblast, and appear to be fused with the latter. The line of junction between them can, however, almost always be made out.

Soon after the formation of this primitive streak a groove is formed along its central line by a pushing inwards of the epiblast. [Pg 42] The epiblast is not thinner where it lines the groove, but the mass of mesoblast below the groove is considerably thinner than at its two sides. This it is which produces the peculiar appearance of the primitive groove when the blastoderm is viewed by transmitted light as a transparent line in the middle of an opaque one.

This groove, as I said above, is placed at right angles to the long axis of the egg, and nearer the hind end, that is, the narrower end of the pellucid area. It was called the primitive groove by the early embryologists, and they supposed that the neural canal arose from the closure of its edges above. It is always easy to distinguish this groove, in transverse sections, by several well-marked characters. In the first place, the epiblast and mesoblast always appear more or less fused together underneath it; in the second place, the epiblast does not become thinner where it lines the groove; and in the third place, the mesoblast beneath it never shews any signs of being differentiated into any organ.

As Dursy has pointed out, there is frequently to be seen in fresh specimens, examined as transparent objects, a narrow opaque line running down the centre of this groove. I do not know what this line is caused by, as there does not appear to be any structural feature visible in sections to which it can correspond.

From the twelfth to the sixteenth hour the primitive groove grows rapidly, and by the sixteenth hour is both absolutely and considerably longer than it was at the twelfth hour, and also proportionately longer as compared with the length of the pellucid area.

There is a greater interval between its end and that of the pellucid area in front than behind.

At about the sixteenth hour, or a little later, a thickening of the mesoblast takes place in front of the primitive groove, forming an opaque streak, which in fresh specimens looks like a continuation from the anterior extremity of the primitive groove (vide Pl. 1, fig. 8). From hardened specimens, however, it is easy to see that the connection of this streak with the primitive groove is only an apparent one. Again, it is generally possible to see that in the central line of this streak there is a narrow [Pg 43] groove. I do not feel certain that there is no period when this groove may not be present, but its very early appearance has not been recognized either by Dursy or by Waldeyer. Moreover, both these authors, as also His, seem to have mistaken the opaque streak spoken of above for the notochord. This, however, is not the case, and the notochord does not make its appearance till somewhat later. The mistake is of very minor importance, and probably arose in Dursy's case from his not sufficiently making use of sections. At about the time the streak in front of the primitive groove makes its appearance a semicircular fold begins to be formed near the anterior extremity of the pellucid area, against which the opaque streak, or as it had, perhaps, better be called, the medullary streak, ends abruptly.

This fold is the head fold, and the groove along the medullary streak is the medullary groove, which subsequently forms the cavity of the medullary or neural canal.

Everything which I have described above can without difficulty be made out from the examination of fresh and hardened specimens under the simple microscope; but sections bring out still more clearly these points, and also shew other features which could not have been brought to light without their aid. In Pl. 1, figs. 6 and 7, two sections of an embryo of about eighteen hours are shewn. The first of these passes through the medullary groove, and the second of them through the extreme anterior end of the primitive groove. The points of difference in the two sections are very obvious.

From fig. 6 it is clear that a groove has already been formed in the medullary streak, a fact which was not obvious in the fresh specimen. In the second place the mesoblast is thickened both under the groove and also more especially in the medullary folds at the sides of the groove; but shews hardly a sign of the differentiation of the notochord. So that it is clear that the medullary streak is not the notochord, as was thought to be the case by the authors above mentioned. In the third place there is no adhesion between the epiblast and the mesoblast. In all the sections I have cut through the medullary groove I have found this feature to be constant; while (for instance, as in Pl. 1, figs. 7, 9, 17) all sections through the primitive groove [Pg 44] shew most clearly an adhesion between the epiblast and mesoblast. This fact is both strongly confirmatory of the separate origins of the medullary and primitive grooves, and is also important in itself, as leaving no loophole for supposing that in the region of embryo there is any separation of the cells from the epiblast to form the mesoblast.

By this time the primitive groove has attained its maximum growth, and from this time begins both absolutely to become smaller, and also gradually to be pushed more and more backwards by the growth of the medullary groove.

The specimen figured in Pl. 1, fig. 18, magnified about ten diameters, shews the appearance presented by an embryo of twenty-three hours. The medullary groove (mc) has become much wider and deeper than it was in the earlier stage; the medullary folds (A) are also broader and more conspicuous. The medullary groove widens very much posteriorly, and also the medullary folds separate far apart to enclose the anterior end of the primitive groove (pr).

All this can easily be seen with a simple microscope, but the sections taken from the specimen figured also fully bear out the interpretations given above, and at the same time shew that the notochord has at this age begun to appear. The sections marked 13-17 pass respectively through the lines with corresponding numbers in fig. 18. Section 1 (fig. 13) passes through the middle of the medullary canal.

In it the following points are to be noted. (1) That the epiblast becomes very much thinner where it lines the medullary canal (mc), a feature never found in the epiblast lining the primitive groove. (2) That the mesoblast is very much thickened to form the medullary folds at A, A, while there is no adherence between it and the epiblast, below the primitive groove. (3) The notochord (ch) has begun to be formed, though its separation from the rest of the mesoblast is not as yet very distinct[8].

In fig. 14 the medullary groove has become wider and the medullary folds broader, the notochord has also become more expanded: the other features are the same as in section 1. In the third section (fig. 15) the notochord is still more expanded; [Pg 45] the bottom of the now much expanded medullary groove has become raised to form the ridge which separates the medullary from the primitive groove. The medullary folds are also flatter and broader than in the previous section. Section 4 (fig. 16) passes through the anterior end of the primitive groove. Here the notochord is no longer visible, and the adherence between the mesoblast and epiblast below the primitive groove comes out in marked contrast with the entire separation of the two layers in the previous sections.

The medullary folds (A) are still visible outside the raised edges of the primitive groove, and are as distinctly as possible separate and independent formations, having no connection with the folds of the primitive groove. In the last section (fig. 17), which is taken some way behind section 4, no trace of the medullary folds is any longer to be seen, and the primitive groove has become deeper. This series of sections, taken in conjunction with the specimen figured in fig. 18, must remove all possible doubt as to the total and entire independence of the primitive and medullary grooves. They arise in different parts of the blastoderm; the one reaches its maximum growth before the other has commenced to be formed; and finally, they are distinguished by almost every possible feature by which two such grooves could be distinguished.

Soon after the formation of the notochord, the protovertebræ begin to be formed along the sides of the medullary groove (Pl. 1, fig. 19, pv). Each new protovertebra (of those which are formed from before backwards) arises just in front of the anterior end of the primitive groove. As growth continues, the primitive groove becomes pushed further and further back, and becomes less and less conspicuous, till at about thirty-six hours only a very small and curved remnant is to be seen behind the sinus rhomboidalis; but even up to the forty-ninth Dursy has been able to distinguish it at the hinder end of the embryo.

The primitive groove in the chick is, then, a structure which appears very early, and soon disappears without entering directly into the formation of any part of the future animal, and without, so far as I can see, any function whatever. It is clear, therefore, that the primitive groove must be the rudiment of some ancestral feature; but whether it is a rudiment of some [Pg 46] structure which is to be found in reptiles, or whether of some earlier form, I am unable to decide. It is just possible that it is the last trace of that involution of the epiblast by which the hypoblast is formed in most of the lower animals. The fact that it is formed in the hinder part of the pellucid area perhaps tells slightly in favour of this hypothesis, since the point of involution of the epiblast not unfrequently corresponds with the position of the anus.

EXPLANATION OF PLATE 1, Figs. 6-8 and 13-19.

Figs. 6 and 7 are sections through an embryo rather earlier than the one drawn in fig. 8. Fig. 6 passes through the just commencing medullary groove (md), which appears in fresh specimens, as in fig. 8, merely as an opaque streak coming from the end of the primitive groove. The notochord is hardly differentiated, but the complete separation of mesoblast and hypoblast under the primitive groove is clearly shewn. Fig. 7 passes through the anterior end of the primitive groove (pr), and shews the fusion between the mesoblast and epiblast, which is always to be found under the primitive groove.

Fig. 8 is a view from above of a twenty hours' blastoderm, seen as a transparent object. Primitive groove (pr). Medullary groove (md), which passes off from the anterior end of the primitive groove, and is produced by the thickening of the mesoblast. Head fold (pf).

Figs. 13-17 are sections through the blastoderm, drawn in fig. 18 through the lines 1, 2, 3, 4, 5 respectively.

The first section (fig. 13) passes through the true medullary groove (mc); the two medullary folds (A, A) are seen on each side with the thickened mesoblast, and the mesoblast cells are beginning to form the notochord (nc) under the medullary groove. There is no adherence between the mesoblast cells and the epiblast under the medullary groove.

The second (fig. 14) section passes through the medullary groove where it has become wider. Medullary folds, A, A; notochord, ch.

In the third section (fig. 15) the notochord (ch) is broader, and the epiblast is raised in the centre, while the medullary folds are seen far apart at A.

In section fig. 16 the medullary folds (A) are still to be seen enclosing the anterior end of the primitive groove (pr). Where the primitive groove appears there is a fusion of the epiblast and mesoblast, and no appearance of the notochord.

In the last section, fig. 17, no trace is to be seen of the medullary folds.

Figs. 18 and 19 are magnified views of two hardened blastoderms. Fig. 18 is twenty-three hours old; fig. 19 twenty-five hours. They both shew how the medullary canal arises entirely independently of the primitive groove and in front of it, and also how the primitive groove gets pushed backwards by the growth of the medullary groove. pv, Protovertebræ; other references as above. Fig. 18 is the blastoderm from which sections figs. 13-17 were cut.

[7] From the Quarterly Journal of Microscopical Science, Vol. XIII, 1873.

[8] In the figure the notochord has been made too distinct.

[Pg 47]

IV. The Development of the Blood-vessels of the Chick[9].

With Plate 2.

The development of the first blood-vessels of the yolk-sac of the chick has been investigated by a large number of observers, but with very discordant results. A good historical résumé of the subject will be found in a paper of Dr Klein (liii. Band der K. Akad. der Wissensch. Wien), its last investigator.

The subject is an important one in reference to the homologies of the blood-vascular system of the vertebrata. As I shall shew in the sequel (and on this point my observations agree with those of Dr Klein), the blood-vessels of the chick do not arise as spaces or channels between the cells of the mesoblast; on the contrary, they arise as a network formed by the united processes of mesoblast-cells, and it is through these processes, and not in the spaces between them, that the blood flows. It is, perhaps, doubtful whether a system of vessels arising in this way can be considered homologous with any vascular system which takes its origin from channels hollowed out in between the cells of the mesoblast.

My own researches chiefly refer to the development of the blood-vessels in the pellucid area. I have worked but very slightly at their development in the vascular area; but, as far as my observations go, they tend to prove that the mode of their origin is the same, both for the pellucid and the vascular area.

The method which I have principally pursued has been to examine the blastoderm from the under surface. It is very difficult to obtain exact notions of the mode of development of [Pg 48] the blood-vessels by means of sections, though these come in as a valuable confirmation of the other method.

For the purpose of examination I have employed (1) fresh specimens; (2) specimens treated with spirit, and then mounted in glycerine; (3) specimens treated with chloride of gold for about half a minute, and then mounted in glycerine; and (4) specimens treated with osmic acid.

All these methods bring out the same appearances with varying clearness; but the successful preparations made by means of the gold chloride are the best, and bring out the appearances with the greatest distinctness.

The first traces of the blood-vessels which I have been able to distinguish in the pellucid area are to be seen at about the thirtieth hour or slightly earlier, at about the time when there are four to five protovertebræ on each side.

Fig. 1 shews the appearance at this time. Immediately above the hypoblast there are certain cells whose protoplasm sends out numerous processes. These processes vary considerably in thickness and size, and quickly come in contact with similar processes from other cells, and unite with them.

I have convinced myself, by the use of the hot stage, that these processes continually undergo alteration, sometimes uniting with other processes, sometimes becoming either more elongated and narrower or broader and shorter. In this way a network of somewhat granular protoplasm is formed with nuclei at the points from which the processes start.

From the first a difference may be observed in the character of this network in different parts of the pellucid area. In the anterior part the processes are less numerous and thicker, the nuclei fewer, and the meshes larger; while in the posterior part the processes are generally very numerous, and at first thin, the meshes small, and the nuclei more frequent. As soon as this network commences to be formed the nuclei begin to divide. I have watched this take place with the hot stage. It begins by the elongation of the nucleus and division of the nucleolus, the parts of which soon come to occupy the two ends of the nucleus. The nucleus becomes still longer and then narrows in the centre and divides. By this means the nuclei become much more numerous, and are found in almost all the larger [Pg 49] processes. Whether they are carried out into the processes by the movement of the surrounding protoplasm, or whether they move through the protoplasm, I have been unable to determine; the former view, however, seems to be the most probable.

It is possible that some nuclei arise spontaneously in the protoplasm, but I am much more inclined to think that they are all formed by the division of pre-existing nuclei—a view favoured by the number of nuclei which are seen to possess two nucleoli. Coincidently with the formation of the new nuclei the protoplasm of the processes, as well as that surrounding the nuclei at the starting-points of the processes, begins to increase in quantity.

At these points the nuclei also increase more rapidly than elsewhere, but at first the resulting nuclei seem to be all of the same kind.

In the anterior part of the pellucid area (fig. 4) the increase in the number of nuclei and in the amount of protoplasm at the starting-points of the protoplasm is not very great, but in the posterior part the increase in the amount of the protoplasm at these points is very marked, and coincidently the increase in number of the nuclei is also great. This is shewn in figs. 2 and 3. These are both taken from the tail end of an embryo of about thirty-three hours, with seven or eight protovertebræ. Fig. 3 shews the processes beginning to increase in thickness, and also the protoplasm at the starting-points increasing in quantity; at the same time the nuclei at these points are beginning to become more numerous. Fig. 3 is taken from a slightly higher level, i.e. slightly nearer the epiblast. In it the protoplasm is seen to have increased still more in quantity, and to be filled with nuclei. These nuclei have begun to be slightly coloured, and one of them is seen to possess two nucleoli.

Very soon after this a change in the nuclei begins to be observed, more especially in the hinder part of the embryo. While before this time they were generally elongated, some of them now become more nearly circular. In addition to this, they begin to have a yellowish tinge, and the nuclei, when treated with gold (for in the fresh condition it is not easy to [Pg 50] see them distinctly), have a more jagged and irregular appearance than the nucleoli of the other nuclei.

This change takes place especially at the starting-points of the processes, so that the appearance presented (fig. 5) is that of spherical masses of yellowish nuclei connected with other similar spherical masses by protoplasmic processes, in which nuclei of the original type are seen imbedded. These masses are surrounded by a thin layer of protoplasm, at the edge of which a normal nucleus may here and there be detected, as at fig. 5, a and a´, the latter possessing two nucleoli. Some of these processes are still very delicate, and it is exceedingly probable that they undergo further changes of position before the final capillary system is formed.

These differentiated nuclei are the first stage in the formation of the blood-corpuscles. From their mode of formation it is clear that the blood-corpuscles of the Sauropsida are to be looked upon as nuclei containing nucleoli, rather than as cells containing nuclei; indeed, they seem to be merely ordinary nuclei with red colouring matter.

This would make them truly instead of only functionally homologous with the red corpuscles of the Mammalia, and would well agree with the fact that the red corpuscles of Mammalia, in their embryonic condition, possess what have previously been called nuclei, but which might perhaps more properly be called nucleoli.

In the anterior part of the blastoderm the processes, as I have stated, are longer and thinner, and the spaces enclosed between them are larger. This is clearly brought out in Pl. 2, fig. 4. But, besides these large spaces, there are other smaller spaces, such as that at v. It is, on account of the transparency of the protoplasm, very difficult to decide whether these are vacuoles or simply spaces enclosed by the processes, but I am inclined to think that they are merely spaces. The difficulty of exactly determining this point is increased by the presence of numerous white-yolk spherules in the hypoblast above, which considerably obscure the view. At about the same time that the blood-corpuscles appear in the posterior end of the pellucid area, or frequently a little later, they begin to be formed in the anterior part also. The [Pg 51] masses of them are, however, far smaller and far fewer than in the posterior part of the embryo. It is at the tail end of the pellucid area that the chief formation of blood-corpuscles takes place.

The part of the pellucid area intermediate in position between the anterior and posterior ends of the embryo is likewise intermediate as regards the number of corpuscles formed and the size of the spaces between the processes; the spaces being here larger than at the posterior extremity, but smaller than the spaces in front. Close to the sides of the embryo the spaces are, however, smaller than in any other part of the pellucid area. It is, however, in this part that the first formation of blood-corpuscles takes place, and that the first complete capillaries are formed.

We have then somewhat round protoplasmic masses filled with blood-corpuscles and connected by means of processes, a few of which may begin to contain blood-corpuscles, but the majority of which only contain ordinary nuclei. The next changes to be noticed take place in the nuclei which were not converted into blood-corpuscles, but which were to be seen in the protoplasm surrounding the corpuscles. They become more numerous and smaller, and, uniting with the protoplasm in which they were imbedded, become converted into flat cells (spindle-shaped in section), and in a short time form an entire investment for the masses of blood-corpuscles. The same change also occurs in the protoplasmic processes which connect the masses of corpuscles. In the case of those processes which contain no corpuscles the greater part of their protoplasm seems to be converted into the protoplasm of the spindle-shaped cells. The nuclei arrange themselves so as completely to surround the exterior of the protoplasmic processes. In this way each process becomes converted into a hollow tube, completely closed in by cells formed from the investment of the original nuclei by the protoplasm which previously formed the solid processes. The remainder of the protoplasm probably becomes fluid, and afterwards forms the plasma in which the corpuscles float. While these changes are taking place the formation of the blood-corpuscles does not stand still, and by the time a system of vessels, enclosed by cellular walls, is formed out of [Pg 52] the protoplasmic network, a large number of the connecting processes in this network have become filled with blood-corpuscles. The appearances presented by the network at a slightly later stage than this is shewn in Pl. 2, fig. 6, but in this figure all the processes are seen to be filled with blood-corpuscles.

This investment of the masses of corpuscles by a cellular wall occurs much earlier in some specimens than in others, both in relation to the time of incubation and to the completion of the network. It is generally completed in some parts by the time there are eight or nine protovertebræ, and is almost always formed over a great part of the pellucid area by the thirty-sixth hour. The formation of the corpuscles, as was pointed out above, occurs earliest in the central part of the hour-glass shaped pellucid area, and latest in its anterior part. In the hinder part of the pellucid area the processes, as well as their enlarged starting-points, become entirely filled with corpuscles; this, however, is by no means the case in its anterior part. Here, although the corpuscles are undoubtedly developed in parts as shewn in fig. 7, yet a large number of the processes are entirely without them. Their development, moreover, is in many cases very much later. When the development has reached the stage described, very little is required to complete the capillary system. There are always, of course, a certain number of the processes which end blindly, and others are late in their development, and are not by this time opened; but, as a general rule, when the cellular investment is formed for the masses of corpuscles, there is completed an open network of tubes with cellular walls, which are more or less filled with corpuscles. These become quickly driven into the opaque area in which at that time more corpuscles may almost always be seen than in the pellucid area.

By the formation of a network of this kind it is clear that there must result spaces enclosed between the walls of the capillaries; these spaces have under the microscope somewhat the appearance of being vesicles enclosed by walls formed of spindle-shaped cells. In reality they are only spaces enclosed at the sides, and, as a general rule, not above and below. They have been mistaken by some observers for vesicles in [Pg 53] which the corpuscles were supposed to be developed, and to escape by the rupture of the walls into the capillary spaces between. This mistake has been clearly pointed out by Klein (loc. cit.).

At the time when these spaces are formed, and especially in the hinder two-thirds of the pellucid area, and in the layer of blood-vessels immediately above the hypoblast, a formation takes place which forms in appearance a secondary investment of the capillaries. Dr Klein was the first to give a correct account of this formation. It results from the cells of the mesoblast in the meshes of the capillary system. Certain of these cells become flattened, and send out fine protoplasmic processes. They arrange themselves so as completely to enclose the spaces between the capillaries, forming in this way vesicles.

Where seen on section (vide fig. 6) at the edge of the vesicles these cells lining the vesicles appear spindle-shaped, and look like a secondary investment of the capillaries. This investment is most noticeable in the hinder two-thirds of the pellucid area; but, though less conspicuous, there is a similar formation in its anterior third, where there would seem to be only veins present. Dr Klein (loc. cit., fig. 12) has also drawn this investment in the anterior third of the pellucid area. He has stated that the vessels in the mesoblast between the splanchnopleure and the somatopleure, and which are enclosed by prolongations from the former, do not possess this secondary investment; he has also stated that the same is true for the sinus terminalis; but I am rather doubtful whether the generalisation will hold, that veins and arteries can from the first be distinguished by the latter possessing this investment. I am also rather doubtful whether the spaces enclosed by the protoplasmic threads between the splanchnopleure and somatopleure are the centres of vessels at all, since I have never seen any blood-corpuscles in them.

It is not easy to learn from sections much about the first stages in the formation of the capillaries, and it is impossible to distinguish between a completely-formed vessel and a mere spherical space. The fine protoplasmic processes which connect the masses of corpuscles can rarely be seen in sections, except when they pass vertically, as they do occasionally (vide Pl. 2, fig. 9) in the opaque area, joining the somatopleure and the [Pg 54] splanchnopleure. Dr Klein considers these latter processes to be the walls of the vessels, but they appear rather to be the processes which will eventually become new capillaries.

From sections, however, it is easy to see that the appearances of the capillaries in the vascular area are similar to the appearances in the pellucid area, from which it is fair to conclude that their mode of formation is the same in both. It is also easy to see that the first formation of vessels occurs in the splanchnopleure, and that even up to the forty-fifth hour but few or no vessels are found in the somatopleure. The mesoblast of the somatopleure is continued into the opaque area as a single layer of spindle-shaped cells.

Sections clearly shew in the case of most of the vessels that the secondary investment of Klein is present, even in the case of those vessels which lie immediately under the somatopleure.

In reference to the origin of particular vessels I have not much to say. Dr Klein's account of the origin of the sinus terminalis is quite correct. It arises by a number of the masses of blood-corpuscles, similar to those described above, becoming connected together by protoplasmic processes. The whole is subsequently converted into a continuous vessel in the usual way.

From the first the sinus terminalis possesses cellular walls, as is clear from its mode of origin. I am inclined to think that Klein is right in saying that the aortæ arise in a similar manner, but I have not worked out their mode of origin very fully.

It will be seen from the account given above that, in reference to the first stages in the development of the blood-vessels, my observations differ very considerably from those of Dr Klein; as to the later stages, however, we are in tolerable agreement. We are in agreement, moreover, as to the fundamental fact that the blood-vessels are formed by a number of cells becoming connected, and by a series of changes converted into a network of vessels, and that they are not in the first instance merely channels between the cells of the mesoblast.

By the forty-fifth hour colourless corpuscles are to be found in the blood whose exact origin I could not determine; probably they come from the walls of the capillaries.

[Pg 55] In the vessels themselves the coloured corpuscles undergo increase by division, as has already been shewn by Remak. Corpuscles in the various stages of division may easily be found. They do not appear to show very active amœboid movements in the vessels, though their movements are sometimes very active when removed from the body.

To recapitulate—some of the cells of the mesoblast of the splanchnopleure send out processes, these processes unite with the processes from other cells, and in this way a network is formed. The nuclei of the original cells divide, and at the points from which the processes start their division is especially rapid. Some of them acquire especially at these points a red colour, and so become converted into blood-corpuscles; the others, together with part of the protoplasm in which they are imbedded, become converted into an endothelium both for the processes and the masses of corpuscles; the remaining protoplasm becomes fluid, and thus the original network of the cells becomes converted into a network of hollow vessels, filled with fluid, in which corpuscles float.

In reference to the development of the heart, my observations are not quite complete. It is, however, easy to prove from sections (vide figs. 10 and 11, Pl. 2) that the cavity of the heart is produced by a splitting or absorption of central cells of the thickened mesoblast of the splanchnopleure, while its muscular walls are formed from the remaining cells of this thickened portion. It is produced in the following way:—When the hypoblast is folded in to form the alimentary canal the mesoblast of the splanchnopleure follows it closely, and where the splanchnopleure turns round to assume its normal direction (fig. 11) its mesoblast becomes thickened. This thickened mass of mesoblast is, as can easily be seen from figs. 10 and 11, Pl. 2, entirely distinct from the mesoblast which forms the outside walls of the alimentary canal. At the point where this thickening occurs an absorption takes place to form the cavity of the heart. The method in which the cavity is formed can easily be seen from figs. 10 and 11. It is in fig. 11 shewn as it takes place in the mesoblast on each side, the folds of the splanchnopleure not having united in the middle line; and hence a pair of cavities are formed, one on each side. It [Pg 56] is, however, probable that, in the very first formation of the heart, the cavity is single, being formed after the two ends of the folded mesoblast have united (vide hz, fig. 10). In some cases the two folds of the mesoblast appear not at first to become completely joined in the middle line; in this case the cavity of the heart is still complete from side to side, but the mesoblast-cells which form its muscular walls are deficient above. By the process of absorption, as I said, a cavity is produced in the thickened part of the mesoblast of the splanchnopleure, a cavity which is single in front, but becomes divided further behind, where the folds of the mesoblast have not united, into two cavities, to form the origin of the omphalomeseraic veins. As the folding proceeds backwards the starting-point of the omphalomeseraic veins is also pushed backwards, and the cavities which were before separated become joined together. From its first formation the heart is lined internally by an endothelium; this is formed of flattened cells, spindle-shaped in section. The exact manner of the origin of this lining I have not been able to determine; it is, however, probable that some of the central mesoblast-cells are directly converted into the cells of the endothelium.

I have obtained no evidence enabling me to determine whether Dr Klein is correct in stating that the cells of the mesoblast in the interior of the heart become converted partly into blood-corpuscles and partly into a cellular lining forming the endothelium of the heart, in the same way that the blood-vessels in the rest of the blastoderm are formed. But I should be inclined to think that it is very probable—certainly more probable than that the cavity of the heart is formed by a process of splitting taking place. Where I have used the word absorption in speaking of the formation of the cavity of the heart, I must be understood as implying that certain of the interior cells become converted into the endothelium, while others either form the plasma or become blood-corpuscles.

The originally double formation of the hinder part of the heart probably explains Dr Afanassiev's statement (Bullétin de l'Académ. Impériale de St Pétersb., tom. xiii, pp. 321-335), that he finds the endothelium of the heart originally dividing its interior into two halves; for when the partition of the mesoblast [Pg 57] which separated at first the two halves of the heart became absorbed, the endothelium lining of each of the originally separate vessels would remain complete, dividing the cavity of the heart into two parts. The partition in the central line is, however, soon absorbed.

The account given above chiefly differs from that of Remak by not supposing that the mesoblast-cells which form the heart are in any way split off from the wall of the alimentary canal.

There can be no doubt that His is wrong in supposing that the heart originates from the mesoblast of the splanchnopleure and somatopleure uniting to form its walls, thus leaving a cavity between them in the centre. The heart is undoubtedly formed out of the mesoblast of the splanchnopleure only.

Afanassiev's observations are nearer to the truth, but there are some points in which he has misinterpreted his sections.

Sections Pl. 2, figs. 10 and 11, explain what I have just said about the origin of the heart. Immediately around the notochord the mesoblast is not split, but a very little way outside it is seen to be split into two parts so and sp; the former of these follows the epiblast, and together with it forms the somatopleure, which has hardly begun to be folded at the line where the sections are taken. The latter (sp) forms with the hypoblast (hy) the splanchnopleure, and thus has become folded in to form the walls of the alimentary canal (d). In fig. 11 the folds have not united in the central line, but in fig. 10 they have so united. In fig. 11, where the mesoblast, still following the hypoblast, turns back to assume its normal direction, it is seen to be thickened and to have become split, so that a cavity (of) (of the omphalomeseraic vein) is formed in it on each side, lined by endothelium.

In the section immediately behind section fig. 11 the mesoblast was thickened, but had not become split.

In fig. 10 the hypoblast folds are seen to have united in the centre, so as to form a completely closed digestive canal (d); the folds of the mesoblast have also united, so that there is only a single cavity in the heart (hz), lined, as was the case with the omphalomeseraic veins, by endothelium.

In conclusion, I have to thank Dr Foster for his assistance and suggestions throughout the investigations which have formed [Pg 58] the subject of these three short papers, and which were well carried on in the apartments used by him as a Physiological Laboratory.

EXPLANATION OF PLATE 2.

Fig. 1 is taken from the anterior part of the pellucid area of a thirty hours' chick, with four protovertebræ. At n is a nucleus with two nucleoli.

Figs. 2 and 3 are taken from the posterior end of the pellucid area of a chick with eight protovertebræ. In fig. 3 the nuclei are seen to have considerably increased in number at the points of starting of the protoplasmic processes. At n is seen a nucleus with two nucleoli.

Fig. 4 is taken from the anterior part of the pellucid area of an embryo of thirty-six hours. It shews the narrow processes characteristic of the anterior part of the pellucid area, and the fewer nuclei. Small spaces, which have the appearance of vacuoles, are shewn at v.

Fig. 5 is taken from the posterior part of the pellucid area of a thirty-six hours' embryo. It shews the nuclei, with somewhat irregular nucleoli, which have begun to acquire the red colour of blood-corpuscles; the protoplasmic processes containing the nuclei; the nuclei in the protoplasm surrounding the corpuscles, as shewn at a, a´.

Fig. 6 shews fully formed blood-vessels, in part filled with blood-corpuscles and in part empty. The walls of the capillaries, formed of cells, spindle-shaped in section, are shewn, and also the secondary investment of Klein at k, and at b is seen a narrow protoplasmic process filled with blood-corpuscles.

Fig. 7 is taken from the anterior part of the pellucid area of a thirty-six hours' embryo. It shews a collection of nuclei which are beginning to become blood-corpuscles.

Figs. 1-5 are drawn with an 1/8 object-glass. Fig. 6 is on a much smaller scale. Fig. 7 is intermediate.

Fig. 8.—A transverse section through the dorsal region of a forty-five hours' embryo; ao, aorta with a few blood-corpuscles. v, Blood-vessels, all of them being formed in the splanchnopleure, and all of them provided with the secondary investment of Klein; pe, pellucid area; op, opaque area.

Fig. 9.—Small portion of a section through the opaque area of a thirty-five hours' embryo, showing protoplasmic processes, with nuclei passing from the somatopleure to the splanchnopleure.

Fig. 10.—Section through the heart of a thirty-four hours' embryo. a. Alimentary canal; hb, hind brain; nc, notochord; e, epiblast; so, mesoblast of the somatopleure; sp, mesoblast of the splanchnopleure; hy, hypoblast; hz, cavity of the heart.

[Pg 59] Fig. 11.—Section through the same embryo as fig. 10, and passing through the orifice of the omphalomeseraic vein. of, Omphalomeseraic vein; other references as above.

These two sections shew that the heart is entirely formed from the mesoblast of the splanchnopleure, and that it is formed by the splitting of that part of the mesoblast which has turned to assume its normal direction after being folded in to form the muscular wall of the alimentary canal. In fig. 11 the cavities so formed on each side have not yet united, but in fig. 10 they have united. When the folding becomes more complete the cavities (of, of) in fig. 11 will unite, and in this way the origin of the omphalomeseraic veins will be carried further backwards. In the section immediately behind section 11 the mesoblast had become thickened, but had not split.

[9] From the Quarterly Journal of Microscopical Science, Vol. XIII, 1873.

[Pg 60]

V. A Preliminary Account of the Development of the Elasmobranch Fishes[10].

With Plates 3 and 4.

During the spring of the present year I was studying at the Zoological Station, founded by Dr Dohrn at Naples, and entirely through its agency was supplied with several hundred eggs of various species of Dog-fish (Selachii)—a far larger number than any naturalist has previously had an opportunity of studying. The majority of the eggs belonged to an oviparous species of Mustelus, but in addition to these I had a considerable number of eggs of two or three species of Scyllium, and some of the Torpedo. Moreover, since my return to England, Professor Huxley has most liberally given me several embryos of Scyllium stellare in a more advanced condition than I ever had at Naples, which have enabled me to fill up some lacunæ in my observations.

On many points my investigations are not yet finished, but I have already made out a number of facts which I venture to believe will add to our knowledge of vertebrate embryology; and since it is probable that some time will elapse before I am able to give a complete account of my investigations, I have thought it worth while preparing a preliminary paper in which I have briefly, but I hope in an intelligible manner, described some of the more interesting points in the development of the Elasmobranchii. The first-named species (Mustelus sp.?) was alone used for the early stages, for the later ones I have also employed the other species, whose eggs I have had; but as far as I have [Pg 61] seen at present, the differences between the various species in early embryonic life are of no importance.

Without further preface I will pass on to my investigations.

The Egg-shell.

In the eggs of all the species of Dog-fishes which I have examined the yolk lies nearest that end of the quadrilateral shell which has the shortest pair of strings for attachment. This is probably due to the shape of the cavity of the shell, and is certainly not due to the presence of any structures similar to chalazæ.

The Yolk.

The yolk is not enclosed in any membrane comparable to the vitelline membrane of Birds, but lies freely in a viscid albumen which fills up the egg-capsule. It possesses considerable consistency, so that it can be removed into a basin, in spite of the absence of a vitelline membrane, without falling to pieces. This consistency is not merely a property of the yolk-sphere as a whole, but is shared by every individual part of it.

With the exception of some finely granular matter around the blastoderm, the yolk consists of rather small, elliptical, highly refracting bodies, whose shape is very characteristic and renders them easily recognizable. A number of striæ like those of muscle are generally visible on most of the spherules, which give them the appearance of being in the act of breaking up into a series of discs; but whether these striæ are normal, or produced by the action of water I have not determined.

Position of the Blastoderm.

The blastoderm is always situated, immediately after impregnation, near the pole of the yolk which lies close to the end of the egg-capsule. Its position varies a little in the different species and is not quite constant in different eggs of the same species. But this general situation is quite invariable. It is of about the same proportional size as the blastoderm of a bird.

Segmentation.

In a fresh specimen, in which segmentation has only just commenced, the blastoderm or germinal disc appears as a circular [Pg 62] disc, distinctly marked off by a dark line from the rest of the yolk. This line, as is proved by sections, is the indication of a very shallow groove. The appearance of sharpness of distinction between the germ and the yolk is further intensified by their marked difference of colour, the germ itself being usually of a darker shade than the remainder of the yolk; while around its edge, and apparently sharply separated from it by the groove before mentioned, is a ring of a different shade which graduates at its outer border into the normal shade of the yolk.

These appearances are proved by transverse sections to be deceptive. There is no sharp line either at the sides or below separating the blastoderm from the yolk. In the passage between the fine granular matter of the germ to the coarser yolk-spheres every intermediate size of granule is present; and, though the space between the two is rather narrow, in no sense of the word can there be said to be any break or line between them.

This gradual passage stands in marked contrast with what we shall find to be the case at the close of the segmentation. In the youngest egg which I had, the germinal disc was already divided into four segments by two furrows at right angles. These furrows, however, did not reach its edge; and from my sections I have found that they were not cut off below by any horizontal furrow. So that the four segments were continuous below with the remainder of the germ without a break.

In the next youngest specimen which I had, there were already present eighteen segments, somewhat irregular in size, but which might roughly be divided into an outer ring of larger spheres, separated, as it were, by a circular furrow from an inner series of smaller segments. The furrows in this case reached quite to the edge of the germinal disc.

The remarks I made in reference to the earlier specimen about the separation of the germ from the yolk apply in every particular to the present one. The external limit of the blastoderm was not defined by a true furrow, and the segmentation furrows still ended below without meeting any horizontal furrows, so that the blastoderm was not yet separated by any line from the remainder of the yolk, and the segments of which it was composed were still only circumscribed upon five sides. In [Pg 63] this particular the segmentation in these animals differs materially from that in the Bird, where the horizontal furrows appear very early.

In each segment a nucleus was generally to be seen in sections. I will, however, reserve my remarks upon the nature of the nuclei till I discuss the nuclei of the blastoderm as a whole.

For some little time the peripheral segments continue larger than the more central ones, but this difference of size becomes less and less marked, and before the segments have become too small to be seen with the simple microscope, their size appears to be uniform over the whole surface of the blastoderm.

In the blastoderms somewhat older than the one last described the segments have already become completely separate masses, and each of them already possesses a distinct nucleus. They form a layer one or two segments deep. The limits of the blastoderm are not, however, defined by the already completed segments, but outside these new segments continue to be formed around nuclei which appear in the yolk. At this stage there is, therefore, no line of demarcation between the germ and the yolk, but the yolk is being bored into, so to speak, by a continuous process of fresh segmentation.

The further segmentation of the already existing spheres, and the formation of new ones from the yolk below and to the sides, continues till the central cells acquire their final size, the peripheral ones being still large, and undefined towards the yolk. These also soon reach the final size, and the blastoderm then becomes rounded off towards the yolk and sharply separated from it.

The Nuclei of the Yolk.

Intimately connected with the segmentation is the appearance and history of a number of nuclei which arise in the yolk surrounding the blastoderm.

When the horizontal furrows appear which first separate the blastoderm from the yolk, the separation does not occur along the line of passage from the fine to the coarse yolk, but in the former at some distance from this line.

The blastoderm thus rests upon a mass of finely granular material, from which, however, it is sharply separated. At this [Pg 64] time there appear in this finely granular material a number of nuclei of a rather peculiar character.

They vary immensely in size—from that of an ordinary nucleus to a size greater than the largest blastoderm-cell.

In Pl. 3, fig. 1, n, is shewn their distribution in this finely granular matter and their variation in size. But whatever may be their size, they always possess the same characteristic structure. This is shewn in Pl. 3, figs. 1 and 2, n.

They are rather irregular in shape, with a tendency when small to be roundish, and are divided by a number of lines into distinct areas, in each of which a nucleolus is to be seen. The lines dividing them into these areas have a tendency (in the smaller specimens) to radiate from the centre, as shewn in Pl. 3, fig. 1.

These nuclei colour red with hematoxylin and carmine and brown with osmic acid, while the nucleoli or granules contained in the areas also colour very intensely with all the three above-named reagents.

With such a peculiar structure, in favourable specimens these nuclei are very easily recognised, and their distribution can be determined without difficulty. They are not present alone in the finely granular yolk, but also in the coarsely granular yolk adjoining it. They form very often a special row, sometimes still more markedly than in Pl. 3, fig. 1, along the floor of the segmentation cavity. They are not, however, found alone in the yolk. All the blastoderm-cells in the earlier stages possess precisely similar nuclei! From the appearance of the first nucleus in a segmentation-sphere till a comparatively late period in development, every nucleus which can be distinctly seen is found to be of this character. In Pl. 3, fig. 2, this is very distinctly shewn.

(1) We have, then, nuclei of this very peculiar character scattered through the sub-germinal granular matter, and also universally present in the cells of the blastoderm. (2) These nuclei are distributed in a special manner under the floor of the segmentation cavity on which new cells are continually appearing. Putting these two facts together, there would be the strongest presumption that these nuclei do actually become the nuclei of cells which enter the blastoderm, and such is [Pg 65] actually the case. In my account of the segmentation I have, indeed, already mentioned this, and I will return to it, but before doing so will enter more fully into the distribution of these nuclei in the yolk.

They appear in small numbers around the blastoderm at the close of segmentation, and round each one of them there may at this time be seen in osmic acid specimens, and with high powers, a fine network similar to but finer than that represented in Pl. 3, fig. 2. This network cannot, as a general rule, be traced far into the yolk, but in some exceptionally thin specimens it may be seen in any part of the fine granular yolk around the blastoderm, the meshes of the network being, however, considerably coarser between than around the nuclei. This network may be seen in the fine granular material around the germ till the latest period of which I have yet cut sections of the blastoderm. In the later specimens, indeed, it is very much more distinctly seen than in the earlier, owing to the fact that in parts of the blastoderm, especially under the embryo, the yolk-granules have disappeared partly or entirely, leaving only this fine network with the nuclei in it.

A specimen of this kind is represented in Pl. 3, fig. 2, where the meshes of the network are seen to be finer immediately around the nuclei, and coarser in the intervals. The specimen further shows in the clearest manner that this network is not divided into areas, each representing a cell and each containing a nucleus. I do not know to what extent this network extends into the yolk. I have never yet seen the limits of it, though it is very common to see the coarsest yolk-granules lying in its meshes. Some of these are shewn in Pl. 3, fig. 2, yk.

This network of lines[11] (probably bubbles) is characteristic of many cells, especially ova. We are, therefore, forced to believe that the fine granular and probably coarser granular yolk of this meroblastic egg consists of an active organized basis with [Pg 66] passive yolk-spheres imbedded in it. The organized basis is especially concentrated at the germinal pole of the egg, but becomes less and less in quantity, as compared with the yolk-spheres, the further we depart from this.

Admitting, as I think it is necessary to do, the organized condition of the whole yolk-sphere, there are two possible views as to its nature. We may either take the view that it is one gigantic cell, the ovum, which has grown at the expense of the other cells of the egg-follicle, and that these cells in becoming absorbed have completely lost their individuality; or we may look upon the true formative yolk (as far as we can separate it from the remainder of the food-yolk) as the remains of one cell (the primitive ovum), and the remainder of the yolk as a body formed from the coalescence of the other cells of the egg-follicle, which is adherent to, but has not coalesced with, the primitive ovum, the cells in this case not having completely lost their individuality; and to these cells, the nuclei, I have found, must be supposed to belong.

The former view I think, for many reasons, the most probable. The share of these nuclei in the segmentation, and the presence of similar nuclei in the cells of the germ, both support it, and are at the same time difficulties in the way of the other view. Leaving this question which cannot be discussed fully in a preliminary paper like the present one, I will pass on to another important question, viz.:

How do these nuclei originate? Are they formed by the division of the pre-existing nuclei, or by an independent formation? It must be admitted that many specimens are strongly in favour of the view that they increase by division. In the first place, they are often seen two together; examples of this will be seen in Pl. 3, fig. 1. In the second place, I have found several specimens in which five or six appear close together, which look very much as if there had been an actual division into six nuclei. It is, however, possible in this case that the nuclei are really connected below and only appear separate, owing to the crenate form of the mass. Against this may be put the fact that the division of a nucleus is by no means so common as has been sometimes supposed, that in segmentation it has very rarely been observed [Pg 67] that the nucleus of a sphere first divides[12], and that then segmentation takes place, but segmentation generally occurs and then a new nucleus arises in each of the newly formed spheres. Such nuclei as I have described are rare; they have, however, been observed in the egg of a Nephelis (one of the Leeches), and have in that case been said to divide. Dr Kleinenberg, however, by following a single egg through the whole course of its development, has satisfied himself that this is not the case, and that, further, these nuclei in Nephelis never form the nuclei of newly developing cells.

I must leave it an open question, and indeed one which can hardly be solved from sections, whether these nuclei arise freely or increase by division, but I am inclined to believe that both processes may possibly take place. In any case their division does not appear to determine the segmentation or segregation of the protoplasm around them.

As was mentioned in my account of the segmentation, these nuclei first appear during that process, and become the nuclei of the freshly formed segmentation spheres. At the close of segmentation a few of them are still to be seen around the blastoderm, but they are not very numerous.

From this period they rapidly increase in number, up to the commencement of the formation of the embryo as a body distinct from the germ. Though before this period they probably become the nuclei of veritable cells which enter the germ, it is not till this period, when the growth of the blastoderm becomes very rapid and it commences to spread over the yolk, that these new cells are formed in large numbers. I have many specimens of this age which shew the formation of these new cells with great clearness. This is most distinctly to be seen immediately below the embryo, where the yolk-spherules are few in number. At the opposite end of the blastoderm I believe that more of these cells are formed, but, owing to the presence of numerous yolk-spherules, it is much more difficult to make certain of this.

[Pg 68] As to the final destination of these cells, my observations are not yet completed. Probably a large number of them are concerned in the formation of the vascular system, but I will give reasons later on for believing that some of them are concerned in the formation of the walls of the digestive canal and of other parts.

I will conclude my account of these nuclei by briefly summarizing the points I have arrived at in reference to them.

A portion, or more probably the whole, of the yolk of the Dog-fish consists of organized material, in which nuclei appear and increase either by division or by a process of independent formation, and a great number of these subsequently become the nuclei of cells formed around them, frequently at a distance from the germ, which then travel up and enter it.

The formation of cells in the yolk, apart from the general process of segmentation, has been recognised by many observers. Kupffer (Archiv. für Micr. Anat., Bd. IV. 1868) and Owsjannikow (Entwicklung der Coregonus, Bulletin der Akad. St Petersburgh, Vol. XIX.) in osseous fishes[13], Ray Lankester (Annals and Mag. of Nat. Hist. Vol. IX. 1873, p. 81) in Cephalopoda, Götte (Archiv. für Micr. Anat. Vol. X.) in the chick, have all described a new formation of cells from the so-called food-yolk. The organized nature of the whole or part of this, previous to the formation of the cells from it, has not, however, as a rule, been distinctly recognised. In the majority of cases, as, for instance, in Loligo, the nucleus is not the first thing to be formed, but a plastide is first formed, in which a nucleus subsequently makes its appearance.

[Pg 69] Formation of the Layers.

Leaving these nuclei, I will now pass on to the formation of the layers.

At the close of segmentation the surface of the blastoderm is composed of cells of a uniform size, which, however, are too small to be seen by the aid of the simple microscope.

The cells of this uppermost layer are somewhat columnar, and can be distinguished from the remainder of the cells of the blastoderm as a separate layer. This layer forms the epiblast; and the Dog-fish agree with Birds, Batrachians, and Osseous fish in the very early differentiation of it.

The remainder of the cells of the blastoderm form a mass, many cells deep, in which it is impossible as yet or till a very considerably later period to distinguish two layers. They may be called the lower layer cells. Some of them near the edge of this mass are still considerably larger than the rest, but they are, as a whole, of a fairly uniform size. Their nuclei are of the same character as the nuclei in the yolk.

There is one point to be noticed in the shape of the blastoderm as a whole. It is unsymmetrical, and a much larger number of its cells are found collected at one end than at the other. This absence of symmetry is found in all sections which are cut parallel to the long axis of the egg-capsule. The thicker end is the region where the embryo will subsequently appear.

This very early appearance of distinction in the blastoderm between the end at which the embryo will appear, and the non-embryonic end is important, especially as it shews the affinity of the modes of development of Osseous fishes and the Elasmobranchii. Oellacher (Zeitschrift für Wiss. Zoologie, Vol. XXXIII. 1873) has shewn, and, though differing from him on many other points, on this point Götte (Arch. für Micr. Anat. Vol. IX. 1873) agrees with him, that a similar absence of symmetry by which the embryonic end of the germ is marked off, occurs almost immediately after the end of segmentation in Osseous fishes. In the early stages of development there are [Pg 70] a number of remarkable points of agreement between the Osseous fish and the Dog-fish, combined with a number of equally remarkable points of difference. Some of these I shall point out as I proceed with my description.

The embryonic end of the germ is always the one which points towards the pole of the yolk farthest removed from the egg-capsule.

The germ grows, but not very rapidly, and without otherwise undergoing any very appreciable change, for some time.

The growth at these early periods appears to be particularly slow, especially when compared with the rapid manner in which some of the later stages of the development are passed through.

The next important change which occurs is the formation of the so-called segmentation cavity.

This forms a very marked feature throughout the early stages. It appears, however, to have somewhat different relations to the blastoderm than the homologous structure in other vertebrates. In its earliest stage which I have observed, it appears as a small cavity in the centre of the lower layer cells. This grows rapidly, and its roof becomes composed of epiblast and only a thin lining of lower layer cells, while its floor is formed by the yolk (Pl. 3, fig. 3, sg). In the next and third stage (Pl. 3, fig. 4, sg) its floor is formed by a thin layer of cells, its roof remaining as before. It has, however, become a less conspicuous formation than it was; and in the last (fourth) stage in which it can be distinguished it is very inconspicuous, and almost filled up by cells.

What I have called the second stage corresponds to a period in which no trace of the embryo is to be seen. In the third stage the embryonic end of the blastoderm projects outwards to form a structure which I shall speak of as the embryonic rim, and in the fourth and last stage a distinct medullary groove is formed. For a considerable period during the second stage the segmentation cavity remains of about the same size; during the third stage it begins to be encroached upon, and becomes smaller both absolutely, and relatively to the increased size of the germ.

[Pg 71] The segmentation cavity of the Dog-fish most nearly agrees with that of Osseous fishes in its mode of formation and relation to the embryo.

Dog-fish resemble Osseous fish in the fact that their embryos are entirely formed from a portion of the germ which does not form part of the roof of the segmentation cavity, so that the cells forming the roof of the segmentation cavity take no share at any time in the formation of their embryos. They further agree with Osseous fish (always supposing that the descriptions of Oellacher, loc. cit., and Götte, Archiv. für Micr. Anat. Bd. IX. are correct) in the floor of the segmentation cavity being formed at one period by yolk. Together with these points of similarity there are some important differences.

(1) The segmentation cavity in the Osseous fish from the first arises as a cavity between the yolk and the blastoderm, and its floor is never at any period covered with cells. In the Dog-fish, as we have said above, both in the earlier and later periods the floor is covered with cells.

(2) The roof in the Dog-fish is invariably formed by the epiblast and a row of flattened lower layer cells.

According to both Götte and Oellacher the roof of the segmentation cavity in Osseous fishes is in the earlier stages formed alone of the two layers which correspond with the single layer forming the epiblast in the Dog-fish. In Osseous fishes it is very difficult to distinguish the various layers, owing to the similarity of their component cells. In Dog-fish this is very easy, owing to the great distinctness of the epiblast, and it appears to me, on this account, very probable that the two above-named observers may be in error as to the constitution of its roof in the Osseous fish. With both the Bird and the Frog the segmentation cavity of the Dog-fish has some points of agreement, and some points of difference, but it would take me too far from my present subject to discuss them.

When the segmentation cavity is first formed, no great changes have taken place in the cells forming the blastoderm. The upper layer—the epiblast—is composed of a single layer of columnar cells, and the remainder of the cells of blastoderm, [Pg 72] forming the lower layer, are of a fairly uniform size, and polygonal from mutual pressure. The whole edge of the blastoderm is thickened, but this thickening is especially marked at its embryonic end.

This thickened edge of the blastoderm is still more conspicuous in the next and second stage (Pl. 3, fig. 3).

In the second stage the chief points of progress, in addition to the increased thickness of the edge of the blastoderm, are—

(1) The increased thickness and distinctness of the epiblast, caused by its cells becoming more columnar, though it remains as a one-cell-thick layer.

(2) The disappearance of the cells from the floor of the segmentation cavity.

The lower layer cells have undergone no important changes, and the blastoderm has increased very little if at all in size.

From Pl. 3, fig. 3, it is seen that there is a far larger collection of cells at the embryonic than at the opposite end.

Passing over some rather unimportant stages, I will come to the next important one.

The general features of this (the third) stage in a surface view are—

(1) The increase in size of the blastoderm.

(2) The diminution in size of the segmentation cavity, both relatively and absolutely.

(3) The appearance of a portion of the blastoderm projecting beyond the rest over the yolk. This projecting rim extends for nearly half the circumference of the yolk, but is most marked at the point where the embryo will shortly appear. I will call it the embryonic rim.

These points are still better seen from sections than from surface views, and will be gathered at once from an inspection of Pl. 3, fig. 4.

The epiblast has become still more columnar, and is markedly thicker in the region where the embryo will appear. But its most remarkable feature is that at the outer edge of the embryonic rim (er) it turns round and becomes continuous with the lower layer cells. This feature is most important, and involves some peculiar modifications in the development. [Pg 73] I will, however, reserve a discussion of its meaning till the next stage.

The only other important feature of this stage is the appearance of a layer of cells on the floor of the segmentation cavity.

Does this layer come from an ingrowth from the thickened edge of the blastoderm, or does it arise from the formation of new cells in the yolk?

It is almost impossible to answer this question with certainty. The following facts, however, make me believe that the newly formed cells do play an important part in the formation of this layer.

(1) The presence at an earlier date of almost a row of nuclei under the floor of the segmentation cavity (Pl. 3, fig. 1).

(2) The presence on the floor of the cavity of such large cells as those represented in fig. 1, bd, cells which are very different, as far as the size and granules are concerned, from the remainder of the cells of the blastoderm.

On the other hand, from this as well as other sections, I have satisfied myself that there is a distinct ingrowth of cells from the embryonic swelling. It is therefore most probable that both these processes, viz. a fresh formation and an ingrowth, have a share in the formation of the layer of cells on the floor of the segmentation cavity.

In the next stage we find the embryo rising up as a distinct body from the blastoderm, and I shall in future speak of the body, which now becomes distinct as the embryo. It corresponds with what Kupffer (loc. cit.) in his paper on the Osseous Fishes has called the embryonic keel.This starting-point for speaking of the embryo as a distinct body is purely arbitrary and one merely of convenience. If I wished to fix more correctly upon a period which could be spoken of as marking the commencing formation of the embryo, I should select the time when structures first appear to mark out the portion of the germ from which the embryo becomes formed; this period would be in the Elasmobranchii, as in the Osseous fish, at the termination of segmentation, when the want of symmetry between the embryonic end of the germ and the opposite end first appears.

[Pg 74] I described in the last stage the appearance of the embryonic rim. It is in the middle point of this, where it projects most, that the formation of the embryo takes place. There appear two parallel folds extending from the edge of the blastoderm towards the centre, and cut off at their central end by another transverse fold. These three folds raise up, between them, a flat broadish ridge, the embryo (Pl. 3, fig. 5). The head end of the embryo is the end nearest the centre of the blastoderm, the tail end being the one formed by its (the blastoderm's) edge.

Almost from its first appearance this ridge acquires a shallow groove—the medullary groove (Pl. 3, fig. 5, mg)—along its middle line, where the epiblast and hypoblast are in absolute contact (vide fig. 6a, 7a, 7b, &c.) and where the mesoblast (which is already formed by this stage) is totally absent. This groove ends abruptly a little before the front end of the embryo, and is deepest in the middle and wide and shallow behind.

On each side of it is a plate of mesoblast equivalent to the combined vertebral and lateral plates of the Chick. These, though they cannot be considered as entirely the cause of the medullary groove, may perhaps help to make it deeper. In the parts of the germ outside the embryo the mesoblast is again totally absent, or, more correctly, we might say that outside the embryo the lower layer cells do not become differentiated into hypoblast and mesoblast, and remain continuous only with the lower of the two layers into which the lower layer cells become differentiated in the body of embryo. This state of things is not really very different from what we find in the Chick. Here outside the embryo (i.e. in the opaque area) there is a layer of cells in which no differentiation into hypoblast and mesoblast takes place, but the layer remains continuous rather with the hypoblast than the mesoblast.

There is one peculiarity in the formation of the mesoblast which I wish to call attention to, i.e. its formation as two lateral masses, one on each side of the middle line, but not continuous across this line (vide figs. 6a and 6b, and 7a and 7b). Whether this remarkable condition is the most primitive, [Pg 75] i.e. whether, when in the stage before this the mesoblast is first formed, it is only on each side of the middle line that the differentiation of the lower layer cells into hypoblast and mesoblast takes place, I do not certainly know, but it is undoubtedly a very early condition of the mesoblast. The condition of the mesoblast as two plates, one on each side of the neural canal, is precisely similar to its embryonic condition in many of the Vermes, e.g. Euaxes and Lumbricus. In these there are two plates of mesoblast, one on each side of the nervous cord, which are known as the Germinal streaks (Keimstreifen) (vide Kowalevsky Würmern u. Arthropoden; Mém. de l'Acad. Imp. St Pétersbourg, 1871).

From longitudinal sections I have found that the segmentation cavity has ceased by this stage to have any distinct existence, but that the whole space between the epiblast and the yolk is filled up with a mass of elongated cells, which probably are solely concerned in the formation of the vascular system. The thickened posterior edge of the blastoderm is still visible.

At the embryonic end of the blastoderm, as I pointed out in an earlier stage, the epiblast and the lower layer cells are perfectly continuous.

Where they join the epiblast, the lower layer cells become distinctly divided, and this division commenced even in the earlier stage, into two layers; a lower one, more directly continuous with the epiblast, consisting of cells somewhat resembling the epiblast-cells, and an upper one of more flattened cells (Pl. 3, fig. 4, m). The first of these forms the hypoblast, and the latter the mesoblast. They are indicated by hy and m in the figures. The hypoblast, as I said before, remains continuous with the whole of the rest of lower layer cells of the blastoderm (vide fig. 7b). This division into hypoblast and mesoblast commences at the earlier stage, but becomes much more marked during this one.

In describing the formation of the hypoblast and mesoblast in this way I have assumed that they are formed out of the large mass of lower layer cells which underlie the epiblast at the embryonic end of the blastoderm. But there is another and, in some ways, rather a tempting view, viz. [Pg 76] to suppose that the epiblast, where it becomes continuous with the hypoblast, in reality becomes involuted, and that from this involuted epiblast are formed the whole mesoblast and hypoblast.

In this case we would be compelled to suppose that the mass of lower layer cells which forms the embryonic swelling is used as food for the growth of the involuted epiblast, or else employed solely in the growth over the yolk of the non-embryonic portion of the blastoderm; but the latter possibility does not seem compatible with my sections.

I do not believe that it is possible, from the examination of sections alone, to decide which of these two views (viz. whether the epiblast is involuted, or whether it becomes merely continuous with the lower layer cells) is the true one. The question must be decided from other considerations.

The following ones have induced me to take the view that there is no involution, but that the mesoblast and hypoblast are formed from the lower layer cells.

(1) That it would be rather surprising to find the mass of lower layer cells which forms the embryo swelling playing no part in the formation of embryo.

(2) That the view that it is the lower layer cells from which the hypoblast and mesoblast are derived agrees with the mode of formation of these two layers in the Bird, and also in the Frog; since although, in the latter animal, there is an involution, this is not of the epiblast, but of the larger cells of the lower pole of the yolk, which in part correspond with what I have called the lower layer cells in the Dog-fish.

If the view be accepted that it is from the lower layer cells that the hypoblast and mesoblast are formed, it becomes necessary to explain what the continuity of the hypoblast with the epiblast means.

The explanation of this is, I believe, the keystone to the whole position. The vertebrates may be divided as to their early development into two classes, viz. those with holoblastic ova, in which the digestive canal is formed by an involution with the presence of an anus of Rusconi.

This class includes Amphioxus, the Lamprey, the Sturgeon, and Batrachians.

[Pg 77] The second class are those with meroblastic ova and no anus of Rusconi, and with an alimentary canal formed by the infolding of the sheet of hypoblast, the digestive canal remaining in communication with the food-yolk for the greater part of embryonic life by an umbilical canal.

This class includes the Elasmobranchii, Osseous fish, Reptiles, and Aves.

The mode of formation of the alimentary canal in the first class is clearly the more primitive; and it is equally clear that its mode of formation in the second class is an adaptation due to the presence of the large quantity of food-yolk.

In the Dog-fish I believe that we can see, to a certain extent, how the change from the one to the other of these modes of development of the alimentary canal took place.

In all the members of the first class, viz. Amphioxus, the Lamprey, the Sturgeon, and the Batrachians, the epiblast becomes continuous with the hypoblast at the so-called anus of Rusconi, and the alimentary canal, potentially in all and actually in the Sturgeon (vide Kowalevsky, Owsjannikow, and Wagner, Bulletin der Acad. d. St Petersbourg, Vol. XIV. 1870, "Entwicklung der Störe"), communicates freely at its extreme hind end with the neural canal. The same is the case in the Dog-fish. In these, when the folding in to form the alimentary canal on the one hand, and the neural on the other, takes place, the two foldings unite at the corner, where the epiblast and hypoblast are in continuity, and place the two tubes, the neural and alimentary, in free communication with each other[14].

There is, however, nothing corresponding with the anus of Rusconi, which merely indicates the position of the involution of the digestive canal, and subsequently completely closes up, though it nearly coincides in position with the true anus in the Batrachians, &c.

This remarkable point of similarity between the Dog-fish's development and the normal mode of development in the first class (the holoblastic) of vertebrates, renders it quite clear that the continuity of the epiblast and hypoblast in the Dogfish [Pg 78] is really the remnant of a more primitive condition, when the alimentary canal was formed by an involution. Besides the continuity between neural and alimentary canals, we have other remnants of the primitive involution. Amongst these the most marked is the formation of the embryonic rim, which is nothing less than the commencement of an involution. Its form is due to the flattened, sheet-like condition of the germ. In the mode in which the alimentary canal is closed in front I shall shew there are indications of the primitive mode of formation of the alimentary canal; and in certain peculiarities of the anus, which I shall speak of later, we have indications of the primitive anus of Rusconi; and finally, in the general growth of the epiblast (small cells of the upper pole of the Batrachian egg) over the yolk (lower pole of the Batrachian egg), we have an example of the manner in which the primitive involution, to form the alimentary canal, invariably disappears when the quantity of yolk in an egg becomes very great.

I believe that in the Dog-fish we have before our eyes one of the steps by which a direct mode of formation comes to be substituted for an indirect one by involution. We find, in fact, in the Dog-fish, that the cells from which are derived the mesoblast and hypoblast come to occupy their final position in the primitive arrangement of the cells during segmentation, and not by a subsequent and secondary involution.

This change in the mode of formation of the alimentary canal is clearly a result of change of mechanical conditions from the presence of the large food-yolk.

Excellent parallels to it will be found amongst the Mollusca. In this class the presence or absence of food-yolk produces not very dissimilar changes to those which are produced amongst vertebrates from the same cause.

The continuity of the hypoblast and epiblast at the embryonic rim is a remnant which, having no meaning or function, except in reference to the earlier mode of development, is likely to become lost, and in Birds no trace of it is any longer to be found.

I will not in the present preliminary paper attempt hypothetically to trace the steps by which the involution gradually [Pg 79] disappeared, though I do not think it would be very difficult to do so. Nor will I attempt to discuss the question whether the condition with a large amount of food-yolk (as seems more probable) was twice acquired—once by the Elasmobranchii and Osseous fishes, and once by Reptiles and Birds—or whether only once, the Reptiles and Birds being lineal descendants of the Dog-fish.

In reference to the former point, however, I may mention that the Batrachians and Lampreys are to a certain extent intermediate in condition between the Amphioxus and the Dog-fishes, since in them the yolk becomes divided during segmentation into lower layer cells and epiblast, but a modified involution is still retained, while the Dog-fish may be looked upon as intermediate between Birds and Batrachians, the continuity at the hind end between the epiblast and hypoblast being retained by them, though not the involution.

It may be convenient here to call attention to some of the similarities and some of the differences which I have not yet spoken of between the development of Osseous fish and the Dog-fish in the early stages. The points of similarity are—(1) The swollen edge of the blastoderm. (2) The embryo-swelling. (3) The embryo-keel. (4) The spreading of the blastoderm over the yolk-sac from a point corresponding with the position of the embryo, and not with the centre of the germ. The growth is almost nothing at that point, and most rapid at the opposite pole of the blastoderm, being less and less rapid along points of the circumference in proportion to their proximity to the embryonic swelling. (5) The medullary groove.

In external appearance the early embryos of Dog-fish and Teleostei are very similar; some of my drawings could almost be substituted for those given by Oellacher. This similarity is especially marked at the first appearance of the medullary groove. In the Dog-fish the medullary groove becomes converted into the medullary canal in the same way as in Birds and all other vertebrates, except Osseous fishes, where it comes to nothing, and is, in fact, a rudimentary structure. But in spite of Oellacher's assertions to the contrary, I am convinced from the similarity of its position and appearance to the true medullary groove in the Dog-fish, that the groove which appears [Pg 80] in Osseous fishes is the true medullary groove; although Oellacher and Kuppfer appear to have conclusively proved that it does not become converted into the medullary canal. The chief difference between the Dog-fish and Osseous fish, in addition to the point of difference about the medullary groove, is that the epiblast is in the Dog-fish a single layer, and not divided into nervous and epidermic layers as in Osseous fish, and this difference is the more important, since, throughout the whole period of development till after the commencement of the formation of the neural canal, the epiblast remains in Dog-fish as a one-cell-deep layer of cells, and thus the possibility is excluded of any concealed division into a neural and epidermic layer, as has been supposed to be the case by Stricker and others in Birds.

Development of the Embryo.

After the embryo has become definitely established, for some time it grows rapidly in length, without externally undergoing other important changes, with the exception of the appearance of two swellings, one on each side of its tail.

These swellings, which I will call the Caudal lobes (figs. 8 and 9, ts), are also found in Osseous fishes, and have been called by Oellacher the Embryonal saum. They are caused by a thickening of mesoblast on each side of the hind end of the embryo, at the edge of the embryonic rim, and form a very conspicuous feature throughout the early stages of the development of the Dog-fish, and are still more marked in the Torpedo (Pl. 3, fig. 9). Although from the surface the other changes which are visible are very insignificant, sections shew that the notochord is commencing to be formed.

I pointed out that beneath the medullary groove the epiblast and hypoblast were not separated by any interposed mesoblast. Along the line (where the mesoblast is deficient) which forms the long axis of the embryo, a rod-like thickening of the hypoblast appears (Pl. 3, figs. 7a and 7b, ch and ch´), first at the head end of the embryo, and gradually extends backwards. This is the rudiment of the notochord; it remains attached for some time to the hypoblast, and becomes separated from it first at [Pg 81] the head end of the embryo, and the separation is then carried backwards. This thickening of the hypoblast projects up and comes in contact with the epiblast, and in the later stages with bad (especially chromic-acid) specimens the line of separation between the epiblast and the thickening may become a little obscured, and might possibly lead to the supposition that a structure similar to that which has been called the axis cord was present. In all my best (osmic-acid) specimens the line of junction is quite clear; and any one who is aware how easily two separate masses of cells may be made indistinguishably to fuse together from simple pressure will not be surprised to find the occasional obscurity of the line of junction between the epiblast and hypoblast. In the earlier stage of the thickening there is never in the osmic-acid preparations any appearance of fusion except in very badly prepared ones. Its mode of formation will be quite clear without further description from an inspection of Pl. 3, figs. 7a and 7b, ch and ch´. Both are taken from one embryo. In fig. 7b, the most anterior of the two, the notochord has become quite separated from the hypoblast. In fig. 7a, ch, there is only a very marked thickening of hypoblast, which reaches up to the epiblast, but the thickening is still attached to the hypoblast. Had I had space to insert a drawing of a third section of the same embryo there would only have been a slight thickening of the hypoblast. In the earlier stage it will be seen, by referring to figs. 6a and 6b, that there is no sign of a thickening of the hypoblast. My numerous sections (all made from embryos hardened in osmic acid) shewing these points are so clear that I do not think there can be any doubt whatever of the notochord being formed as a thickening of the hypoblast. Two interpretations of this seem possible.

I mentioned that the mesoblast appeared to be primitively formed as two independent sheets, split off, so to speak, from the hypoblast, one on each side of the middle line of the embryo. If we looked upon the notochord as a third median sheet of mesoblast, split off from the hypoblast somewhat later than the other two, we should avoid having to admit its hypoblastic origin.

Professor Huxley, to whom I have shewn my specimens, strongly advocates this view.

[Pg 82] The other possibility is that the notochord is primitively a true hypoblastic structure which has only by adaptation become an apparently mesoblastic one in the higher vertebrates. In favour of this view are the following considerations:

(1) That this is the undoubtedly natural interpretation of the sections. (2) That the notochord becomes separated from the hypoblast after the latter has acquired its typical structure, and differs in that respect from the two lateral sheets of mesoblast, which are formed coincidently with the hypoblast by a homogeneous mass of cells becoming differentiated into two distinct layers. (3) That the first mode of looking at the matter really proves too much, since it is clear that by the same method of reasoning we could prove the mesoblastic origin of any organ derived from the hypoblast and budded off into the mesoblast. We would merely have to assert that it was really a mass of mesoblast budded off from the hypoblast rather later than the remainder of the mesoblast. Still, it must be admitted that the first view I have suggested is a possible, not to say a probable one, though the mode of arguing by which it can be upheld may be rather dangerous if generally applied. We ought not, however, for that reason necessarily to reject it in the present case. As Mr Ray Lankester pointed out to me, if we accept the hypoblastic origin of the notochord, we should find a partial parallel to it in the endostyle of Tunicates, and it is perhaps interesting to note in reference to it that the notochord is the only unsegmented portion of the axial skeleton.

Whether the strong à priori difficulties of the hypoblastic origin of the notochord are sufficient to counterbalance the natural interpretation of my sections, cannot, I think, be decided from the single case of the Dog-fish. It is to be hoped that more complete investigations of the Lamprey, &c., may throw further light upon the question.

Whichever view of the primitive origin of the notochord is the true one, its apparent origin is very instructive as illustrating the possible way in which an organ might come to change the layer to which it primarily belonged.

If the notochord is a true mesoblastic structure, it is easy to be seen how, by becoming separated from the hypoblast a little later than is the case with the Dog-fish, its mesoblastic [Pg 83] origin would become lost; while if, on the other hand, it is primitively a hypoblastic structure, we see from higher vertebrates how, by becoming separated from the hypoblast rather earlier than in the Dog-fish, viz. at the same time as the rest of the mesoblast, its primitive derivation from the hypoblast has become concealed.

The view seemingly held by many embryologists of the present day, that an organ, when it was primitively derived from one layer, can never be apparently formed in another layer, appears to me both unreasonable on à priori grounds, and also unsupported by facts.

I see no reason for doubting that the embryo in the earliest periods of development is as subject to the laws of natural selection as is the animal at any other period. Indeed, there appear to me grounds for the thinking that it is more so. The remarkable differences in allied species as to the amount of food-yolk, which always entail corresponding alterations in the development—the different modes of segmentation in allied species, such as are found in the Amphipoda and Isopoda—the suppression of many stages in freshwater species, which are retained in the allied marine species—are all instances of modifications due to natural selection affecting the earliest stages of development. If such points as these can be affected by natural selection I see no reason why the arrangement of individual cells (or rather primitive elements) should not also be modified; why, in fact, a mass of cells which was originally derived from one layer, but in the course of development became budded off from that layer and entered another layer, should not by a series of small steps cease ever to be attached to the original layer, but from the first moment it can be distinguished should be found as a separate mass in the second layer.

The change of layers will, of course, only take place where some economy is effected by it. The variations in the mode of development of the nervous system may probably be explained in this way.

If we admit that organs can undergo changes, as to the primitive layer from which they are derived, important consequences must follow.

It will, for instance, by no means be sufficient evidence of [Pg 84] two organs not being homologous that they are not developed from the same layer. It renders the task of tracing out the homologies from development much more difficult than if the ordinary view of the invariable correspondence of the three layers throughout the animal kingdom be accepted. Although I do not believe that this correspondence is invariable or exact, I think that we both find and should expect to find that it is, roughly speaking, fairly so.

Thus, the muscles, internal skeleton, and connective tissue are always placed in the adult between the skin (epidermis) and the epithelium of the alimentary canal.

We should therefore expect to find them, and, as a matter of fact, we always do find them, developed from a middle layer when this is present.

The upper layer must always and does always form the epidermis, and similarly the lower layer or hypoblast must form a part of the epithelium of the alimentary canal. A full discussion of this question would, however, lead me too far away from my present subject.

The only other point of interest which I can touch on in this stage is the commencing closure of the alimentary canal in the region of the head. This is shewn in Pl. 3, figs. 6a, 6b, 7b, n.a. From these figures it can be seen that the closing does not take place as much by an infolding as by an ingrowth from the side walls of the alimentary canal towards the middle line. In this abnormal mode of closing of the alimentary canal we have again, I believe, an intermediate stage between the mode of formation of the alimentary canal in the Frog and the typical folding in which occurs in Birds. There is, however, another point in reference to it which is still more interesting. The cells to form the ingrowth from the bottom (ventral) wall of the alimentary canal are derived by a continuous fresh formation from the yolk, being formed around the nuclei spoken of above (vide p. 63 et seq.). All my sections shew this with more or less clearness, especially those a little later than fig. 6b, in which the lower wall of the alimentary canal is nearly completed. This is the more interesting since, from the mode of formation of the alimentary canal in the Batrachians, &c., we might expect that the cells from the yolk would take [Pg 85] a share in its formation in the Dog-fish. I have not as yet made out for certain the share which is taken by these freshly formed cells of the yolk in the formation of any other organ.

By the completion of its lower wall in the way described, the throat early becomes a closed tube, its closing taking place before any other important changes are visible in the embryo from surface views.

A considerable increase in length is attained before other changes than an increase in depth of the medullary groove and a more complete folding off of the embryo from the blastoderm take place. The first important change is the formation of the protovertebræ.

These are formed by the lateral plates of mesoblast, which I said were equivalent at once to the vertebral and lateral plates in the Bird, becoming split by transverse divisions into cubical masses.

At the time when this occurs, and, indeed, up till a considerably later period, the mesoblast is not split into somatopleure and splanchnopleure, and it is not divided into vertebral and lateral plates. The transverse lines of division of the protovertebræ do not, however, extend to the outer edge of the undivided lateral plates.

The differences between this mode of formation of the protovertebræ and that occurring in Birds are too obvious to require pointing out. I will speak of them more fully when I have given the whole history of the protovertebræ of the Dog-fish.

I will only now say that I have had in the early stages to investigate the formation of the protovertebræ entirely by means of sections, the objects being too opaque to be otherwise studied.

The next change of any importance is the commencement of the formation of the head. The region of the head first becomes distinguishable by the flattening out of the germ at its front end.

The flattened-out portion of the germ grows rapidly, and forms a spatula-like termination to the embryo (Pl. 3, fig. 8).

In the region of the head the medullary groove is at first totally absent (vide section, Pl. 3, fig. 8a).

Indeed, as can be seen from fig. 8b, the laminæ dorsales, so [Pg 86] far from bending up at this stage, actually bend down in the opposite direction.

I am at present quite unable even to form a guess what this peculiar feature of the brain means. It, no doubt, has some meaning in reference to the vertebrate ancestry if we could only discover it. The peculiar spatula-like flattened condition of the head is also (vide loc. ant. cit.) apparently found in the Sturgeons; it must therefore almost undoubtedly be looked upon as not merely an accidental peculiarity.

While these changes have been taking place in the head not less important changes have occurred in the remainder of the body. In the first place the two caudal lobes have increased in size, and have become, as it were, pushed in together, leaving a groove between them (fig. 8, ts). They are very conspicuous objects, and, together with the spatula-like head, give the whole embryo an almost comical appearance. The medullary canal has by this time become completely closed in the region of the tail (figs. 8 and 8b).

It is still widely open in the region of the back, and, though more nearly closed again in the neck, is, as I have said, flattened out to nothing in the head.

The groove[15] between the two caudal lobes must not be confused (as may easily be done) with the medullary groove, which by the time the former groove has become conspicuous is a completely closed canal.

The vertebral plates are not divided (vide fig. 7) into a somatopleuric and splanchnopleuric layer by this stage, except in the region of the head (vide fig. 8b, pp), where there is a distinct space between the two layers, which is undoubtedly homologous with the pleuro-peritoneal cavity of the hinder portion of the body.

It is probably the same cavity which Oellacher (loc. cit.) calls in Osseous fishes the pericardial cavity. In the Dog-fish, at least, it has no connection with the pericardium. Of its subsequent history I shall say a few words when I come to speak of the later stages.

[Pg 87] The embryo does not take more than twenty-four hours in passing from this stage, when the head is a flat plate, to the stage when the whole neural canal (including the region of the head) is closed in. The other changes, in addition to the closing in of the neural canal, are therefore somewhat insignificant. The folding off of the embryo from the germ has, however, progressed considerably, and a portion of the hind gut is closed in below. This is accomplished, not by a tail-fold, as in Birds, but by two lateral folds, which cause the sides of the body to meet and coalesce below. At the extreme hind end, where the epiblast is continuous with the hypoblast, the lateral folds turn round, so to speak, and become continuous with the medullary folds, so that when the various folds meet each other an uninterrupted canal is found passing round from the neural into the alimentary canal, and placing these two in communication at the tail end of the body. Since I have already mentioned this, and spoken of its significance, I will not dwell on it further here.

The cranial flexure commences coincidently with the closing in of the neural canal in the region of the brain, and the division into fore, mid, and hind brain becomes visible at the same time as or even before the closing of the canal occurs. The embryo has now become more or less transparent, and protovertebræ, of which about twenty are present, can now be seen in the fresh specimens. The heart, however, is not yet formed.

Up to this period, a period at which the embryo becomes very similar in external appearance to any other vertebrate embryo, I have followed in my description a chronological order. I shall now cease to do so, since it would be too long for a preliminary notice of this kind, but shall confine myself to the history of a few organs whose development is either more important or more peculiar than that of the others.

The Protovertebræ.

I have thought it worth while to give a short history of the development of the protovertebræ, firstly, because it is very easy to follow this in the Dog-fish, and, secondly, because [Pg 88] I believe that the Dog-fish have more nearly retained the primitive condition of the protovertebræ than any other vertebrate whose embryology has hitherto been described with sufficient detail.

I intend to describe, at the same time, the development of the spinal nerves.

I left each lateral mass of mesoblast in my last stage as a plate which had not yet become split into a somatic and a splanchnic sheet (Pl. 3, fig. 8a, vp), but which had become cut by transverse lines (not, indeed, extending to the outer limit of the sheet, but as yet not cut off by longitudinal lines of cleavage) into segments, which I called protovertebræ.

This sheet of mesoblast is fairly thick at its proximal (upper) end, but thins off laterally to a sheet two cells deep, and its cells are so arranged as to foreshadow its subsequent splitting into somatic and splanchnic sheets. Its upper (proximal) end is at this stage level with the bottom of the neural canal, but soon begins to grow upwards, and at the same time the splitting into somatopleure and splanchnopleure commences (Pl. 3, fig. 10, so and sp).

The separation between the two sheets is first visible in its uppermost part, and thence extends outwards. By this means each of the protovertebræ becomes divided into two sheets, which are only connected at their upper ends and outside the region of the body. I speak of the whole lateral sheet as being composed of protovertebræ, because at this time no separation into vertebral and lateral plates can be seen; but I may anticipate matters by saying that only the upper portion of the sheet from the level of the top of the digestive canal, becomes subsequently the true protovertebræ. From this it is clear that the pleuro-peritoneal cavity extends primitively quite up to the top of the protovertebræ; and that thus a portion of a sheet of mesoblast, at first perfectly continuous with the splanchnic sheet from which is derived the muscular wall of the alimentary canal, is converted into a part of the voluntary muscular system of the body, having no connection whatever with the involuntary muscular system of the digestive tract.

The pleuro-peritoneal cavity is first distinctly formed at a [Pg 89] time when only two visceral clefts are present. Before the appearance of a third visceral cleft in a part of the innermost layer of each protovertebræ (which may be called the splanchnic layer, from its being continuous with the mesoblast of the splanchnopleure), opposite the bottom of the neural tube, some of the cells commence to become distinguishable from the rest, and to form a separate mass. This mass becomes much more distinct a little later, its cells being characterised by being spindle-shaped, and having an elongated nucleus which becomes deeply stained by reagents (Pl. 4, fig. 11, mp´). Coincidently with its appearance the young Dog-fish commences spontaneously to move rapidly from side to side with a kind of serpentine motion, so that, even if I had not traced the development of this differentiated mass of cells till it becomes a band of muscles close to the notochord, I should have had little doubt of its muscular nature. It is indicated in figs. 11, 12, 13, by the letters mp´. Its early appearance is most probably to be looked upon as an adaptation consequent upon the respiratory requirements of the young Dog-fish necessitating movements within the egg.

Shortly after this date, at a period when three visceral clefts are present, I have detected the first traces of the spinal nerves.

At this time they appear in sections as small elliptical masses of cells, entirely independent of the protovertebræ, and closely applied to the upper and outer corners of the involuted epiblast of the neural canal (Pl. 4, fig. 11, spn). These bodies are far removed from any mesoblastic structures, and at the same time the cells composing them are not similar to the cells composing the walls of the neural canal, and are not attached to these, though lying in contact with them. I have not, therefore, sufficient evidence at present to enable me to say with any certainty where the spinal nerves are derived from in the Dog-fish. They may be derived from the involuted epiblast of the neural canal, and, indeed, this is the most natural interpretation of their position.

On the other hand, it is possible that they are formed from wandering cells of the mesoblast—a possibility which, with our present knowledge of wandering cells, must not be thrown aside as altogether improbable.

[Pg 90] In any case, it is clear that the condition in the Bird, where the spinal nerves are derived from tissue of the protovertebræ, is not the primitive one. Of this, however, I will speak again when I have concluded my account of the development of the protovertebræ.

About the same time that the first rudiments of the nerves appear, the division of the mesoblast of the sides of the body into a vertebral and a lateral portion occurs. This division first appears in the region where the oviduct (Müller's duct) is formed (Pl. 4, fig. 11, ov).

At this part opposite the level of the dorsal aorta the two sheets, viz. the splanchnic and the somatic, unite together, and thus each lateral sheet of mesoblast becomes divided into an upper portion (fig. 11, mp), split up by transverse partitions into protovertebræ, and a lower portion not so split, but consisting of an outer layer, the true somatopleure, and an inner layer, the true splanchnopleure. These two divisions of the primitive plate are thus separated by the line at which a fusion between the mesoblast of the somatopleure and splanchnopleure takes place. The mass of cells resulting from the fusion at this point corresponds with the intermediate cell-mass of Birds (vide Waldeyer, Eierstock und Ei).

At the same time, in the upper of these two sheets (the protovertebræ), the splanchnic layer sends a growth of cells inwards towards the notochord and the neural canal. This growth is the commencement of the large quantity of mesoblastic tissue around the notochord, which is in part converted into the axial skeleton, and in part into the connective tissue adjoining this.

This mass of cells is at first quite continuous with the splanchnic layer of the protovertebræ, and I see no reason for supposing that it is not derived from the growth of the cells of this layer. The ingrowth to form it first appears a little after the formation of the dorsal aorta; but, as far as I have been able to see, its cells have no connection with the walls of the aorta.

What I have said as to the development of the skeleton-forming layer will be quite clear from figs. 11 and 12a; and from these it will also be clear, especially from fig. 11a, that [Pg 91] the outermost layer of this mass of cells, which was the primitive splanchnic layer of the protovertebræ, still retains its epithelial character, and so can easily be distinguished from those cells which will form the skeleton. In the next stage which I have figured (fig. 12a), this outer portion of the splanchnic layer is completely separated from the skeleton-forming cells, and at the same time, having united below as well as above with the outer (somatic) layer of the two layers of which the protovertebræ are formed, the two together form an independent mass (fig. 12, mp), similar in appearance and in every way homologous with the muscle-plate of Birds.

On the inner side of this, which we may now call the muscle-plate, is seen the bundle of earlier-developed muscles (fig. 12, mp´) which I spoke of before.

The section represented in fig. 12 is from a very considerably later embryo than that represented in fig. 11, so that the skeleton-forming cells, few in number in the earlier section, have become very numerous in the later one, and have grown up above the neural canal, and also below the notochord, between the digestive canal and the aorta. They have, moreover, changed their character; they were round before, now they have become stellate. As to their further history, it need only be said that the layer of them immediately around the notochord and neural canal forms the cartilaginous centra and arches of the vertebræ, and that the remaining portion of them, which becomes much more insignificant in size as compared with the muscles, forms the connective tissue of the skeleton and of the parts around and between the muscles.

A muscle-plate itself is at this stage (shewn in fig. 12) composed of an inner and an outer layer of columnar cells (splanchnic and somatic) united at the upper and lower ends of the plate, and on the inner of the two lies the more developed mass of muscles before spoken of (mp´).

Each of these plates now grows both upwards and downwards; and at the same time connective-tissue cells appear between the plates and epidermis; but from where they come I do not know for certain; very probably they are derived from the somatic layer of the muscle-plate.

While the muscle-plates continue to grow both upwards and [Pg 92] downwards, the cells of which they are composed commence to become elongated and soon acquire an unmistakably muscular character (Pl. 4, fig. 13, mp).

Before this has occurred the inner mass of muscles has also undergone further development and become a large and conspicuous band of muscles close to the notochord (fig. 13, mp´).

At the same time that the muscle-plates acquire the true histological character of muscle, septa of connective tissue grow in and divide them into a number of distinct segments which subsequently form separate bands of muscle. I will not say more in reference to the development of the muscular system than that the whole of the muscles of the body (apart from the limbs, the origin of whose muscular system I have not yet investigated) are derived from the muscle-plates which grow upwards above the neural canal and downwards to the ventral surface of the body.

During the time the muscle-plates have been undergoing these changes the nerve masses have also undergone developmental changes.

They become more elongated and fibrous, their main attachment to the neural tube being still at its posterior (dorsal) surface, near which they first appeared. Later still they become applied closely to the sides of the neural tube and send fibres to it below as well as above. Below (ventral to) the neural tube a ganglion appears, forming only a slight swelling, but containing a number of characteristic nerve-cells. The ganglion is apparently formed just below the junction of the anterior and posterior roots, though probably the fibres of the two roots do not mix till below it.

The main points which deserve notice in the development of the protovertebræ are—

(1) That at the time when the mesoblast becomes split horizontally into somatopleure and splanchnopleure the vertebral and lateral plates are one, and the splitting extends to the very top of the vertebral or muscle-plate, so that the future muscle-plates are divided into a splanchnic and somatic layer, the space between which is at first continuous with the pleuro-peritoneal cavity.

[Pg 93] (2) That the following parts are respectively formed by the vertebral and lateral plates:

(a) Vertebral plate. From the splanchnic layer of this, or from cells which appear close to and continuous with it, the skeleton, and connective tissue of the upper part of the body, are derived.

The remainder of the plate, consisting of a splanchnic and somatic layer, is entirely converted into the muscles of the trunk, all of which are derived from it.

(b) Between the vertebral plate and the lateral plate is a mass of cells where, as I mentioned above, the mesoblast of the somatopleure and splanchnopleure fuse together. This mass of cells is the equivalent of the intermediate cell mass of Birds (vide Waldeyer, Eierstock und Ei).

From it are derived the Wolffian bodies and duct, the oviduct, the ovaries and the testis, and the connective tissue of the parts adjoining these.

(c) The lateral plate. From the somatic layer of this is derived the connective tissue of the ventral half of the body; the mesoblast of the limbs, including probably the muscles, and certainly the skeleton. From its splanchnic layer are derived the muscles and connective tissue of the alimentary canal.

(3) The spinal nerves are developed independently of the protovertebræ, so that the protovertebræ of the Elasmobranchii do not appear to be of such a complicated structure as the protovertebræ of Birds.

The Digestive Canal.

I do not intend to enter into the whole history of the digestive canal, but to confine myself to one or two points of interest connected with it. These fall under two heads:

(1) The history of the portion of the digestive canal between the anus and the end of the tail where the digestive canal opens into the neural canal.

(2) Certain less well-known organs derived from the digestive canal.

[Pg 94] The anus is a rather late formation, but its position becomes very early marked out by the hypoblast of the digestive canal approaching at that point close to the surface, whilst receding to some little distance from it on either side. The portion of the digestive tract I propose at present dealing with is that between this point, which I will call, for the sake of brevity, the anus and the hind end of the body. This portion of the canal is at first very short; it is elliptical in section, and of rather a larger bore than the remainder of the canal. Its diameter becomes, however, slightly less as it approaches the tail, dilating again somewhat at its extreme end. It is lined by a markedly columnar epithelium. Though at first very short, its length increases with the growth of the tail, but at the same time its calibre continually becomes smaller as compared with the remainder[TN1] of the alimentary canal.

It commences to become smaller, first of all, near, though not quite, at its extreme hind end, and thus becomes of a conical shape; the base of the cone being just behind the anus, while the apex of the cone is situated a short distance from the hind end of the embryo. The extreme hind end, however, at the same time does not diminish in size, and becomes relatively (if not also absolutely) much larger in diameter than it was at first, as compared with the remainder of the digestive canal. It becomes, in fact, a vesicle or vesicular dilatation at the end of a conical canal.

Just before the appearance of the external gills this part of the digestive canal commences to atrophy. It begins to do so close to the terminal vesicle, which, however, still remains as or more conspicuous than it was before. The lumen of the canal becomes smaller and smaller, and finally it becomes a solid string of cells, and these also soon become indistinguishable and not a trace of the canal is left.

Almost the whole of it has disappeared before the vesicle begins to atrophy, but very shortly after all trace of the rest of the canal has vanished the terminal vesicle also vanishes. This occurs just about the time or shortly after the appearance of the external gills—there being slight differences probably in this respect in the different species.

In this history there are two points of especial interest:

[Pg 95] (1) The terminal vesicle.

(2) The disappearance of a large and well-developed portion of the alimentary canal.

The interest in the terminal vesicle lies in the possibility of its being some rudimentary structure.

In Osseous fishes Kupffer has described the very early appearance of a vesicle near the tail end, which he doubtfully speaks of as the allantois.The figure he gives of it in his earlier paper (Archiv. für Micro. Anat. Vol. II. pl. xxiv, fig. 2) bears a very strong resemblance to my figures of this vesicle at the time when the hind end of the alimentary canal is commencing to disappear; and I feel fairly confident that it is the same structure as I have found in the Dog-fish: but until the relations of the Kupffer's vesicle to the alimentary canal are known, any comparison between it and the terminal vesicle in the Dog-fish must be to a certain extent guess-work.

I have, however, been quite unsuccessful in finding any other vesicular structure which can possibly correspond to the so-called allantoic vesicle of Osseous fish.

The disappearance of a large portion of the alimentary canal behind the anus is very peculiar. In order, however, to understand the whole difficulties of the case I shall be obliged to speak of the relations of the anus of the Dog-fish to the anus of Rusconi in the Lamprey, &c.

In those vertebrates whose alimentary canal is formed by an involution, the anus of Rusconi represents the opening of this involution, and therefore the point where the alimentary canal primitively communicates with the exterior. When, however, the anus of Rusconi becomes closed, the wall of the alimentary canal still remains at that point in close juxtaposition to the surface, and the new and final anus is formed at or close to that point. In the Dog-fish, although the anus of Rusconi is not present, still, during the closing of the alimentary canal, the point which would correspond with this becomes marked out by the alimentary canal there approaching the surface, and it is at this point that the involution to form the true anus subsequently appears.

The anus in the Dog-fish has thus, more than a mere secondary significance. It corresponds with the point of closing of [Pg 96] the primitive involution. If it was not for this peculiarity of the vertebrate anus we would naturally suppose, from the disappearance of a considerable portion of the alimentary canal lying behind its present termination, that in the adult the alimentary canal once extended much farther back than at present, and that the anus we now find was only a secondary anus, and not the primitive one. It is perhaps possible that this hinder portion of the alimentary canal is a result of the combined growth of the tail and the persisting continuity (at the end of the body) of the epiblast with the hypoblast.

Whichever view is correct, it may be well to mention, in order to shew that the difficulty about the anus of Rusconi is no mere visionary one, that Götte (Untersuchung über die Entwicklung der Bombinator igneus, Archiv. für Micro. Anat., vol. V. 1869) has also described the disappearance of the hind portion of the alimentary canal in Batrachians, a rudiment (according to him) remaining in the shape of a lymphatic trunk.

It is, perhaps, possible that we have a further remnant of this hind portion of the alimentary canal amongst the higher vertebrates in the allantois.

Organs developed from the Digestive Canal.

In reference to the development of the liver, pancreas, &c., as far as my observations have at present gone, the Dog-fish presents no features of peculiar interest. The liver is developed as in the Bird, and independently of the yolk.

There are, however, two organs derived from the hypoblast which deserve more attention. Immediately under the notochord, and in contact with it (vide Pl. 3, fig. 10; Pl. 4, figs. 11 and 12, x), a small roundish (in section) mass of cells is to be seen in most of the sections.

Its mode of development is shewn in fig. 10, x. That section shows a mass of cells becoming pinched off from the top of the alimentary canal. By this process of pinching off from the alimentary canal a small rod-like body close under the notochord is formed. It persists till after the appearance of the external gills, but later than that I have not hitherto succeeded in finding any trace of it.

[Pg 97] It was first seen by Götte (loc. cit.) in the Batrachians, and he gave a correct account of its development, and added that it became the thoracic duct.

I have not myself worked out the later stages in the development of this body with sufficient care to be in a position to judge of the correctness of Götte's statements as to its final fate. If it is true that it becomes the thoracic duct it is very remarkable, and ought to throw some light upon the homologies of the lymphatic system.

Some time before the appearance of the external gills another mass of cells becomes, I believe, constricted off from the part of the alimentary canal in the neighbourhood of the anus, and forms a solid rod composed at first of dark granular cells lying between the Wolffian ducts. I have not followed out its development quite completely, but I have very little doubt that it is really constricted off from a portion of the alimentary canal chiefly in front of the point where the anus appears, but also, I believe, from a small portion behind this.

Though the cells of which it is composed are at first columnar and granular (fig. 12, su, r), they soon begin to become altered, and in the latter stage of its development the body forms a conspicuous rounded mass of cells with clear protoplasm, and each provided with a large nucleus. Later still it becomes divided into a number of separate areas of cells by septa of connective tissue, in which (the septa) capillaries are also present. Since I have not followed it to its condition in the adult, I cannot make any definite statements as to the fate of this body; but I think that it possibly becomes the so-called suprarenal organ, which in the Dog-fish forms a yellowish elongated body lying between the two kidneys.

The development of the Wolffian Duct and Body and of the Oviduct.

The development of the Wolffian duct and the Oviduct in the various classes of vertebrates is at present involved in some obscurity, owing to the very different accounts given by different observers.

[Pg 98] The manner of development of these parts in the Dog-fish is different from anything that previous investigators have met with in other classes, but I believe that it gives a clearer insight into the true constitution of these parts than vertebrate embryology has hitherto supplied, and at the same time renders easier the task of understanding the differences in the modes of development in the different classes.

I shall commence with a simple description of the observed facts, and then give my view as to their meaning. At about the time of the appearance of the third visceral cleft, and a short way behind the point up to which the alimentary canal is closed in front, the splanchnopleure and somatopleure fuse together opposite the level of the dorsal aorta.

From the mass of cells formed by this fusion a solid knob rises up towards the epiblast (Pl. 4, fig. 11b, ov), and from this knob a solid rod of cells grows backwards towards the tail (fig. 11c, ov) very closely applied to the epiblast. This description will be rendered clear by referring to figs. 11b and c. Fig. 11b is a section at the level of the knob, and fig. 11c is a section of the same embryo a short way behind this point. So closely does the rod of cells apply itself to the epiblast that it might very easily be supposed to be derived from it. Such, indeed, was at first my view till I cut a section passing through the knob. In order, however, to avoid all possibility of mistake I made sections of a large number of embryos of about the age at which this appears, and invariably found the large knob in front, and from it the solid string growing backwards.

This string is the commencement of the Oviduct or Müller's duct, which in the Dog-fish as in the Batrachians is the first portion of the genito-urinary system to appear, and is in the Dog-fish undoubtedly at first solid. All my specimens have been hardened with osmic acid, and with specimens hardened with this reagent it is quite easy to detect even the very smallest hole in a mass of cells.

As a solid string or rod of cells the Oviduct remains for some time; it grows, indeed, rapidly in length, the extreme hind end of the rod being very small and the front end continuing to remain attached to the knob. The knob, however, travels inwards and approaches nearer and nearer to the true pleuro-peritoneal [Pg 99] cavity, always remaining attached to the intermediate cell mass.

At about the time when five visceral clefts are present the Oviduct first begins to get a lumen and to open at its front end into the pleuro-peritoneal cavity. The cells of the rod are first of all arranged in an irregular manner, but gradually become columnar and acquire a radiating arrangement around a central point. At this point, where the ends of all the cells meet, a very small hole appears, which gradually grows larger and becomes the cavity of the duct (fig. 12, ov). The hole first makes its appearance at the anterior end of the duct, and then gradually extends backwards, so that the hind end is still without a lumen, when the lumen of the front end is of a considerable size.

At the front knob the same alteration in the cells takes place as in the rest of the duct, but the cells become deficient on the side adjoining the pleuro-peritoneal cavity, so that an opening is formed into the pleuro-peritoneal cavity, which soon becomes of a considerable size. Soon after its first formation, indeed, the opening becomes so large that it may be met in from two to three consecutive sections if these are very thin.

Thus is formed the lumen of the Oviduct. The duct still, at this age, ends behind without having become attached to the cloaca, so that at this time the Oviduct is a canal closed behind, but communicating in front by a large opening with the pleuro-peritoneal cavity.

It has during this time been travelling downwards, and is now much nearer the pleuro-peritoneal cavity than the epiblast.

It may be well to point out that the mode of development which I have described is really not very different from an involution, and must, in fact, be only looked upon as a modification of an involution. Many examples from all classes in the animal kingdom could be selected to exemplify how an involution may become simply a solid thickening. In the Osseous fish nearly all the organs which are usually formed by an involution have undergone this change in their mode of development. I shall attempt to give reasons later on for the solid form having been acquired in this particular case of the Oviduct.

[Pg 100] At about the time when a lumen appears in the Oviduct the first traces of the Wolffian duct become visible.

At intervals along the whole length, between the front and hind ends of the Oviduct, involutions arise from the pleuro-peritoneal cavity (fig. 12a, pwd) on the inside (nearer the middle line) of the Oviduct. The upper ends of these numerous involutions unite together and form a string of cells, at first solid, but very soon acquiring a lumen, and becoming a duct which communicates (as it clearly must from its mode of formation), at numerous points with the pleuro-peritoneal cavity. It is very probable that there is one involution to each segment of the body between the front and hind ends of the Oviduct. This duct is the Wolffian duct, which thus, together with the Oviduct, is formed before the appearance of the external gills.

For a considerable period the front end of the Oviduct does not undergo important changes; the hind end, however, comes into connection with the extreme end of the alimentary canal. The two Oviducts do not open together into the cloaca, though, as my sections prove, their openings are very close together. The whole Oviduct, as might be expected, shares in the general growth, and its lumen becomes in both sexes very considerably greater than it was before.

It is difficult to define the period at which I find these changes accomplished without giving drawings of the whole embryo. The stage is one considerably after the external gills have appeared, but before the period at which the growth of the olfactory bulbs renders the head of an elongated shape.

During the same period the Wolffian duct has undergone most important changes. It has commenced to bud off diverticula, which subsequently become the tubules of the Wolffian body (vide fig. 13, wd). I am fairly satisfied that the tubules are really budded off, and are not formed independently in the mesoblast. The Dog-fish agrees so far with Birds, where I have also no doubt the tubules of the Wolffian body are formed as diverticula from the Wolffian duct.

The Wolffian ducts have also become much longer than the Oviduct, and are now found behind the anus, though they do not extend as far forward as does the Oviduct.

[Pg 101] They have further acquired a communication with the Oviduct, in the form of a narrow duct passing from each of them into an Oviduct a short way before the latter opens into the cloacal dilatation of the alimentary canal.

The canals formed by the primitive involution leading from the pleuro-peritoneal cavity into the Wolffian duct have become much more elongated, and at the same time narrower. One of these is shewn in fig. 13, pwd.

Any doubt which could possibly be entertained as to the true character of the ducts whose development I have described is entirely removed by the development of the tubules of the Wolffian body. In the still later stage than this further proofs are furnished involving the function of the Oviduct. At the period when the olfactory lobes have become so developed as to render the head of the typical elongated shape of the adult, I find that the males and females can be distinguished by the presence in the former of the clasping appendages[16]. I find at this stage that in the female the front ends of the Oviducts have approached the middle line, dilated considerably, and commenced to exhibit at their front ends the peculiarities of the adult. In the male they are much less conspicuous, though still present.

At the same time the tubules of the Wolffian body become much more numerous, the Malpighian tufts appear, and the ducts cease almost, if not entirely, to communicate with the pleuro-peritoneal cavity. I have not made out anything very definitely as to the development of the Malpighian tufts, but I am inclined to believe that they arise independently in the mesoblast of the intermediate cell mass.

The facts which I have made out in reference to the development of the Wolffian duct, especially of its arising as a series of involutions from the pleuro-peritoneal cavity, will be found, I believe, of the greatest importance in understanding the true constitution of the Wolffian body. To this I will return directly, but first wish to clear the ground by insisting upon one preliminary point.

From their development the Oviduct and Wolffian body appear to stand to each other in the relation of the Wolffian [Pg 102] duct being the equivalent to a series, so to speak, of Oviducts.

I pointed out before that the mode of development of the Oviduct could only be considered as a modification of a simple involution from the pleuro-peritoneal cavity. Its development, both in the Birds and in the Batrachians as an involution, still more conclusively proves the truth of this view.

The explanation of its first appearing as a solid rod of cells which keeps close to the epiblast is, I am inclined to think, the following. Since the Oviduct had to grow a long way backwards from its primitive point of involution, it was clearly advantageous for it not to bore its way through the mesoblast of the intermediate cell mass, but to pass between this and the epiblast. This modification having been adopted, was followed by the knob forming the origin of the duct coming to be placed at the outside of the intermediate cell mass rather than close to the pleuro-peritoneal cavity, a change which necessitated the mode of development by an involution being dropped and the solid mode of development substituted for it, a lumen being only subsequently acquired.

In support of the modification in the development being due to this cause is the fact that in Birds a similar modification has taken place with the Wolffian duct. The Wolffian duct there arises differently from its mode of development in all the lower vertebrates as a solid rod close to the epiblast[17], instead of as an involution.

If the above explanation about the Oviduct be correct, then it is clear that similar causes have produced a similar modification in development (only with a different organ) in Birds; while, at the same time, the primitive mode of origin of the Oviduct (Müller's duct) has been retained by them.

The Oviduct, then, may be considered as arising by an involution from the pleuro-peritoneal cavity.

The Wolffian duct arises by a series of such involutions, all of which are behind (nearer the tail) the involution to form the Oviduct.

[Pg 103] The natural interpretation of these facts is that in the place of the Oviduct and Wolffian body there were primitively a series of similar bodies (probably corresponding in number with the vertebral segments), each arising by an involution from the pleuro-peritoneal cavity; and that the first of these subsequently became modified to carry eggs, while the rest coalesced to form the Wolffian duct.

If we admit that the Wolffian duct is formed by the coalescence of a series of similar organs, we shall only have to extend the suggestion of Gegenbaur as to the homology of the Wolffian body in order to see its true nature. Gegenbaur looks upon the whole urinogenital system as homologous with a pair of segmental organs. Accepting its homology with the segmental organs, its development in Elasmobranchii proves that it is not one pair, but a series of pairs of segmental organs with which the urinogenital system is homologous. The first of these have become modified so as to form the Oviducts, and the remainder have coalesced to form the Wolffian ducts.

The part of a segmental organ which opens to the exterior appears to be lost in the case of all but the last one, where this part is still retained, and serves as the external opening for all.

Whether the external opening of the first segmental organ (Oviduct) is retained or not is doubtful. Supposing it has been lost, we must look upon the external opening for the Wolffian body as serving also for the Oviduct. In the case of all other vertebrates whose development has been investigated (but the Elasmobranchii), the Wolffian duct arises by a single involution, or, what is equivalent to it, the other involutions having disappeared. This even appears to be the case in the Marsipobranchii. In the adult Lamprey the Wolffian duct terminates at its anterior end by a large ciliated opening into the pleuro-peritoneal cavity. It will, perhaps, be found, when the development of the Marsipobranchii is more carefully studied, that there are primitively a number of such openings[18]. The Oviduct, when present, arises in other vertebrates [Pg 104] as a single involution, strongly supporting the view that its mode of formation in the Dog-fish is fundamentally merely an involution.

The duct of the testes is, I have little doubt, derived from the anterior part of the Wolffian body; if so, it must be looked upon as not precisely equivalent to the Oviduct, but rather to a series of coalesced organs, each equivalent to the Oviduct. The Oviduct is in the Elasmobranchii, as in other vertebrates, primitively developed in both sexes. In the male, however, it atrophies. I found it still visible in the male Torpedos, though much smaller than in the females near the close of intra-uterine life.

Whether or not these theoretical considerations as to the nature of the Wolffian body and Oviduct are correct, I believe that the facts I have brought to light in reference to the development of these parts in the Dog-fish will be found of service to every one who is anxious to discover the true relations of these parts.

Before leaving the subject I will say one or[TN2] two words about the development of the Ovary. In both sexes the germinal epithelium (fig. 13) becomes thickened below the Oviduct, and in both sexes a knob (in section but really a ridge) comes to project into the pleuro-peritoneal cavity on each side of the mesentery (fig. 13, pov). In both sexes, but especially the females, the epithelium on the upper surface of this ridge becomes very much thickened, whilst subsequently it elsewhere atrophies. In the females, however, the thickened epithelium on the knob grows more and more conspicuous, and develops a number of especially large cells with large nuclei, precisely similar to Waldeyer's (loc. cit.) primitive ova of the Bird. In the male the epithelium on the ridge, though containing primitive ova, is not as conspicuous as in the female. Though I have not worked out the matter further than this at present, I still have no doubt that these projecting ridges become the Ovaries.

[Pg 105] The Head.

The study of the development of the parts of the head, on account of the crowding of organs which occurs there, always presents greater difficulties to the investigator than that of the remainder of the body. My observations upon it are correspondingly incomplete. I have, however, made out a few points connected with it in reference to some less well-known organs, which I have thought it worth while calling attention to in this preliminary account.

The continuation of the Pleuro-peritoneal Cavity into the Head.

In the earlier part of this paper (p. 86) I called attention to the extension of the separation between somatopleure and splanchnopleure into the head, forming a space continuous with the pleuro-peritoneal cavity (Pl. 3, fig. 8a, pp); this becomes more marked in the next stage, and, indeed, the pleuro-peritoneal cavity is present for a considerable time in the head before it becomes visible elsewhere. At the time of the appearance of the second visceral cleft it has become for the most part atrophied, but there persist two separated portions of it in front of the first cleft, and also remnants of it less well marked between and behind the two clefts. The visceral clefts necessarily divide it into separate parts.

The two portions in front of the first visceral cleft remain very conspicuous till the appearance of the external gills, and above the hinder one of the two the fifth nerve bifurcates.

These two are shewn as they appear in a surface view in fig. 14, pp. They are in reality somewhat flattened spaces, lined by a mesoblastic epithelium; the epithelium on the inner surface of the space corresponding to the splanchnopleure, and that on the outer to the somatopleure.

I have not followed the history of these later than the time of the appearance of the external gills.

The presence of the pleuro-peritoneal cavity in the head is interesting, as shewing the fundamental similarity between the head and the remainder of the body.

[Pg 106] The Pituitary Body.

All my sections seem to prove that it is a portion of the epiblastic involution to form the mouth which is pinched off to form the pituitary body, and not a portion of the hypoblast of the throat. Since Götte (Archiv. für Micr. Anat. Bd. IX.) has also found that the same is the case with the Batrachians and Mammalia, I have little doubt it will be found to be universally the case amongst vertebrates.

Probably the observations which lead to the supposition that it was the throat which was pinched off to form the pituitary body were made after the opening between the mouth and throat was completed, when it would naturally be impossible to tell whether the pinching off was from the epiblast of the mouth involution or the hypoblast of the throat.

The Cranial Nerves.

The cranial nerves in their early condition are so clearly visible that I have thought it worth while giving a figure of them, and calling attention to some points about their embryonic peculiarities.

From my figure (14) it will be seen that there is behind the auditory vesicle a nervous tract, from which four nerves descend, and that each of these nerves is distributed to the front portion of a visceral arch. When the next and last arch (in this species) is developed, a branch from this nervous mass will also pass down to it. That each of these is of an equal morphological value can hardly be doubted.

The nerve to the third arch becomes the glosso-pharyngeal (fig. 14, gl), the nerves to the other arches become the branchial branches of the vagus nerve (fig. 14, vg). Thus the study of their development strongly supports Gegenbaur's view of the nature of the vagus and glosso-pharyngeal, viz. that the vagus is a compound nerve, each component part of it which goes to an arch being equivalent to one nerve, such as the glosso-pharyngeal.

Of the nerves in front of the auditory sac the posterior is the seventh nerve (fig. 14, VII). Its mode of distribution to [Pg 107] the second arch leaves hardly a doubt that it is equivalent to one such nerve as those distributed to the posterior arches. Subsequently it acquires another branch, passing forwards towards the arch in front.

The most anterior nerve is the fifth (fig. 14, V), of which two branches are at this stage developed. The natural interpretation of its present condition is, that it is equivalent to two nerves, but the absence of relation in its branches to any visceral clefts renders it more difficult to determine the morphology of the fifth nerve than of the other nerves. The front branch of the two is the ophthalmic branch of the adult, and the hind branch the inferior maxillary branch. The latter branch subsequently gives off low down, i.e. near its distal extremity, another branch, the superior maxillary branch.

In its embryonic condition this latter branch does not appear like a third branch of the fifth, equivalent to the seventh or the glosso-pharyngeal nerves, but rather resembles the branch of the seventh nerve which passes to the arch in front, which also is present in all the other cranial nerves.

Modes of Preparation.

Before concluding I will say one or two words as to my modes of preparation.

I have used picric and chromic acids, both applied in the usual way; but for the early stages I have found osmic acid by far the most useful reagent. I placed the object to be hardened, in osmic acid (half per cent.) for two hours and a half, and then for twenty four in absolute alcohol.

I then embedded and cut sections of it in the usual way, without staining further.

I found it advantageous to cut sections of these embryos immediately after hardening, since if kept for long in the absolute alcohol the osmic acid specimens are apt to become brittle.

[Pg 108] LIST OF WORKS REFERRED TO.

Gegenbaur. Anat. der Wirbelthiere, III Heft, Leipzig, 1873.

A. Götte. Archiv. für Micr. Anat., Vol. X. 1873. Der Keim der Forelleneies, Archiv. Für. Micr. Anat., Vol. IX. 1873. Untersuchung über die Entwicklung der Bombinator igneus, Archiv. für Micr. Anat., Vol. V. 1869. Kurze Mittheilungen aus der Entwicklungsgeschichte der Unke, Archiv. für Micr. Anat., Vol. IX. 1873.

Kupffer. Archiv. für Micr. Anat., Vol. II. 1866, p. 473. Ibid. Vol. IV. 1868, p. 209.

Kowalevsky. Entwicklungsgeschichte der Holothurien, Mémoires de l'Acad. Impér. des Sciences de St Petersbourg, vii ser. Vol. XI. 1867.

Kowalevsky, Owsjannikow, und Wagner. Entwicklung der Störe, Bulletin der K. Acad. St Petersbourg, Vol. XIV. 1873.

Kowalevsky. Embryologische Studien an Würmern und Arthropoden, Mémoirs de l'Acad. Impér. des Sciences de St Petersbourg, Vol. XIV. 1871.

E. Ray Lankester. Annals and Mag. of Nat. History, Vol. XI. 1873, p. 81.

W. Müller. Ueber die Persistenz der Urniere bei Myxine Glutinosa, Jenaische Zeitschrift, Vol. VII. 1873.

Oellacher. Zeitschrift für Wiss. Zoologie, Vol. XXIII. 1873.

Owsjannikow. Entwicklung der Coregonus, Bul. der K. Akad. St Petersbourg, Vol. XIX.

Romiti. Archiv. für Micr. Anat., Vol. IX. 1873.

Waldeyer. Eierstock u. Eie.

EXPLANATION OF PLATES 3 AND 4.

COMPLETE LIST OF REFERENCE LETTERS.

al. Alimentary canal. ao. Dorsal aorta. auv. Auditory vesicle. bd. Formative cell probably derived from the yolk. cav. Cardinal vein. ch. Notochord. ch´. Thickening of hypoblast to form the notochord. Eb. Line indicating the edge of the blastoderm. ep. Epiblast. ep´. Epidermis. er. Embryonic rim. es. Embryonic swelling. gl. Glosso-pharyngeal nerve. h. Head. ht. Heart. hy. Hypoblast. ll. Lower layer cells. ly. Line of separation between the blastoderm and the yolk. m. Mesoblast. mc. Medullary canal. mg. Medullary groove. mp. Muscle-plate. mp´. Early formed mass of muscles. n. Peculiar nuclei formed in the yolk. n´. Similar nuclei in the cells of the blastoderm. na. Cells which help to close in the alimentary canal, and which are derived from the yolk. ny. Network of lines present in the food-yolk. ol. Olfactory pit. op. Eye. ov. Oviduct. pn. Pineal gland. pov. Projection which becomes the ovary. pp. Pleuro-peritoneal cavity. pp´. Remains of pleuro-peritoneal cavity in the head. prv. Protovertebræ. pwd. Primary points of involution from the pleuro-peritoneal cavity by the coalescence of which the Wolffian duct is formed. sg. Segmentation cavity. so. Somatopleure. sos. Stalk connecting embryo with yolk-sac. sp. Splanchnopleure. spn. [Pg 109] Spinal nerve. sur. Suprarenal body. ts. Caudal lobes. v. Blood-vessel. vg. Vagus nerve. V. Fifth nerve. VII. Seventh nerve. vc, 1, 2, 3, &c. 1st, 2nd and 3rd &c. visceral clefts. vp. Vertebral plates. wd. Wolffian duct. x. Peculiar body underlying the notochord derived from the hypoblast. yk. Yolk spherules.

All the figures were drawn with the Camera Lucida.

Plate 3.

Fig. 1. Section parallel with the long axis of the embryo through a blastoderm, in which the floor of the segmentation cavity (sg) is not yet completely lined by cells. The roof of the segmentation cavity is broken. (Magnified 60 diam.) The section is intended chiefly to illustrate the distribution of nuclei (n) in the yolk under the blastoderm. One of the chief points to be noticed in their distribution is the fact that they form almost a complete layer under the floor of the segmentation cavity. This probably indicates that the cells whose nuclei they become take some share in forming the layer of cells which subsequently (vide fig. 4) forms the floor of the cavity.

Fig. 2. Small portion of blastoderm and subjacent yolk of an embryo at the time of the first appearance of the medullary groove. (Magnified 300 diam.)

The specimen is taken from a portion of the blastoderm which will form part of the embryo. It shews two large nuclei of the yolk (n) and the network in the yolk between them; this network is seen to be closer around the nuclei than in the intervening space. The specimen further shews that there are no areas representing cells around the nuclei.

Fig. 3. Section parallel with the long axis of the embryo through a blastoderm, in which the floor of the segmentation cavity is not yet covered by a complete layer of cells. (Magnified 60 diam.)

It illustrates (1) the characters of the epiblast, (2) the embryonic swelling (es), (3) the segmentation cavity (sg). It should have been drawn upon the same scale as fig. 4; the line above it represents its true length upon this scale.

Fig. 4. Longitudinal section through a blastoderm at the time of the first appearance of the embryonic rim, and before the formation of the medullary groove. (Magnified 45 diam.)

It illustrates (1) the embryonic rim, (2) the continuity of epiblast and hypoblast at edge of this, (3) the continual differentiation of the lower layer cells, to form, on the one hand, the hypoblast, which is continuous with the epiblast, and on the other the mesoblast, between this and the epiblast; (4) the segmentation cavity, whose floor of cells is now completed.

N.B. The cells at the embryonic end of the blastoderm have been made rather too large.

Fig. 5. Surface view of the blastoderm shortly after the appearance of the medullary groove. To shew the relation of the embryo to the blastoderm.

Fig. 6a and b. Two transverse sections of the same embryo, shortly after the appearance of the medullary groove. (Magnified 96 diam.)

a. In the region of the groove. It shews (1) the two masses of mesoblast on each side, and the deficiency of the mesoblast underneath the medullary groove; (2) the commencement of the closing in of the alimentary canal below, chiefly from cells (na) derived from the yolk.

b. Section in the region of the head where the medullary groove is deficient, other points as above.

[Pg 110] Fig. 7a and b. Two transverse sections of an embryo about the age or rather younger than that represented in fig. 5. (Magnified 96 diam.)

a. Section nearer the tail; it shews the thickening of the hypoblast to form the notochord (ch´).

In b the thickening has become completely separated from the hypoblast as the notochord. In a the epiblast and hypoblast are continuous at the edge of the section, owing to the section passing through the embryonic rim.

Fig. 8. Surface view of a spatula-shaped embryo. The figure shews (1) the flattened head (h) where the medullary groove is deficient, (2) the caudal lobes, with a groove between them; it also shews that at this point, the medullary groove has become roofed over and converted into a canal.

Fig. 8a. Transverse section of fig. 8, passing through the line a. (Magnified 90 diam.) The section shews (1) the absence of the medullary groove in the head and the medullary folds turning down at this time instead of upwards; (2) the presence of the pleuro-peritoneal cavity in the head (pp); (3) the completely closed alimentary canal (al).

Fig. 8b. Transverse section of fig. 8, through the line b. (Magnified 90 diam.) It shews (1) the neural canal completely formed; (2) the vertebral plates of mesoblast not yet split up into somatopleure and splanchnopleure.

Fig. 9. Side view of an embryo of the Torpedo, seen as a transparent object a little older than the embryo represented in fig. 8. (Magnified 20 diam.) The internal anatomy has hardly altered, with the exception of the medullary folds having closed over above the head and the whole embryo having become more folded off from the germ.

The two caudal lobes, and the very marked groove between them, are seen at ts. The front end of the notochord became indistinct, and I could not see its exact termination. The epithelium of the alimentary canal (al) is seen closely underlying the notochord and becoming continuous with the epiblast at the hind end of the notochord.

The first visceral cleft (1vc) and eye (op) are just commencing to be formed, and the cranial flexure has just appeared.

Fig. 10. Section through the dorsal region of an embryo somewhat older than the one represented in fig. 9. (Magnified 96 diam.)

It shews (1) the formation by a pinching off from the top of the alimentary canal of a peculiar body which underlies the notochord (x); (2) the primitive extension of the pleuro-peritoneal cavity up to the top of the vertebral plates.

Plate 4.

Fig. 11a, b, and c. Three sections closely following each other from an embryo in which three visceral clefts are present; a is the most anterior of the three. (Magnified 96 diam.) In all of these the muscle-plates are shewn at mp. They have become separated from the lateral plates in b and c, but are still continuous with them in a. The early formed mass of muscles is also shewn in all the figures (mp´).

The figures further shew (1) the formation of the spinal nerves (spn) as small bodies of cells closely applied to the upper and outer edge of the neural canal.

(2) The commencing formation of the cells which form the axial skeleton from the inner (splanchnopleuric) layer of the muscle-plate. Sections b and c are given more especially to shew the mode of formation of the oviduct (ov).

[Pg 111] In b it is seen as a solid knob (ov), arising from the point where the somatopleure and splanchnopleure[TN3] unite, and in c (the section behind b) as a solid rod (ov) closely applied to the epiblast, which has grown backwards from the knob seen in b.

N.B. In all three sections only one side is completed.

Fig. 12a and b. Two transverse sections of an embryo just before the appearance of the external gills. (Magnified 96 diam.)

In a there is seen to be an involution on each side (pwd), while b is a section from the space between two involutions from the pleuro-peritoneal cavity, so that the Wolffian duct (at first solid) (wd) is not connected as in a with the pleuro-peritoneal cavity. The further points shewn in the sections are—

(1) The commencing formation of the spiral valve (al).

(2) The suprarenal body (sur).

(3) The oviduct (ov), which has acquired a lumen.

(4) The increase in length of the muscle-plates, the spinal nerves, &c.

Fig. 13. Section through the dorsal region of an embryo in which the external gills are of considerable length. (Magnified 40 diam.) The chief points to be noticed:

(1) The formation of the Wolffian body by outgrowths from the Wolffian duct (wd).

(2) One of the still continuing connections (primitive involutions) between the Wolffian duct and the pleuro-peritoneal cavity (pwd).

(3) The oviduct largely increased in size (ov).

N.B. On the left side the oviduct has been accidentally made too small.

(4) The growth downwards of the muscle-plate to form the muscles of the abdomen.

(5) The formation of an outgrowth on each side of the mesentery (pov), which will become the ovary.

(6) The spiral valve (al).

Fig. 14. Transparent view of the head of an embryo shortly before the appearance of the external gills. (Magnified 20 diam.) The chief points to be noticed are—

(1) The relation of the cranial nerves to the visceral clefts and the manner in which the glosso-pharyngeal (gl) and vagus (vg) are united.

(2) The remnants of the pleuro-peritoneal cavity in the head (pp).

(3) The eye (op). The stalk, as well as the bulb of the eye, are supposed to be in focus, so that the whole eye has a somewhat peculiar appearance.

[10] From the Quarterly Journal of Microscopical Science, Vol. XIV. 1874. Read in Section D, at the Meeting of the British Association at Belfast.

[11] The interpretation of this network is entirely due to Dr Kleinenberg, who suggested it to me on my shewing him a number of specimens exhibiting the nuclei and network.

[12] Kowalevsky (Beiträge zur Entwicklungsgeschichte der Holothurien, Mémoirs de l'Ac. Imp. de St Petersbourg, vii ser., Vol. XI. 1867) describes the division of nuclei during segmentation in the Holothurians, and other observers have described it elsewhere.

[13] Götte, at the end of a paper on The Development of the Layers in the Chick (Archiv. für Micr. Anat., Vol. X. 1873, p. 196), mentions that the so-called cells in Osseous fishes which Oellacher states to have migrated into the yolk, and which are clearly the same as those mentioned by Owsjannikow, are really not cells, but large nuclei. If this statement is correct the phenomena in Osseous fishes are precisely the same as those I have described in the Dog-fish.

[14] This has been already made out by Kowalevsky, Würmern u. Arthropoden, loc. cit.

[15] This groove is the only structure which it seems possible to compare with the so-called primitive groove of Birds. It is, however, doubtful whether they are really homologous.

[16] For the specimens of this age I am indebted to Professor Huxley.

[17] If Romiti's observations (Archives für Mikr. Anatom. Vol. IX. p. 200) are correct, then the ordinary view of the Wolffian duct arising in Birds as a solid rod at the outer corner of the protovertebræ will have to be abandoned.

[18] While correcting the proofs of this paper I have come across a memoir of W. Müller (Ueber die Persistenz der Urniere bei Myxine Glutinosa, Jenaische Zeitschrift, Vol. VII. 1873), in which he mentions that in Myxine the upper end of the Wolffian duct communicates by numerous openings with the pleuro-peritoneal cavity; this gives to the suggestion in the text a foundation of fact.

[Pg 112]

VI. A Comparison of the Early Stages in the Development of Vertebrates[19].

With Plate 5.

If the genealogical relationships of animals are to be mainly or largely determined on embryological evidence, it becomes a matter of great importance to know how far evidence of this kind is trustworthy.

The dependence to be placed on it has been generally assumed to be nearly complete. Yet there appears to be no à priori reason why natural selection should not act during the embryonic as well as the adult period of life; and there is no question that during their embryonic existence animals are more susceptible to external forces than after they have become full grown: indeed, an immense mass of evidence could be brought to shew that these forces do act upon embryos, and produce in them great alterations tending to obscure the genealogical inferences to be gathered from their developmental histories. Even the time-honoured layers form to this no exception. In Elasmobranchii, for instance, we find the notochord derived from the hypoblast and the spinal ganglia derived from the involuted epiblast of the neural canal, whilst in the higher vertebrates both of these organs are formed in the mesoblast. Such instances are leading embryologists to recognise the fact that the so-called layers are not quite constant and must not be absolutely depended upon in the determination of homologies. But though it is necessary to recognise the fact that great changes do occur in animals during their embryonic life, it is not necessary to conclude that all embryological evidence is thereby vitiated; but rather it becomes incumbent on us to attempt to determine which embryological features are ancestral and which secondary. For this purpose it is requisite [Pg 113] to ascertain what are the general characters of secondary features and how they are produced. Many vertebrates have in the first stages of their development a number of secondary characters which are due to the presence of food material in the ovum; the present essay is mainly an attempt to indicate how those secondary characters arose and to trace their gradual development. At the same time certain important ancestral characters of the early phases of the development of vertebrates, especially with reference to the formation of the hypoblast and mesoblast, are pointed out and their meaning discussed.

There are three orders of vertebrates of which no mention has been made, viz., the Mammals, the Osseous fishes, and the Reptiles. The first of these have been passed over because the accounts of their development are not sufficiently satisfactory, though as far as can be gathered from Bischoff's account of the dog and rabbit there would be no difficulty in shewing their relations with other vertebrates.

We also require further investigations on Osseous fishes, but it seems probable that they develop in nearly the same manner as the Elasmobranchii.

With reference to Reptiles we have no satisfactory investigations.

* * * * *

Amphioxus is the vertebrate whose mode of development in its earliest stages is simplest, and the modes of development of other vertebrates are to be looked upon as modifications of this due to the presence of food material in their ova. It is not necessary to conclude from this that Amphioxus was the ancestor of our present vertebrates, but merely that the earliest stages of development of this vertebrate ancestor were similar to those of Amphioxus.

The ovum of Amphioxus contains very little food material and its segmentation is quite uniform. The result of segmentation is a vesicle whose wall is formed of a single layer of cells. These are all of the same character, and the cavity of the vesicle called the segmentation cavity is of considerable size. A section of the embryo, as we may now call the ovum, is represented in Plate 5, fig. A I.

[Pg 114] The first change which occurs is the pushing in of one half of the wall of the vesicle towards the opposite half. At the same time by the narrowing of its mouth the hollow hemisphere so formed becomes again a vesicle[20].

Owing to its mode of formation the wall of this secondary vesicle is composed of two layers which are only separated by a narrow space, the remnant of the segmentation cavity.

Two of the stages in the formation of the secondary vesicle by this process of involution are shewn in Plate X, fig. A II , and A III. In the second of these the general growth has been very considerable, rendering the whole animal much larger than before. The cavity of this vesicle, A III , is that of the commencing alimentary canal whose final form is due to changes of shape undergone by this primitive cavity. The inner wall of the vesicle becomes converted into the wall of the alimentary canal or hypoblast, and also into part or the whole of the mesoblast.

During the involution the cells which are being involuted undergo a change of form, and before the completion of the process have acquired a completely different character to the cells forming the external wall of the secondary vesicle or epiblast. This change of character in the cells is already well marked in fig. A II. It is of great importance, since we shall find that some of the departures from this simple mode of development, which characterise other vertebrates, are in part due to the distinction between the hypoblast and epiblast cells appearing during segmentation, and not subsequently as in Amphioxus during the involution of the hypoblast.

Kowalevsky (Entwicklungsgeschichte des Amphioxus) originally believed that the narrow mouth of the vesicle (according to Mr Lankester's terminology blastopore) became the anus of the adult. He has since, and certainly correctly, given up this view. The opening of the involution becomes closed up and the adult anus is no doubt formed as in all other vertebrates by a pushing in from the exterior, though it probably corresponds in position very closely with the point of closing up of the original involution.

[Pg 115] The mode of formation of the mesoblast is not certainly known in Amphioxus; we shall find, however, that for all other vertebrates it arises from the cells which are homologous with the involuted cells of this animal.

Since food material is a term which will be very often employed, it will be well to explain exactly the sense in which it will be used. It will be used only with reference to those passive highly refractive particles which are found embedded in most ova.

In some eggs, of which the hen's egg may be taken as a familiar example, the yolk-spherules or food material form the larger portion of the ovum, and a distinction is frequently made between the germinal disc and the yolk.

This distinction is, however, apt to lead to a misconception of the true nature of the egg. There are strong grounds for believing that the so-called yolk, equally with the germinal disc, is composed of an active protoplasmic basis endowed with the power of growth, in which passive yolk-spherules are embedded; but that the part ordinarily called the yolk contains such a preponderating amount of yolk-spherules that the active basis escapes detection, and does not exhibit the same power of growth as the germinal disc.

With the exception of mammals, whose development requires to be more completely investigated, Amphioxus is as far as we know the only vertebrate whose ovum does not contain a large amount of food material.

In none of these (vertebrate) yolk-containing ova is the food material distributed uniformly. It is always concentrated much more at one pole than at the other, and the pole at which it is most concentrated may be conveniently called the lower pole of the egg.

In eggs in which the distribution of food material is not uniform segmentation does not take place with equal rapidity through all parts of the egg, but its rapidity is, roughly speaking, inversely proportional to the quantity of food material.

When the quantity of food material in a part of the egg becomes very great, segmentation does not occur at all; and even in those cases where the quantity of food yolk is not too great to prevent segmentation the resulting segmentation [Pg 116] spheres are much larger than where the yolk-granules are more sparsely scattered.

The Frog is the vertebrate whose development comes nearest to that of Amphioxus, as far as the points we are at present considering are concerned. But it will perhaps facilitate the understanding of their relations shortly to explain the diagrammatic sections which I have given of an animal supposed to be intermediate in its development between the Frog and Amphioxus. Plate 5, fig. B I, represents a longitudinal section of this hypothetical egg at the close of segmentation. The lower pole, coloured yellow, represents the part containing more yolk material, and the upper pole, coloured blue, that with less yolk. Owing to the presence of this yolk the lower pole even at the close of segmentation is composed of cells of a different character to those of the upper pole. In this respect this egg can already be distinguished from that of Amphioxus, in which no such difference between the two poles is apparent at the corresponding period (Plate 5, fig. A I).

The segmentation cavity in this ovum is not quite so large proportionately as in Amphioxus, and the encroachment upon it is due to the larger bulk of the lower pole of the egg. In fig. B II the involution of the lower pole has already commenced; this involution is (1) not quite symmetrical, and (2) on the ventral side (the left side) the epiblast cells forming the upper part of the egg are growing round the cells of the lower pole of the egg or lower layer cells. Both of these peculiarities are founded upon what happens in the Frog and the Selachian, but it is to be noticed that the change from the lower layer cells being involuted towards the epiblast cells, to the epiblast cells growing round the lower layer cells, is a necessary consequence of the increased bulk of the latter.

In this involution not only are the cells of the lower pole pushed on, but also some of those of the upper or yellow portion; so that in this as in all other cases the true distinction between the epiblast and hypoblast does not appear till the involution to form the latter is completed. In the next stage, B III, the involution has become nearly completed and the opening to the exterior or blastopore quite constricted.

The segmentation cavity has been entirely obliterated, as [Pg 117] would have been found to be the case with Amphioxus had the stage a little older than that on Plate 5, A III, been represented. The cavity marked (al), as was the case with Amphioxus, is that of the alimentary canal.

The similarities between the mode of formation of the hypoblast and alimentary canal in this animal and in Amphioxus are so striking and the differences between the two cases so slight that no further elucidation is required. One or two points need to be spoken of in order to illustrate what occurs in the Frog. When the involution to form the alimentary canal occurs, certain of the lower layer cells (marked hy) become distinguished from the remainder of the lower layer cells as a separate layer and form the hypoblast which lines the alimentary canal. It is to be noticed that the cells which form the ventral epithelium of the alimentary canal are not so soon to be distinguished from the other lower layer cells as those which form its dorsal epithelium. This is probably a consequence of the more active growth, indicated by the asymmetry of the involution, on the dorsal side, and is a fact with important bearings in the ova with more food material. The cells marked m and coloured red also become distinguished as a separate layer from the remainder of the hypoblast and form the mesoblast. The remainder of the lower layer cells form a mass equivalent to the yolk-sac of many vertebrates, and are not converted directly into the tissues of the animal.

Another point to be noticed is the different relation of epiblast cells to the hypoblast cells at the upper and lower side of the mouth of the involution. Above it, on its dorsal side, the epiblast and hypoblast are continuous with one another. On its ventral side they are primitively not so continuous. This is due to the epiblast, as was before mentioned, growing round the lower layer cells on the ventral side, vide B II, and merely remaining continuous with them on the dorsal. The importance of these two points will appear when we come to speak of other vertebrates.

The next animal whose development it is necessary to speak of is the Frog, and its differences from the mode of development are quite easy to follow and interpret. Segmentation is again not uniform, and results in the formation of an upper layer of [Pg 118] smaller cells and a lower one of larger; in the centre is a segmentation cavity. The stage at the close of segmentation is represented in C I. From the diagram it is apparent that the lower layer cells occupy a larger bulk than they did in the previous animal (Plate 5, B I), and tend to encroach still more upon the segmentation cavity, otherwise the differences between the two are unimportant. There are, however, two points to be noted. In the first place, although the cells of the upper pole are distinguished in the diagrams from the lower by their colour, it is not possible at this stage to say what will become epiblast and what hypoblast. In the second place the cells of the upper pole or epiblast consist of two layers—an outer called the epidermic layer and an inner called the nervous. In the previous cases the epiblast consisted of a single layer of cells. The presence of these two layers is due to a distinction which, arising in most other vertebrates late, in the Frog arises early. In most other vertebrates in the later stages of development the epiblast consists of an outer layer of passive and an inner of active cells. In the Frog and other Batrachians these two layers become distinguished at the commencement of development.

In the next stage (C II) we find that the involution to form the alimentary canal has commenced (al), but that it is of a very different character to the involution in the previous case. It consists in the growing inwards of a number of cells from the point x (C I) towards the segmentation cavity. The cells which grow in this way are partly the blue cells and partly the smaller yellow ones. At first this involuted layer of cells is only separated by a slit from the remainder of the lower layer cells; but by the stage represented in C II this has widened into an elongated cavity (al). In its formation this involution pushes backwards the segmentation cavity, which finally disappears in the stage C III. The point x remains practically stationary, but by the general growth of the epiblast, mesoblast and hypoblast, becomes further removed from the segmentation cavity in C II than in C I. On the opposite side of the embryo to that at which the involution occurs the epiblast cells as before, grow round the lower layer cells. The commencement of this is already apparent in C I, and in C II the process is nearly completed, [Pg 119] though there is still a small mass of yolk filling up the blastopore. The features of this involution are in the main exaggerations of what was supposed to occur in the previous animal. The asymmetry of the involution is so great that it is completely one-sided and results, in the first instance, in a mere slit; and the whole process of enclosing the yolk by epiblast is effected by the epiblast cells on the side of the egg opposite to the involution.

The true mesoblast and hypoblast are formed precisely as in the previous case. The involuted cells become separated into two layers, one forming the dorsal epithelium of the alimentary canal, and a layer between this and the epiblast forming the mesoblast. There is also a layer of mesoblast accompanying the epiblast which encloses the yolk, which is derived from the smaller yellow cells at y (C I). The edge of this mesoblast, m´, forms a thickened ridge, a feature which persists in other vertebrates.

It is a point of some importance for understanding the relation between the mode of formation of the alimentary canal in the Frog and other vertebrates to notice that on the ventral surface the cells which are to form the epithelium of the alimentary canal become distinguished as such very much later than do those to form its dorsal epithelium, and are derived not from the involuted cells but from the primitive large yolk-cells. It is indeed probable that only a very small portion of epithelium of the ventral wall of the mid-gut is in the end derived from these larger yolk-cells. The remainder of the yolk-cells (C III, and C II, yk) form the yolk mass and do not become directly formed into the tissues of the animal.

In the last stage I have represented for the frog, C III , there are several features to be noticed.

The direct connection at their hind-ends between the cavities of the neural and alimentary canals is the most important of these. This is a result of the previous continuity of the epiblast and hypoblast at the point x, and is a feature almost certainly found in Amphioxus, but which I will speak of more fully in my account of the Selachian's development. The opening of the blastopore called the anus of Rusconi is now quite narrowed, it does not become the anus of the adult. It may be noticed that at the front end of the embryo the primitive dorsal [Pg 120] epithelium of the alimentary canal is growing in such a way as to form the epithelium both of the dorsal and ventral surfaces of the fore-gut.

In spite of various features rendering the development of the Frog more difficult of comprehension than that of most other vertebrates, it is easy to see that the step between it and Amphioxus is not a very great one, and will very likely be bridged over at some future time, when our knowledge of the development of other forms becomes greater.

From the Frog to the Selachian is a considerable step, but I have again hypothetically sketched a type intermediate between them whose development agrees in some important points with that of Pelobates fuscus as described by Bambeke. The points of agreement, though not obvious at first sight, I shall point out in the course of my description.

The first stage (D I), at the close of segmentation, deserves careful attention. The segmentation cavity by the increase of the food yolk is very much diminished in size, and, what is still more important, has as it were sunk down so as to be completely within the lower layer cells. The roof of the segmentation cavity is thus formed of epiblast and lower layer cells, a feature which Bambeke finds in Pelobates fuscus and which is certainly found in the Selachians. In the Frog we found that the segmentation cavity began to be encroached on by the lower layer cells, and from this it is only a small step to find these cells creeping still further up and forming the roof of the cavity. In the lower layer cells themselves we find an important new feature, viz. that during segmentation they become divided in two distinct parts—one of these where the segments owing to the presence of much food yolk are very large, and the other where the segments are much smaller.

The separation between these two is rather sharp. Even this separation was foreshadowed in the Frog's egg, in which a number of lower layer cells were much smaller and more active at the two sides of the segmentation cavity than elsewhere. The segmentation cavity at first lies completely within the region of the small spheres. The larger cells serve almost entirely as food yolk. The epiblast, as is normal with vertebrates, consists of a single layer of columnar cells.

[Pg 121] In the next stage (D II) the formation of the alimentary canal (al) has commenced, but it is to be observed that there is in this case no true involution.

As an accompaniment to the encroachment upon the segmentation cavity, which was a feature of the last stage, the cells to form the walls of the alimentary canal have come to occupy their final position during segmentation and without the intermediation of an involution, and traces only of the involution, are to be found in (1) a split in the lower layer cells which passes along the line separating the small and the large lower layer cells; and (2) in the epiblast becoming continuous with the hypoblast on the dorsal side of the mouth of this split. It is even possible that at this point a few cells (though certainly only a very small number) of those marked blue in D I become involuted. This point in this, as in all other cases, is the tail end of the embryo. The other features of this stage are as follows:—(1) The segmentation cavity has become smaller and less conspicuous than it was. (2) The epiblast cells have begun to grow round the yolk even in a more conspicuous manner than they did in the Frog, and are accompanied by a layer of mesoblast cells which again becomes thickened at its edge. The mesoblast cells in the region of the body are formed in the same way as before, viz. by the separation of a layer to form the epithelium of the alimentary canal, the other cells remaining as mesoblast; and as in the Frog, or in a more conspicuous manner, we find that the dorsal surface only of the alimentary cavity has a wall formed of a distinct layer of cells, but on the ventral side the cavity is at first closed in by the large spheres of the yolk only. The formation of the alimentary canal by a split and not by an involution is exactly what Bambeke finds in Pelobates.

The next stage, D III, is about an equivalent age to C III in the Frog. It exhibits the same connection between the neural and the alimentary canals as was found there.

The alimentary canal is beginning to become closed in below, and this occurs near the two ends earlier than in the middle. The cells to form the ventral wall are derived from the large yolk-cells. The non-formation of the ventral wall of the alimentary canal so soon in the middle as at the ends is an [Pg 122] early trace of the umbilical canal found in Birds and Selachians, by which the alimentary tract is placed in communication with the yolk-sac. The segmentation cavity has by this stage completely vanished, and the epiblast with its accompanying mesoblast has spread completely round the yolk material so as to form the ventral wall of the body.

Though in some points this manner of development may seem to differ from that of the Frog, there is really a fundamental agreement between the two, and between this mode of development and that of the Selachians we shall find the agreement to be very close.

After segmentation we find that the egg of a Selachian consists of two parts—one of these called the germinal disc or blastoderm, and the other the yolk. The former of these corresponds with the epiblast and the part of the lower pole composed of smaller segments in the last-described egg, and the latter to the larger segments of the lower pole. This latter division, owing to the quantity of yolk which it contains, has not undergone segmentation, but its homology with the larger segments of the previous eggs is proved (1) by its containing a number of nuclei (E I, n), which become the nuclei of true cells and enter the blastoderm, and (2) by the presence in it of a number of lines forming a network similar to that of many cells. The segmentation cavity, as before, lies completely within the lower layer cells.

The next stage, E II, is almost precisely similar to the second stage of the last egg. As there, the primitive involution is merely represented by a split separating the yolk and the germinal disc, and on the dorsal side alone is there a true cellular wall for this split, and at the dorsal mouth of the split the alimentary epithelium becomes continuous with the epiblast.

The segmentation cavity has become diminished, and round the yolk the epiblast, accompanied by a layer of mesoblast, is commencing to grow. In this growth all parts of the blastoderm take a share except that part where the epiblast and hypoblast are continuous. This manner of growth is precisely what occurs in the Frog, though there it is not so easily made out; and not all the investigators who have studied the Frog have understood the exact meaning of the appearances they have [Pg 123] seen and drawn. This similarity of relation of the epiblast to the yolk in the two cases is a further confirmation of the identity of the Selachian's yolk with the large yolk-spheres of the previous eggs.

The next stage, E III, is in many ways identical with the corresponding stage in the last-described egg, and in the same way as in that case the neural and alimentary canals are placed in communication with each other.

The mode in which this occurs will be easily gathered from a comparison of E II and E III. It is the same for the Selachians and Batrachians. The neural canal (nc) is by the stage figured E III , completely formed in the way so well known in the Bird, and between the roof of the canal and the external epiblast a layer of mesoblast has already grown in. The floor of the neural canal is the same layer marked ep in E II , and therefore remains continuous with the hypoblast at x; and when by a simultaneous process the roof of the neural canal and the ventral wall of the alimentary become formed by the folding over of one continuous layer (the epiblast and hypoblast continuous at the point x), the two canals, viz. the neural and alimentary, are necessarily placed in communication at their hind-ends, as is seen in the diagram.

There are several important points of difference between E III and D III. In the first place, owing to the larger size of the yolk mass in E III , the epiblast, accompanied by mesoblast, has not proceeded nearly so far round it as in the previous case. It is also worth notice that at the right as well as at the left end of the germinal disc the epiblast is commencing to grow round the yolk. The yolk has, however, become surrounded to a much smaller extent on the right hand than on the left. Since, in the earlier stage, the epiblast became continuous with the hypoblast at x, it is not from sections obvious how this occurs. I have therefore appended a diagram to explain it (E´). The blastoderm rests like a disc on the yolk and grows over it on all sides, except at the point where the epiblast and hypoblast are continuous (x). This point becomes as it were left in a bay. Next the two sides of the bay coalesce, the bay becomes obliterated, and the effect produced is exactly as if the blastoderm had grown round the yolk at the point x (corresponding with the [Pg 124] tail of the embryo) as well as everywhere else. It thus comes about that the final point where the various parts of the blastoderm meet and completely enclose the yolk mass does not correspond with the anus of Rusconi of the Frog, but is at some little distance from the hind-end of the embryo. In other words, the position of the blastopore in the Selachian is not the same as in the Frog.

Another point deserving attention is the formation of the ventral wall of the alimentary canal. This takes place in two ways—partly by a folding-in at the sides and end, and partly from cells formed around the nuclei (n) in the yolk. From these a large portion of the ventral wall of the mid-gut is formed.

The folding-in of the sheet of hypoblast to assist in the closing-in of the ventral wall of the alimentary canal is a consequence of the flattened form of the original alimentary slit which is far too wide to form the cavity of the final canal. In the Bird whose development must next be considered this folding-in is a still more prominent feature in the formation of the alimentary canal. As in the last case, the alimentary canal is widely open in the middle to the yolk at the time when its two ends are closed below and shut off from it; still later this opening becomes very narrow and forms the duct of the so-called umbilical cord which places the yolk-sac in communication with the alimentary canal. As the young animal becomes larger the yolk-sac ceases to communicate directly with the alimentary canal, and is carried about by it for some time as an appendage and only at a later period shrivels up.

The mesoblast is formed in a somewhat different way in the Sharks than in other vertebrates. It becomes split off from the hypoblast, not in the form of a single sheet as in other vertebrates, but as two lateral sheets, one on each side of the middle line and separated from one another by a considerable interval; whilst the notochord is derived not as in other vertebrates from the mesoblast, but from the hypoblast (vide F. M. Balfour, Development of Selachians[21], Journal of Microscopical Science, Oct., 1874).

[Pg 125] Between the Selachians and the Aves there is a considerable gulf, which it is more difficult satisfactorily to bridge over than in the previous cases; owing to this I have not attempted to give any intermediate stage between them.

The first stage of the Bird (F I) is very similar in many respects to the corresponding stage in the Selachian. The segmentation cavity is, however, a less well-defined formation, and it may even be doubted whether a true segmentation cavity, homologous with the segmentation cavity in the previously described eggs, is present. On the floor of the cavity which is formed by the yolk are a few larger cells known as formative cells which, according to Götte's observations, are derived from the yolk, in a somewhat similar manner to the cells which were formed around the nuclei in the Selachian egg, and which helped to form the ventral wall of the alimentary canal. Another point to be noticed is that the segmentation cavity occupies a central position, and not one to the side as in the Selachian.

The yolk is proportionately quite as large as in the Selachian's egg, but, as in that case, there can be little or no doubt of its being homologous with the largest of the segmentation spheres of the previous eggs. It does not undergo segmentation. The epiblast is composed of columnar cells, and extends a short way beyond the edge of the lower layer cells.

In the next stage the more important departures from the previous type of development become visible.

The epiblast spreads uniformly over the yolk-sac and not on the one side only as in the former eggs.

This is due to the embryo (indicated in F II by a thickening of the cells) lying in the centre and not at the edge of the blastoderm. A necessary consequence of this is, that the epiblast does not, as in the previous cases, become continuous with the hypoblast at the tail end of the embryo. This continuity, being of no functional importance, could easily be dispensed with, and the central position of the embryo may perhaps be explained by supposing the process, by which in the Selachian egg the blastopore ceases to correspond in position with the opening of the alimentary slit or anus of Rusconi (vide E´), to occur quite early during segmentation instead of at a late period of development. [Pg 126] For the possibility of such a change in the date of formation, the early appearance of the nervous and epidermic layers in the Frog affords a parallel.

The epiblast in its growth round the yolk is only partially accompanied by mesoblast, which, however, is thickened at its extreme edge as in the Frog. Owing to the epiblast not becoming continuous with the hypoblast at the tail end of the embryo, the alimentary slit is not open to the exterior. The hypoblast is formed by some of the lower layer cells becoming distinguished as a separate layer; the remainder of the lower layer cells become the mesoblast.

The formation of the mesoblast and hypoblast out of the lower layer cells has been accepted for the Bird by most observers, but has been disputed by several, and recently by Kölliker. These have supposed that the mesoblast is derived from the epiblast. I feel convinced that these observers are in the wrong, and that the mesoblast is genuinely derived from the lower layer cells.

The greater portion of the alimentary cavity consists of the original segmentation cavity (vide diagrams). This feature of the segmentation cavity of Birds sharply distinguishes it from any segmentation cavity of other eggs, and renders it very doubtful whether the similarly named cavities of the Bird and of other vertebrates are homologous. On the floor of the cavity are still to be seen some of the formative cells, but observers have not hitherto found that they take any share in forming the ventral wall of the alimentary canal.

The features of the next stage are the necessary consequences of those of the last.

The ventral wall of the alimentary canal is entirely formed by a folding-in of the sheet of hypoblast.

The more rapid folding-in at the head still indicates the previous more vigorous growth there, otherwise there is very little difference between the forms of the fold at the head and tail. The alimentary canal does not of course, at this or any period, communicate with the neural tube, since the epiblast and hypoblast are never continuous. The other features, such as the growth of the epiblast round the yolk-sac, are merely continuations of what took place in the last stage.

[Pg 127] In the development of a yolk-sac as a distinct appendage, and its absorption within the body, at a later period, the bird fundamentally resembles the Dog-fish.

Although there are some difficulties in deriving the type of development exhibited by the Bird directly from that of the Selachian, it is not very difficult to do so directly from Amphioxus. Were the alimentary involution to remain symmetrical as in Amphioxus, and the yolk-containing part of the egg to assume the proportions it does in the Bird, we should obtain a mode of development which would not be very dissimilar to that of the Bird. The epiblast would necessarily overgrow the yolk uniformly on all sides and not in the unsymmetrical fashion of the Selachian egg. A confirmation of this view might perhaps be sought for in the complete difference between the types of circulation of the yolk-sac in Birds and Selachians; but this is not so important as might at first sight appear, since it is not from the Selachian egg but from some Batrachian that it would be necessary to derive the Reptiles' and Birds' eggs.

If this view of the Bird's egg be correct, we are compelled to suppose that the line of ancestors of Birds and Reptiles did not include amongst them the Selachians and the Batrachians, or at any rate Selachians and Batrachians which develop on the type we now find.

The careful investigation of the development of some Reptiles might very probably throw light upon this important point. In the meantime it is better to assume that the type of development of Birds is to be derived from that of the Frog and Selachians.

Summary.—If the views expressed in this paper are correct, all the modes of development found in the higher vertebrates are to be looked upon as modifications of that of Amphioxus. It is, however, rather an interesting question whether it is possible to suppose that the original type was not that of Amphioxus, but of some other animal, say, for instance, that of the Frog, and that this varied in two directions,—on the one hand towards Amphioxus, in the reverse direction to the course of variation presupposed in the text; and on the other hand in the direction towards the Selachians as before.

The answer to this question must in my opinion be in the [Pg 128] negative. It is quite easy to conceive the food material of the Frog's egg completely vanishing, but although this would entail simplifications of development and possibly even make segmentation uniform, there would, as far as I can see, be no cause why the essential features of difference between the Frog's mode of development and that of Amphioxus should change. The asymmetrical and slit-like form of involution on the one side and the growth of the epiblast over the mesoblast on the other side, both characteristics of the present Frog's egg, would still be features in the development of the simplified egg.

In the Mammal's egg we probably have an example of a Reptile's egg simplified by the disappearance of the food material; and when we know more of Mammalian embryology it will be very interesting to trace out the exact manner in which this simplification has affected the development. It is also probable that the eggs of Osseous fish are fundamentally simplified Selachian eggs; in which case we already know that the diminution of food material has affected but very slightly the fundamental features of development.

One common feature which appears prominently in reviewing the embryology of vertebrates as a whole is the derivation of the mesoblast from the hypoblast; in other words, we find that it is from the layer corresponding to that which becomes involuted in Amphioxus so as to line the alimentary cavity that the mesoblast is split off.

That neither the hypoblast or mesoblast can in any sense be said to be derived from the epiblast is perfectly clear. When the egg of Amphioxus is in the blastosphere stage we cannot speak of either an epiblast or hypoblast. It is not till the involution or what is equivalent has occurred, converting the single-walled vesicle into a double-walled one, that we can speak of these two layers. It might seem scarcely necessary to insist upon this point, so clear is it without explanation, were it not that certain embryologists have made a confusion about it.

The derivation of the mesoblast from the hypoblast is the more interesting, since it is not confined to the vertebrates, but has a very wide extension amongst the invertebrates. In the cases (whose importance has been recently insisted upon by Professor Huxley), of the Asteroids, the Echinoids, Sagitta, and [Pg 129] others, in which the body-cavity arises as an outgrowth of the alimentary canal and the somatopleure and splanchnopleure are formed from that outgrowth, it is clear without further remark that the mesoblast is derived from the hypoblast. For the Echinoderms in which the water-vascular system and muscular system arise as a solid outgrowth of the wall of the alimentary canal there can also be no question as to the derivation of the mesoblast from the hypoblast.

Amongst other worms, in addition to Sagitta, the investigations of Kowalevsky seem to shew that in Lumbricus the mesoblast is derived from the hypoblast.

Amongst Crustaceans, Bobretsky's[22] observations on Oniscus (Zeitschrift für wiss. Zoologie, 1874) lead to the same conclusion.

In Insects Kowalevsky's observations lead to the conclusion that mesoblast and hypoblast arise from a common mass of cells; Ulianin's observations bring out the same result for the abnormal Poduridæ, and Metschnikoff's observations shew that this also holds for Myriapods.

In Molluscs the point is not so clear.

In Tunicates, even if we are not to include them amongst vertebrates[23], the derivation of mesoblast from hypoblast is without doubt.

Without going further into details it is quite clear that the derivation of the mesoblast from the hypoblast is very general amongst invertebrates.

It will hardly be disputed that primitively the muscular system of the body-wall could not have been derived from the layer of cells which lines the alimentary canal. We see indeed in Hydra and the Hydrozoa that in its primitive differentiation, as could have been anticipated beforehand, the muscular system of the body is derived from the epiblast cells. What, then, is the explanation of the widespread derivation of the mesoblast, including the muscular system of the body, from the hypoblast?

[Pg 130] The explanation of it may, I think, possibly be found, and at all events the suggestion seems to me sufficiently plausible to be worth making, in the fact that in many cases, and probably this applies to the ancestors of the vertebrates, the body-cavity was primitively a part of the alimentary.

Mr Lankester, who has already entered into this line of speculation, even suggests (Q. J. of Micr. Science, April, 1875) that this applies to all higher animals. It might then be supposed that the muscular system of part of the alimentary canal took the place of the primitive muscular system of the body; so that the whole muscular system of higher animals would be primitively part of the muscular system of the digestive tract.

I put this forward merely as a suggestion, in the truth of which I feel no confidence, but which may perhaps induce embryologists to turn their attention to the point. If we accept it for the moment, the supplanting of the body muscular system by that of the digestive tract may hypothetically be supposed to have occurred in the following way.

When the diverticulum or rather paired diverticula were given off from the alimentary canal they would naturally become attached to the body-wall, and any contractions of their intrinsic muscles would tend to cause movements in the body-wall. So far there is no difficulty, but there is a physiological difficulty in explaining how it can have happened that this secondary muscular system can have supplanted the original muscular system of the body.

The following suggestions may lessen this difficulty, though perhaps they hardly remove it completely. If we suppose that the animal in which these diverticula appeared had a hard test and was not locomotive, the intrinsic muscular system of the body would naturally completely atrophy. But since the muscular system of the diverticula from the stomach would be required to keep up the movement of the nutritive fluid, it would not atrophy, and were the test subsequently to become soft and the animal locomotive, would naturally form the muscular system of the body. Or even were the animal locomotive in which the diverticula appeared, it is conceivable that the two systems might at first coexist together; that either (1) subsequently [Pg 131] owing to the greater convenience of early development, the two systems might acquire a development from the same mass of cells and those the cells of the inner or hypoblast layer, so that the derivation of the body muscles from the hypoblast would only be apparent and not real, or (2) owing to their being better nourished as they would necessarily be, and to their possibly easier adaptability to some new form of movement of the animal, the muscle-cells of the alimentary canal might become developed exclusively whilst the original muscular system atrophied.

I only hold this view provisionally till some better explanation is given of the cases of Sagitta and the Echinoderms, as well as of the nearly universal derivation of the mesoblast from the hypoblast. The cases of this kind may be due to some merely embryonic changes and have no meaning in reference to the adult condition, but I think that we have no right to assume this till some explanation of the embryonic can be suggested.

For vertebrates, I have shewn that in Selachians the body-cavity at first extends quite to the top of what becomes the muscle plate, so that the line or space separating the two layers of the muscle plate (vide Balfour, 'Development of Elasmobranch Fishes[24],' Quart. Journ. of Micro. Science for Oct., 1874. Plate XV, fig. 11a, 11b, 12a, mp.) is a portion of the original body-cavity. If this is a primitive condition, which is by no means certain, we have a condition which we might expect, in which both the inner and the outer wall of the primitive body-cavity assists in forming the muscular system of the body.

It is very possible that the formation of the mesoblast as two masses, one on each side of the middle line as occurs in Selachians, and which as I pointed out in the paper quoted above also takes place in some worms, is a remnant of the primitive formation of the body-cavity as paired outgrowth of the alimentary canal. This would also explain the fact that in Selachians the body-cavity consists at first of two separate portions, one on each side of the alimentary canal, which only subsequently [Pg 132] become united below and converted into a single cavity (vide loc. cit.[25], Plate XIV, fig. 8b, pp).

In the Echinoderms we find instances where the body-cavity and water-vascular system arise as an outgrowth from the alimentary canal, which subsequently becomes constricted off from the latter (Asteroids and Echinoids), together with other instances (Ophiura, Synapta) where the water-vascular system and body-cavity are only secondarily formed in a solid mass of mesoblast originally split off from the walls of the alimentary canal.

These instances shew us how easily a change of this kind may take place, and remove the difficulty of understanding why in vertebrates the body-cavity never communicates with the alimentary.

The last point which I wish to call attention to is the blastopore or anus of Rusconi.

This is the primitive opening by which the alimentary canal communicates with the exterior, or, in other words, the opening of the alimentary involution. It is a distinctly marked structure in Amphioxus and the Batrachians, and is also found in a less well-marked form in the Selachians; in Birds no trace of it is any longer to be seen. In all those vertebrates in which it is present, it closes up and does not become the anus of the adult. The final anus nevertheless corresponds very closely in position with the anus of Rusconi. Mr Lankester has shewn (Quart. Journ. of Micro. Science for April, 1875) that in invertebrates as well as vertebrates the blastopore almost invariably closes up. It nevertheless corresponds as a rule very nearly in position either with the mouth or with the anus.

If this opening is viewed, as is generally done, as really being the mouth in some cases and the anus in others, it becomes very difficult to believe that the blastopore can in all cases represent the same structure. In a single branch of the animal kingdom it sometimes forms the mouth and sometimes the anus: thus for instance in Lumbricus it is the mouth (according to Kowalevsky), in Palæmon (Bobretzky) the anus. Is it credible that the mouth and anus have become changed, the one for the other?

If, on the other hand, we accept the view that the blastopore [Pg 133] never becomes either the one or the other of these openings, it is, I think, possible to account for its corresponding in position with the mouth in some cases or the anus in others.

That it would soon come to correspond either with the mouth or anus (probably with the earliest formed of these in the embryo), wherever it was primitively situated, follows from the great simplification which would be effected by its doing so. This simplification consists in the greater facility with which the fresh opening of either mouth or anus could be made where the epiblast and hypoblast were in continuity than elsewhere. Even a change of correspondence from the position of the mouth to that of the anus or vice versa could occur. The mode in which this might happen is exemplified by the case of the Selachians. I pointed out in the course of this paper how the final point of envelopment of the yolk became altered in Selachians so as to cease to correspond with the anus of Rusconi; in other words, how the position of the blastopore became changed. In such a case, if the yolk material again became diminished, the blastopore would correspond in position with neither mouth nor anus, and the causes which made it correspond in position with the anus before, would again operate, and make it correspond in position perhaps with the mouth. Thus the blastopore might absolutely cease to correspond in position with the anus and come to correspond in position with the mouth.

It is hardly possible to help believing that the blastopore primitively represented a mouth. It may perhaps have lost this function owing to an increase of food yolk in the ovum preventing its being possible for the blastopore to develop directly into a mouth, and necessitating the formation of a fresh mouth. If such were the case, there would be no reason why the blastopore should ever again serve functionally as a mouth in the descendants of the animal which developed this fresh mouth.

[Pg 134] EXPLANATION OF PLATE 5.

COMPLETE LIST OF REFERENCES.

al. Cavity of alimentary canal. bl. Blastoderm. ch. Notochord. ep. Epiblast. em. Embryo. f. Formative cells. hy. Hypoblast. ll. Lower layer cells. m. Mesoblast. n. Nuclei of yolk of Selachian egg. nc. Neural canal. sg. Segmentation cavity. x. Point where epiblast and hypoblast are continuous at the mouth of the alimentary involution. This point is always situated at the tail end of the embryo. yk. Yolk.

Epiblast is coloured blue, mesoblast red, and hypoblast yellow. The lower layer cells before their separation into hypoblast and mesoblast are also coloured green.

A I, A II, A III. Diagrammatic sections of Amphioxus in its early stages (founded upon Kowalevsky's observations).

B I, B II, B III. Diagrammatic longitudinal sections of an hypothetical animal, intermediate between Amphioxus and Batrachians, in its early stages.

C I, C II, C III. Diagrammatic longitudinal sections of Bombinator igneus in its early stages (founded upon Götte's observations). in C III the neural canal is completed, which was not the case in B III. The epiblast in C III has been diagrammatically represented as a single layer.

D I, D II, D III. Diagrammatic longitudinal sections of an animal, intermediate between Batrachians and Selachians, in its early stages.

E I, E II, E III. Diagrammatic longitudinal sections of a Selachian in its early stages.

E´. Surface view of the yolk of a Selachian's egg to shew the manner in which it is enclosed by the Blastoderm. The yolk is represented yellow and the Blastoderm blue.

F I, F II, F III. Diagrammatic longitudinal sections of a Bird in its early stages.

[19] From the Quarterly Journal of Microscopical Science, Vol. XV. 1875.

[20] I have been able to make at Naples observations which confirm the account of the invagination of Amphioxus as given by Kowalevsky, though my observations are not nearly so complete as those of the Russian naturalist.

[21] Paper No. V, p. 82 et seq. in this edition.

[22] He says, p. 182: Bevor aber die Hälfte der Eioberfläche von den Embryonalzellen bedeckt ist, kommt die erste gemeinsame Anlage des mittleren und unteren Keimblattes zum Vorschein.

[23] Anton Dohrn, Der Ursprung des Wirbelthieres. Leipzig, 1875.

[24] Paper No. V, p. 60 et seq. of this edition, pl. 4, figs. 11a, 11b, 12a, mp.

[25] Pl. 3 of this edition, fig. 8b, pp.

[Pg 135]

VII. On the Origin and History of the Urinogenital Organs of Vertebrates[26].

Recent discoveries[27] as to the mode of development and anatomy of the urinogenital system of Selachians, Amphibians, and Cyclostome fishes, have greatly increased our knowledge of this system of organs, and have rendered more possible a comparison of the types on which it is formed in the various orders of vertebrates.

[Pg 136] The following paper is an attempt to give a consecutive history of the origin of this system of organs in vertebrates and of the changes which it has undergone in the different orders.

For this purpose I have not made use of my own observations alone, but have had recourse to all the Memoirs with which I am acquainted, and to which I have access. I have commenced my account with the Selachians, both because my own investigations have been directed almost entirely to them, and because their urinogenital organs are, to my mind, the most convenient for comparison both with the more complicated and with the simpler types.

On many points the views put forward in this paper will be found to differ from those which I expressed in my paper (loc. cit.) which give an account of my original[28] discovery of the segmental organs of Selachians, but the differences, with the exception of one important error as to the origin of the Wolffian duct, are rather fresh developments of my previous views from the consideration of fresh facts, than radical changes in them.

* * * * *

In Selachian embryos an intermediate cell-mass, or middle plate of mesoblast is formed, as in birds, from a partial fusion of the somatic and splanchnic layers of the mesoblast at the outer border of the protovertebræ. From this cell-mass the whole of the urinogenital system is developed.

At about the time when three visceral clefts have appeared, there arises from the intermediate cell-mass, opposite the fifth protovertebra, a solid knob, from which a column of cells grows backwards to opposite the position of the future anus (Fig. 1. pd.).

Fig. 1. Two sections of a Pristiurus Embryo with three visceral clefts.

The sections are to shew the development of the segmental duct (pd) or primitive duct of the kidneys. In A (the anterior of the two sections) this appears as a solid knob projecting towards the epiblast. In B is seen a section of the column which has grown backwards from the knob in A.

spn. rudiment of a spinal nerve; mc. medullary canal; ch. notochord; X. string of cells below the notochord; mp. muscle-plate; mp´. specially developed portion of muscle-plate; ao. dorsal aorta; pd. segmental duct; so. somatopleura; sp. splanchnopleura; pp. pleuro-peritoneal or body-cavity; ep. epiblast; al. alimentary canal.

This knob projects outwards toward the epiblast, and the column lies at first between the mesoblast and epiblast. The knob and column do not long remain solid. The knob becoming hollow acquires a wide opening into the pleuro-peritoneal or body-cavity, and the column a lumen; so that by the time that five visceral clefts have appeared, the two together form a [Pg 137] duct closed behind, but communicating in front by a wide opening with the pleuro-peritoneal cavity.

Before these changes are accomplished, a series of solid[29] outgrowths of elements of the 'intermediate cell-mass' appear at the uppermost corner of the body-cavity. These soon become hollow and appear as involutions from the body-cavity, curling round the inner and dorsal side of the previously formed duct.

One involution of this kind makes its appearance for each protovertebra, and the first belongs to the protovertebra immediately behind the anterior end of the duct whose development has just been described. In Pristiurus there are in all 29 of these at this period. The last two or three arise from that portion of the body-cavity, which at this stage still exists behind the anus. The first-formed duct and the subsequent involutions are the rudiments of the whole of the urinary system. [Pg 138] The duct is the primitive duct of the kidney[30]; I shall call it in future the segmental duct; and the involutions are the commencements of the segmental tubes which constitute the body of the kidney. I shall call them in future segmental tubes.

Soon after their formation the segmental tubes become convoluted, and their blind ends become connected with the segmental duct of the kidney. At the same time, or rather before this, the blind posterior termination of each of the segmental ducts of the kidneys unites with and opens into one of the horns of the cloaca. At this period the condition of affairs is represented in Fig. 2.

Fig. 2. Diagram of the primitive condition of the Kidney in a Selachian Embryo.

pd. segmental duct. It opens at o into the body-cavity and at its other extremity into the cloaca; x. line along which the division appears which separates the segmental duct into the Wolffian duct above and the Müllerian duct below; st. segmental tubes. They open at one end into the body-cavity, and at the other into the segmental duct.

There is at pd, the segmental duct of the kidneys, opening in front (o) into the body-cavity, and behind into the cloaca, and there are a series of convoluted segmental tubes (st), each opening at one end into the body-cavity, and at the other into the duct (pd).

The next important change which occurs is the longitudinal division of the segmental duct of the kidneys into Müller's duct, or the oviduct, and the duct of the Wolffian bodies or Leydig's duct. The splitting[31] is effected by the growth of a wall of cells [Pg 139] which divides the duct into two parts (fig. 3, wd. and md.). It takes place in such a way that the front end of the segmental duct, anterior to the entrance of the first segmental tube, together with the ventral half of the rest of the duct, is split off from its dorsal half as an independent duct (vide fig. 2, x).

Fig. 3. Transverse section of a Selachian Embryo illustrating the formation of the Wolffian and Müllerian ducts by the longitudinal splitting of the segmental duct.

mc. medullary canal; mp. muscle-plate; ch. notochord; ao. aorta; cav. cardinal vein; st. segmental tube. On the one side the section passes through the opening of a segmental tube into the body-cavity. On the other this opening is represented by dotted lines, and the opening of the segmental tube into the Wolffian duct has been cut through; wd. Wolffian duct; md. Müllerian duct. The Müllerian duct and the Wolffian duct together constitute the primitive segmental duct; gr. The germinal ridge with the thickened germinal epithelium; l. liver; i. intestine with spiral valve.

The dorsal portion also forms an independent duct, and into it the segmental tubes continue to open. Such at least is the [Pg 140] method of splitting for the female—for the male the splitting is according to Professor Semper, of a more partial character, and consists for the most part in the front end of the duct only being separated off from the rest. The result of these changes is the formation in both sexes of a fresh duct which carries off the excretions of the segmental involutions, and which I shall call the Wolffian duct—while in the female there is formed another complete and independent duct, which I shall call the Müllerian duct, or oviduct, and in the male portions only of such a duct.

The next change which takes place is the formation of another duct from the hinder portion of the Wolffian duct, which receives the secretion of the posterior segmental tubes. This secondary duct unites with the primary or Wolffian duct near its termination, and the primary ducts of the two sides unite together to open to the exterior by a common papilla.

Slight modifications of the posterior terminations of these ducts are found in different genera of Selachians (vide Semper, Centralblatt für Med. Wiss. 1874, No. 59), but they are of no fundamental importance.

These constitute the main changes undergone by the segmental duct of the kidneys and the ducts derived from it; but the segmental tubes also undergo important changes. In the majority of Selachians their openings into the body-cavity, or, at any rate, the openings of a large number of them, persist through life; but the investigations of Dr Meyer[32] render it very probable that the small portion of each segmental tube adjoining the opening becomes separated from the rest and becomes converted into a sort of lymph organ, so that the openings of the segmental tubes in the adult merely lead into lymph organs and not into the gland of the kidneys.

These constitute the whole changes undergone in the female, but in the male the open ends of a varying number (according to the species) of the segmental tubes become connected with the testis and, uniting with the testicular follicles, serve to carry away the seminal fluid[33]. The spermatozoa have therefore to [Pg 141] pass through a glandular portion of the kidneys before they enter the Wolffian duct, by which they are finally carried away to the exterior.

In the adult female, then, there are the following parts of the urinogenital system (fig. 4):

(1) The oviduct, or Müller's duct (fig. 4, md.), split off from the segmental duct of the kidneys. Each oviduct opens at its upper end into the body-cavity, and behind the two oviducts have independent communications with the cloaca. The oviducts serve simply to carry to the exterior the ova, and have no communication with the glandular portion of the kidneys.

Fig. 4. Diagram of the arrangement of the Urinogenital Organs in an adult Female Selachian.

md. Müllerian duct; wd. Wolffian duct; st. segmental tubes; d. duct of the posterior segmental tubes; ov. ovary.

(2) The Wolffian ducts (fig. 4, wd.) or the remainder of the segmental ducts of the kidneys. Each Wolffian duct ends blindly in front, and the two unite behind to open by a common papilla into the cloaca.

This duct receives the secretion of the whole anterior end of the kidneys[34], that is to say, of all the anterior segmental tubes.

(3) The secondary duct (fig. 4, d.) belonging to the lower portion of the kidneys opening into the former duct near its termination.

(4) The segmental tubes (fig. 4, st) from whose convolutions and outgrowths the kidney is formed. They may be divided [Pg 142] into two parts, according to the duct by which their secretion is carried off.

In the male the following parts are present:

(1) The Müllerian duct (fig. 5, md.), consisting of a small remnant, attached to the liver, which represents the foremost end of the oviduct of the female.

(2) The Wolffian duct (fig. 5, wd), which precisely corresponds to the Wolffian duct of the female, except that, in addition to functioning as the duct of the anterior part of the kidneys, it also serves to carry away the semen. In the female it is straight, but has in the adult male a very tortuous course (vide fig. 5).

Fig. 5. Diagram of the arrangement of the Urinogenital Organs in an adult male selachian.

md. rudiment of Müllerian duct; wd. Wolffian duct, which also serves as vas deferens; st. segmental tubes. The ends of three of those which in the female open into the body-cavity, have in the male united with the testicular follicles, and serve to carry away the products of the testis; d. duct of the posterior segmental tubes; t. testis.

(3) the duct (fig. 5, d.) of the posterior portion of the kidneys, which has the same relations as in the female.

(4) The segmental tubes (fig. 5, st.). These have the same relations as in the female, except that the most anterior two, three or more, unite with the testicular follicles, and carry away the semen into the Wolffian duct.

* * * * *

The mode of arrangement and the development of these parts suggest a number of considerations.

In the first place it is important to notice that the segmental tubes develop primitively as completely independent [Pg 143] organs[35], one of which appears in each segment. If embryology is in any way a repetition of ancestral history, it necessarily follows that these tubes were primitively independent of each other. Ancestral history, as recorded in development, is often, it is true, abridged; but it is clear that though abridgement might prevent a series of primitively separate organs from appearing as such, yet it would hardly be possible for a primitively compound organ, which always retained this condition, to appear during development as a series of separate ones. These considerations appear to me to prove that the segmented ancestors of vertebrates possessed a series of independent and segmental excretory organs.

Both Professor Semper and myself, on discovering these organs, were led to compare them and state our belief in their identity with the so-called segmental organs of Annelids.

This view has since been fairly generally accepted. The segmental organs of annelids agree with those of vertebrates in opening at one end into the body-cavity, but differ in the fact that each also communicates with the exterior by an independent opening, and that they are never connected with each other.

On the hypothesis of the identity of the vertebrate segmental tubes with the annelid segmental organs, it becomes essential to explain how the external openings of the former may have become lost.

This brings us at once to the origin of the segmental duct of the kidneys, by which the secretion of all the segmental tubes was carried to the exterior, and it appears to me that a right understanding of the vertebrate urinogenital system depends greatly upon a correct view of the origin of this duct. I would venture to repeat the suggestion which I made in my original paper (loc. cit.) that this duct is to be looked upon as the most anterior of the segmental tubes which persist in vertebrates. [Pg 144] In favour of this view are the following anatomical and embryological facts. (1) It develops in nearly the same manner as the other segmental tubes, viz. in Selachians as a solid outgrowth from the intermediate cell-mass, which subsequently becomes hollowed so as to open into the body-cavity: and in Amphibians and Osseous and Cyclostome fishes as a direct involution from the body-cavity. (2) In Amphibians, Cyclostomes and Osseous fishes its upper end develops a glandular portion, by becoming convoluted in a manner similar to the other segmental tubes. This glandular portion is often called either the head-kidney or the primitive kidney. It is only an embryonic structure, but is important as demonstrating the true nature of the primitive duct of the kidneys.

We may suppose that some of the segmental tubes first united, possibly in pairs, and that then by a continuation of this process the whole of them coalesced into a common gland. One external opening sufficed to carry off the entire secretion of the gland, and the other openings therefore atrophied.

This history is represented in the development of the dog-fish in an abbreviated form, by the elongation of the first segmental tube (segmental duct of the kidney) and its junction with each of the posterior segmental tubes. Professor Semper looks upon the primitive duct of the kidneys as a duct which arose independently, and was not derived from metamorphosis of the segmental organs. Against this view I would on the one hand urge the consideration, that it is far easier to conceive of the transformation by change of function (comp. Dohrn, Functionswechsel, Leipzig, 1875) of a segmental organ into a segmental duct, than to understand the physiological cause which should lead, in the presence of so many already formed ducts, to the appearance of a totally new one. By its very nature a duct is a structure which can hardly arise de novo. We must even suppose that the segmental organs of Annelids were themselves transformations of still simpler structures. On the other hand I would point to the development in this very duct amongst Amphibians and Osseous fishes of a glandular portion similar to that of a segmental tube, as an à posteriori proof of its being a metamorphosed segmental tube. The development in insects of a longitudinal tracheal duct by the coalescence of a [Pg 145] series of transverse tracheal tubes affords a parallel to the formation of a duct from the coalescence of a series of segmental tubes.

Though it must be admitted that the loss of the external openings of the segmental organs requires further working out, yet the difficulties involved in their disappearance are not so great as to render it improbable that the vertebrate segmental organs are descended from typical annelidan ones.

The primitive vertebrate condition, then, is probably that of an early stage of Selachian development while there is as yet a segmental duct,—the original foremost segmental tube opening in front into the body-cavity and behind into the cloaca; with which duct all the segmental tubes communicate. Vide Fig. 2.

The next condition is to be looked upon as an indirect result of the segmental duct serving as well for the products of the generative organs as the secretions of the segmental tubes.

As a consequence of this, the segmental duct became split into a ventral portion, which served alone for the ova, and a dorsal portion which received the secretion of the segmental tubes. The lower portion, which we have called the oviduct, in some cases may also have received the semen as well as the ova. This is very possibly the case with Ceratodus (vide Günther, Trans. of Royal Society, 1871), and the majority of Ganoids (Hyrtl, Denkschriften Wien, Vol. VIII.). In the majority of other cases the oviduct exists in the male in a completely rudimentary form; and the semen is carried away by the same duct as the urine.

In Selachians the transportation of the semen from the testis to the Wolffian duct is effected by the junction of the open ends of two or three or more segmental tubes with the testicular follicles, and the modes in which this junction is effected in the higher vertebrates seem to be derivatives from this. If the views here expressed are correct it is by a complete change of function that the oviduct has come to perform its present office. And in the bird and higher vertebrates no trace, or only the very slightest (vide p. 165) of the primitive urinary function is retained during embryonic or adult life.

The last feature in the anatomy of the Selachians which [Pg 146] requires notice is the division of the kidney into two portions, an anterior and posterior. The anatomical similarity between this arrangement and that of higher vertebrates (birds, &c.) is very striking. The anterior one precisely corresponds, anatomically, to the Wolffian body, and the posterior one to the true permanent kidney of higher vertebrates: and when we find that in the Selachians the duct for the anterior serves also for the semen as does the Wolffian duct of higher vertebrates, this similarity seems almost to amount to identity. A discussion of the differences in development in the two cases will come conveniently with the account of the bird; but there appear to me the strongest grounds for looking upon the kidneys of Selachians as equivalent to both the Wolffian bodies and the true kidneys of the higher vertebrates.

The condition of the urinogenital organs in Selachians is by no means the most primitive found amongst vertebrates.

The organs of both Cyclostomous and Osseous fishes, as well as those of Ganoids, are all more primitive; and in the majority of points the Amphibians exhibit a decidedly less differentiated condition of these organs than do the Selachians.

In Cyclostomous fishes the condition of the urinary system is very simple. In Myxine (vide Joh. Müller Myxinoid fishes, and Wilhelm Müller, Jenaische Zeitschrift, 1875, Das Urogenitalsystem des Amphioxus u. d. Cyclostomen) there is a pair of ducts which communicate posteriorly by a common opening with the abdominal pore. From these ducts spring a series of transverse tubules, each terminating in a Malpighian corpuscle. These together constitute the mass of the kidneys. About opposite the gall-bladder the duct of the kidney (the segmental duct) narrows very much, and after a short course ends in a largish glandular mass (the head-kidney), which communicates with the pericardial cavity by a number of openings.

In Petromyzon the anatomy of the kidneys is fundamentally the same as in Myxine. They consist of the two segmental ducts, and a number of fine branches passing off from these, which become convoluted but do not form Malpighian tufts. The head-kidney is absent in the adult.

W. Müller (loc. cit.) has given a short but interesting account of the development of the urinary system of Petromyzon. He [Pg 147] finds that the segmental ducts develop first of all as simple involutions from the body-cavity. The anterior end of each then develops a glandular portion which comes to communicate by a number of openings with the body-cavity. Subsequently to the development of this glandular portion the remainder of the kidneys appears in the posterior portion of the body-cavity; and before the close of embryonic life the anterior glandular portion atrophies.

The comparison of this system with that of a Selachian is very simple. The first developed duct is the segmental duct of a Selachian, and the glandular portion developed at its anterior extremity, which is permanent in Myxine but embryonic in Petromyzon, is, as W. Müller has rightly recognized, equivalent to the head-kidney of Amphibians, which remains undeveloped in Selachians. It is, according to my previously stated view, the glandular portion of the first segmental organ or the segmental duct. The series of orifices by which this communicates with the body-cavity are due to the division of the primary opening of the segmental duct. This is shewn both by the facts of their development in Petromyzon given by Müller, as well as by the occurrence of a similar division of the primary orifice in Amphibians, which is mentioned later in this paper. In a note in my original paper (loc. cit.) I stated that these openings were equivalent to the segmental involutions of Selachians. This is erroneous, and was due to my not having understood the description given in a preliminary paper of Müller (Jenaische Zeitschrift, 1873). The large development of this glandular mass in the Cyclostome and Osseous fishes and in embryo Amphibians, implies that it must at one time have been important. Its earlier development than the remainder of the kidneys is probably a result of the specialized function of the first segmental organ.

The remainder of the kidney in Cyclostomes is equivalent to the kidney of Selachians. Its development from segmental involutions has not been recognized. If these segmental involutions are really absent it may perhaps imply that the simplicity of the Cyclostome kidneys, like that of so many other of their organs, is a result of degeneration rather than a primitive condition.

[Pg 148] In Osseous fishes the segmental duct of the kidneys develops, as the observations of Rosenberg[36] (Teleostierniere, Inaug. Disser. Dorpat, 1867) and Oellacher (Zeitschrift für Wiss. Zool. 1873) clearly prove, by an involution from the body-cavity. This involution grows backwards in the form of a duct and opens into the cloaca. The upper end of this duct (the most anterior segmental tube) becomes convoluted, and forms a glandular body, which has no representative in the urinary apparatus of Selachians, but whose importance, as indicating the origin of the segmental duct of the kidneys, I have already insisted upon.

The rest of the kidney becomes developed at a later period, probably in the same way as in Selachians; but this, as far as I know, has not been made out.

The segmental duct of the kidneys forms the duct for this new gland, as in embryo Selachians (Fig. 2), but, unlike what happens in Selachians, undergoes no further changes, with the exception of a varying amount of retrogressive metamorphosis of its anterior end. The kidneys of Osseous fish usually extend from just behind the head to opposite the anus, or even further back than this. They consist for the most part of a broader anterior portion, an abdominal portion reaching from this to the anus, and, as in those cases in which the kidneys extend further back than the anus, of a caudal portion.

The two ducts (segmental ducts of the kidneys) lie, as a rule, in the lower part of the kidneys on their outer borders, and open almost invariably into a urinary bladder. In some cases they unite before opening into the bladder, but generally have independent openings.

This bladder, which is simply a dilatation of the united lower ends of the primitive kidney-ducts, and has no further importance, is almost invariably present, but in many cases lies unsymmetrically either to the right or the left. It opens to the exterior by a very minute opening in the genito-urinary papilla, immediately behind the genital pore. There are, however, a few cases in which the generative and urinary organs have a [Pg 149] common opening. For further details vide Hyrtl, Denk. der k. Akad. Wien, Vol. II.

It is possible that the generative ducts of Osseous fishes are derived from a splitting from the primitive duct of the kidney, but this is discussed later in the paper.

In Osseous fishes we probably have an embryonic condition of the Selachian kidneys retained permanently through life.

* * * * *

In the majority of Ganoids the division of the segmental duct of the kidney into two would seem to occur, and the ventral duct of the two (Müllerian duct), which opens at its upper end into the body-cavity, is said to serve as an excretory duct for both male and female organs.

The following are the more important facts which are known about the generative and urinary ducts of Ganoids.

In Spatularia (vide Hyrtl, Geschlechts u. Harnwerkzeuge bei den Ganoiden, Denkschriften der k. Akad. Wien, Vol. VIII.) the following parts are found in the female.

(1) The ovaries stretching along the whole length of the abdominal cavity.

(2) The kidneys, which are separate and also extend along the greater part of the abdominal cavity.

(3) The ureters lying on the outer borders of the kidneys. Each ureter dilates at its lower end into an elongated wide tube, which continues to receive the ducts from the kidneys. The two ureters unite before terminating and open behind the anus.

(4) The two oviducts (Müllerian ducts). These open widely into the abdominal cavity, at about two-thirds of the distance from the anterior extremity of the body-cavity. Each opens by a narrow pore into the dilated ureter of its side.

In the male the same parts are found as in the female, but Hyrtl found that the Müllerian duct of the left side at its entrance into the ureter became split into two horns, one of which ended blindly. On the right side the opening of the Müllerian duct was normal.

In the Sturgeon (vide J. Müller, Bau u. Grenzen d. Ganoiden, Berlin Akad. 1844; Leydig, Fischen u. Reptilien, and Hyrtl, Ganoiden) the same parts are found as in Spatularia.

[Pg 150] The kidneys extend along the whole length of the body-cavity; and the ureter, which does not reach the whole length of the kidneys, is a thin-walled wide duct lying on the outer side. On laying it open the numerous apertures of the tubules for the kidney are exposed. The Müllerian duct, which opens in both sexes into the abdominal cavity, ends, according to Leydig, in the cases of some males, blindly behind without opening into the ureter, and Müller makes the same statement for both sexes. It was open on both sides in a female specimen I examined[37], and Hyrtl found it invariably so in both sexes in all the specimens he examined.

Both Rathke and Stannius (I have been unable to refer to the original papers) believed that the semen was carried off by transverse ducts directly into the ureter, and most other observers have left undecided the mechanism of the transportation of the semen to the exterior. If we suppose that the ducts Rathke saw really exist they might perhaps be supposed to enter not directly into the ureter, but into the kidney, and be in fact homologous with the vasa efferentia of the Selachians. The frequent blind posterior termination of the Müllerian duct is in favour of the view that these ducts of Rathke are really present.

In Polypterus (vide Hyrtl, Ganoiden) there is, as in other Ganoids, a pair of Müllerian ducts. They unite at their lower ends. The ureters are also much narrower than in previously described Ganoids and, after coalescing, open into the united oviducts. The urinogenital canal, formed by coalescence of the Müllerian ducts and ureters, has an opening to the exterior immediately behind the anus.

In Amia (vide Hyrtl) there is a pair of Müllerian ducts which, as well as the ureters, open into a dilated vesicle. This vesicle appears as a continuation of the Müllerian ducts, but receives a number of the efferent ductules of the kidneys. There is a single genito-urinary pore behind the anus.

In Ceratodus (Günther, Phil. Trans. 1871) the kidneys are small and confined to the posterior extremity of the abdomen. The generative organs extend however along the greater part of [Pg 151] the length of the abdominal cavity. In both male and female there is a long Müllerian duct, and the ducts of the two sides unite and open by a common pore into a urinogenital cloaca which communicates with the exterior by the same opening as the alimentary canal. In both sexes the Müllerian duct has a wide opening near the anterior extremity of the body-cavity. The ureters coalesce and open together into the urinogenital cloaca dorsal to the Müllerian ducts. It is not absolutely certain that the semen is transported to the exterior by the Müllerian duct of the male, which is perhaps merely a rudiment as in Amphibia. Dr Günther failed however to find any other means by which it could be carried away.

The genital ducts of Lepidosteus differ in important particulars from those of the other Ganoids (vide Müller, loc. cit. and Hyrtl, loc. cit.).

In both sexes the genital ducts are continuous with the investments of the genital organs.

In the female the dilated posterior extremities of the ureters completely invest for some distance the generative ducts, whose extremities are divided into several processes, and end in a different way on the two sides. A similar division and asymmetry of the ducts is mentioned by Hyrtl as occurring in the male of Spatularia, and it seems not impossible that on the hypothesis of the genital ducts being segmental tubes these divisions may be remnants of primitive glandular convolutions. The ureters in both sexes dilate as in other Ganoids at their posterior extremities, and unite with one another. The unpaired urinogenital opening is situated behind the anus. In the male the dilated portion of the ureters is divided into a series of partitions which are not present in the female.

Till the embryology of the secretory system of Ganoids has been worked out, the homologies of their generative ducts are necessarily a matter of conjecture. It is even possible that what I have called the Müllerian duct in the male is functionless, as with Amphibians, but that, owing to the true ducts of the testis having been overlooked, it has been supposed to function as the vas deferens. Günther's (loc. cit.) injection experiments on Ceratodus militate against this view, but I do not think they can be considered as conclusive as long as the [Pg 152] mechanism for the transportation of the semen to the exterior has not been completely made out. Analogy would certainly lead us to expect the ureter to serve in Ganoids as the vas deferens.

The position of the generative ducts might in some cases lead to the supposition that they are not Müllerian ducts, or, in other words, the most anterior pair of segmental organs but a pair of the posterior segmental tubes.

What are the true homologies of the generative ducts of Lepidosteus, which are continuous with the generative glands, is somewhat doubtful. It is very probable that they may represent the similarly functioning ducts of other Ganoids, but that they have undergone further changes as to their anterior extremities.

It is, on the other hand, possible that their generative ducts are the same structures as those ducts of Osseous fishes, which are continuous with the generative organs. These latter ducts are perhaps related to the abdominal pores, and had best be considered in connection with these; but a completely satisfactory answer to the questions which arise in reference to them can only be given by a study of their development.

In the Cyclostomes the generative products pass out by an abdominal pore, which communicates with the peritoneal cavity by two short tubes[38], and which also receives the ducts of the kidneys.

Gegenbaur suggests that these are to be looked upon as Müllerian ducts, and as therefore developed from the segmental ducts of the kidneys. Another possible view is that they are the primitive external openings of a pair of segmental organs. In Selachians there are usually stated to be a pair of abdominal pores. In Scyllium I have only been able to find, on each side, a large deep pocket opening to the exterior, but closed below towards the peritoneal cavity, so that in it there seem to be no abdominal pores[39]. In the Greenland Shark (Læmargus Borealis) [Pg 153] Professor Turner (Journal of Anat. and Phys. Vol. VIII.) failed to find either oviduct or vas deferens, but found a pair of large open abdominal pores, which he believes serve to carry away the generative products of both sexes. Whether the so-called abdominal pores of Selachians usually end blindly as in Scyllium, or, as is commonly stated, open into the body-cavity, there can be no question that they are homologous with true abdominal powers.

The blind pockets of Scyllium appear very much like the remains of primitive involutions from the exterior, which might easily be supposed to have formed the external opening of a pair of segmental organs, and this is probably the true meaning of abdominal pores. The presence of abdominal pores in all Ganoids in addition to true genital ducts and of these pockets or abdominal pores in Selachians, which are almost certainly homologous with the abdominal pores of Ganoids and Cyclostomes, and also occur in addition to true Müllerian ducts, speak strongly against the view that the abdominal pores have any relation to Müllerian ducts. Probably therefore the abdominal pores of the Cyclostomous fishes (which seem to be of the same character as other abdominal pores) are not to be looked on as rudimentary Müllerian ducts.

We next come to the question which I reserved while speaking of the kidneys of Osseous fishes, as to the meaning of their genital ducts.

In the female Salmon and the male and female Eel, the generative products are carried to the exterior by abdominal pores, and there are no true generative ducts. In the case of most other Osseous fish there are true generative ducts which are continuous with the investment of the generative organs[40] and [Pg 154] have generally, though not always, an opening or openings independent of the ureter close behind the rectum, but no abdominal pores are present. It seems, therefore, that in Osseous fish the generative ducts are complementary to abdominal pores, which might lead to the view that the generative ducts were formed by a coalescence of the investment of the generative glands with the short duct of abdominal pore.

Against this view there are, however, the following facts:

(1) In the cases of the salmon and the eel it is perfectly true that the abdominal pore exactly corresponds with the opening of the genital duct in other Osseous fishes, but the absence of genital ducts in these cases must rather be viewed, as Vogt and Pappenheim (loc. cit.) have already insisted, as a case of degeneration than of a primitive condition. The presence of genital ducts in the near allies of the Salmonidæ, and even in the male salmon, are conclusive proofs of this. If we admit that the presence of an abdominal pore in Salmonidæ is merely a result of degeneration, it obviously cannot be used as an argument for the complementary nature of abdominal pores and generative ducts.

(2) Hyrtl (Denkschriften der k. Akad. Wien, Vol. 1) states that in Mormyrus oxyrynchus there is a pair of abdominal pores in addition to true generative ducts. If his statements are correct, we have a strong argument against the generative ducts of Osseous fishes being related to abdominal pores. For though this is the solitary instance of the presence of both a genital opening and abdominal pores known to me in Osseous fishes, yet we have no right to assume that the abdominal pores of Mormyrus are not equivalent to those of Ganoids and Selachians. It must be admitted, with Gegenbaur, that embryology alone can elucidate the meaning of the genital ducts of Osseous fishes.

In Lepidosteus, as was before mentioned, the generative ducts, though continuous with the investment of the generative bodies, unite with the ureters, and in this differ from the generative ducts of Osseous fishes. The relation, indeed, of the [Pg 155] generative ducts of Lepidosteus to the urinary ducts is very similar to that existing in other Ganoid fishes; and this, coupled with the fact that Lepidosteus possesses a pair of abdominal pores on each side of the anus[41], makes it most probable that its generative ducts are true Müllerian ducts.

* * * * *

In the Amphibians the urinary system is again more primitive than in the Selachians.

The segmental duct of the kidneys is formed[42] by an elongated fold arising from the outer wall of the body-cavity, in the same position as in Selachians. This fold becomes constricted into a canal, closed except at its anterior end, which remains open to the body-cavity. This anterior end dilates, and grows out into two horns, and at the same time its opening into the body-cavity becomes partly constricted, and so divided into three separate orifices, one for each horn and a central one between the two. The horns become convoluted, blood channels appearing between their convolutions, and a special coil of vessels is formed arising from the aorta and projecting into the body-cavity near the openings of the convolutions. These formations together constitute the glandular portion[43] of the original anterior segmental tube or segmental duct of the kidneys. I have already pointed out the similarity which this organ exhibits to the head-kidneys of Cyclostome fishes in its mode of formation, especially with reference to the division of the primitive opening. The lower end of the segmental duct unites with a horn of the cloaca.

After the formation of the gland just described the remainder of the kidney is formed.

[Pg 156] This arises in the same way as in Selachians. A series of involutions from the body-cavity are developed; these soon form convoluted tubes, which become branched and interlaced with one another, and also unite with the primitive duct of the kidneys. Owing to the branching and interlacing of the primitive segmental tubes, the kidney is not divided into distinct segments in the same way as with the Selachians. The mode of development of these segmental tubes was discovered by Götte. Their openings are ciliated, and, as Spengel (loc. cit.) and Meyer (loc. cit.) have independently discovered, persist in most adult Amphibians. As both these investigators have pointed out, the segmental openings are in the adult kidneys of most Amphibians far more numerous than the vertebral segments to which they appertain. This is due to secondary changes, and is not to be looked upon as the primitive state of things. At this stage the Amphibian kidneys are nearly in the same condition as the Selachian, in the stage represented in Fig. 2. In both there is the segmental duct of the kidneys, which is open in front, communicates with the cloaca behind, and receives the whole secretion from the kidneys. The parallelism between the two is closely adhered to in the subsequent modifications of the Amphibian kidney, but the changes are not completed so far in Amphibians as in Selachians. The segmental duct of the Amphibian kidney becomes, as in Selachians, split into a Müllerian duct or oviduct, and a Wolffian duct or duct for the kidney.

The following points about this are noteworthy:

(1) The separation of the two ducts is never completed, so that they are united together behind, and for a short distance, blend and form a common duct; the ducts of the two sides so formed also unite before opening to the exterior.

(2) The separation of the two ducts does not occur in the form of a simple splitting, as in Selachians. But the efferent ductules from the kidney gradually alter their points of entrance into the primitive duct. Their points of entrance become carried backwards further and further, and since this process affects the anterior ducts proportionally more than the posterior, the efferent ducts finally all meet and form a common duct which unites with the Müllerian duct near its posterior extremity. [Pg 157] This process is not always carried out with equal completeness. In the tailless Amphibians, however, the process is generally[44] completed, and the ureters (Wolffian ducts) are of considerable length. Bufo cinereus, in the male of which the Müllerian ducts are very conspicuous, serves as an excellent example of this.

In the Salamander (Salamandra maculosa), Figs. 6 and 7, the process is carried out with greater completeness in the female than in the male, and this is the general rule in Amphibians. In the male Proteus, the embryonic condition would seem to be retained almost in its completeness so that the ducts of the kidney open directly and separately into the still persisting primitive duct of the kidney. The upper end of the duct nevertheless extends some distance beyond the end of the kidney and opens into the abdominal cavity. In the female Proteus, on the other hand, the separation into a Müllerian duct and a ureter is quite complete. The Newt (Triton) also serves as an excellent example of the formation of distinct Müllerian and Wolffian ducts being much more complete in the female than the male. In the female Newt all the tubules from the kidney open into a duct of some length which unites with the Müllerian duct near its termination, but in the male the anterior segmental tubes, including those which, as will be afterwards seen, serve as vasa efferentia of the testis, enter the Müllerian duct directly, while the posterior unite as in the female into a common duct before joining the Müllerian duct. For further details as to the variations exhibited in the Amphibians, the reader is referred to Leydig, Anat. Untersuchung, Fischen u. Reptilien. Ditto, Lehrbuch der Histologie, Menschen u. Thiere. Von Wittich, Siebold u. Kölliker, Zeitschrift, Vol. IV. p. 125.

The different conditions of completeness of the Wolffian ducts observable amongst the Amphibians are instructive in reference to the manner of development of the Wolffian duct in Selachians. The mode of division in the Selachians of the segmental duct of the kidney into a Müllerian and Wolffian [Pg 158] duct is probably to be looked upon as an embryonic abbreviation of the process by which these two ducts are formed in Amphibians. The fact that this separation into Müllerian and Wolffian ducts proceeds further in the females of most Amphibians than in the males, strikingly shews that it is the oviductal function of the Müllerian duct which is the indirect cause of its separation from the Wolffian duct. The Müllerian duct formed in the way described persists almost invariably in both sexes, and in the male sometimes functions as a sperm reservoir; e.g. Bufo cinereus. In the embryo it carries at its upper end the glandular mass described above (Kopfniere), but this generally atrophies, though remnants of it persist in the males of some species (e.g. Salamandra). Its anterior end opens, in most cases by a single opening, into the perivisceral cavity in both sexes, and is usually ciliated. As the female reaches maturity, the oviduct dilates very much; but it remains thin and inconspicuous in the male.

The only other developmental change of importance is the connection of the testes with the kidneys. This probably occurs in the same manner as in Selachians, viz. from the junction of the open ends of the segmental tubes with the follicles of the testes. In any case the vessels which carry off the semen constitute part of the kidney, and the efferent duct of the testis is also that of the kidney. The vasa efferentia from the testis either pass through one or two nearly isolated anterior portions of the kidney (Proteus, Triton) or else no such special portion of the kidney becomes separated from the rest, and the vasa efferentia enter the general body of the kidney.

* * * * *

In the male Amphibian, then, the urinogenital system consists of the following parts (Fig. 6):

(1) Rudimentary Müllerian ducts, opening anteriorly into the body-cavity, which sometimes carry aborted Kopfnieren.

(2) The partially or completely formed Wolffian ducts (ureters) which also serve as the ducts for the testes.

(3) The kidneys, parts of which also serve as the vasa efferentia, and whose secretion, together with the testicular products, is carried off by the Wolffian ducts.

[Pg 159] (4) The united lower parts of Wolffian and Müllerian ducts which are really the lower unsplit part of the segmental ducts of the kidneys.

Fig. 6. Diagram of the Urinogenital Organs of a Male Salamander.

(Copied from Leydig's Histologie des Menschen u. der Thiere.)

md. Müller's duct (rudimentary); y. remnant of the secretory portion of the segmental duct Kopfniere; Wd. Wolffian duct; a less complete structure in the male than in the female; st. segmental tubes or kidney. The openings of these into the body-cavity are not inserted in the figure; t. testis. Its efferent ducts form part of the kidney.

In the female, there are (Fig. 7)

(1) The Müllerian ducts which function as the oviducts.

(2) The Wolffian ducts.

(3) The kidneys.

(4) The united Müllerian and Wolffian ducts as in the male.

Fig. 7. Diagram of the Urinogenital Organs of a Female Salamander.

(Copied from Leydig's Histologie des Menschen u. der Thiere.)

Md. Müller's duct or oviduct; Wd. Wolffian duct or the duct of the kidneys; st. segmental tubes or kidney. The openings of these into the body-cavity are not inserted in the figure; o. ovary.

The urinogenital organs of the adult Amphibians agree in almost all essential particulars with those of Selachians. The [Pg 160] ova are carried off in both by a specialized oviduct. The Wolffian duct, or ureter, is found both in Selachians and Amphibians, and the relations of the testis to it are the same in both, the vasa efferentia of the testes having in both the same anatomical peculiarities.

The following points are the main ones in which Selachians and Amphibians differ as to the anatomy of the urinogenital organs; and in all but one of these, the organs of the Amphibian exhibit a less differentiated condition than do those of the Selachian.

(1) A glandular portion (Kopfniere) belonging to the first segmental organ (segmental duct of the kidneys) is found in all embryo Amphibians, but usually disappears, or only leaves a remnant in the adult. It has not yet been found in any Selachian.

(2) The division of the primitive duct of the kidney into the Müllerian duct and the Wolffian duct is not completed so far in Amphibians as Selachians, and in the former the two ducts are confluent at their lower ends.

(3) The permanent kidney exhibits in Amphibians no distinction into two glands (foreshadowing the Wolffian bodies and true kidneys of higher vertebrates), as it does in the Selachians.

(4) The Müllerian duct persists in its entirety in male Amphibians, but only its upper end remains in male Selachians.

(5) The openings of the segmental tubes into the body-cavity correspond in number with the vertebral segments in most Selachians, but are far more numerous than these in Amphibians. This is the chief point in which the Amphibian kidney is more differentiated than the Selachian.

* * * * *

The modifications in development which the urinogenital system has suffered in higher vertebrates (Sauropsida and Mammalia) are very considerable; nevertheless it appears to me to be possible with fair certainty to trace out the relationship of its various parts in them to those found in the Ichthyopsida. The development of urinogenital organs has been far more fully worked out for the bird than for any other member of the amniotic vertebrates; but, as far as we know, [Pg 161] there are no essential variations except in the later periods of development throughout the division. These later variations, concerning for the most part the external apertures of the various ducts, are so well known and have been so fully described as to require no notice here. The development of these parts in the bird will therefore serve as the most convenient basis for comparison.

In the bird the development of these parts begins by the appearance of a column of cells on the upper surface of the intermediate cell-mass (Fig. 8, W.d). As in Selachians, the intermediate cell-mass is a group of cells between the outer edge of the protovertebræ and the upper end of the body-cavity. The column of cells thus formed is the commencement of the duct of the Wolffian body. Its development is strikingly similar to that of the segmental duct of the kidney in Selachians. I shall attempt when I have given an account of the development of the Müllerian duct to speak of the relations between the Selachian duct and that of the bird.

Romiti (Archiv f. Micr. Anat. Vol. X.) has recently stated that the Wolffian duct develops as an involution from the body-cavity. The fact that the specimens drawn by Romiti to support this view are too old to determine such a point, and the inspection of a number of specimens made by my friend Mr Adam Sedgwick of Trinity College, who, at my request, has been examining the urinogenital organs of the fowl, have led me to the conclusion that Romiti is in error in differing from his predecessors as to the development of the Wolffian duct. The solid string of cells to form the Wolffian duct lies at first close to the epiblast, but, by the alteration in shape which the protovertebræ undergo and the general growth of cells around it, becomes gradually carried downwards till it lies close to the germinal epithelium which lines the body-cavity. While undergoing this change of position it also acquires a lumen, but ends blindly both in front and behind. Towards the end of the fourth day the Wolffian duct opens into a horn of the cloaca. The cells adjoining its inner border commence, as it passes down on the third day, to undergo histological changes, which, by the fourth day, result in the formation of a series of ducts and Malpighian tufts which form the mass of the Wolffian body[45]. [Pg 162]

Fig. 8. Transverse section through the Dorsal region of an Embryo Fowl of 45 h. To shew the mode of Formation of the Wolffian Duct.

A. epiblast; B. mesoblast; C. hypoblast; M.c. medullary canal; Pv. Protovertebræ; W.d. Wolffian duct; so. Somatopleure; Sp. Splanchnopleure; pp. pleuro-peritoneal cavity; ch. notochord; ao. dorsal aorta; v. blood-vessels.

[Pg 163]The Müllerian duct arises in the form of an involution, whether at first solid or hollow, of the germinal epithelium, and, as I am satisfied, quite independently of the Wolffian duct. It is important to notice that its posterior end soon unites with the Wolffian duct, from which however it not long after becomes separated and opens independently into the cloaca. The upper end remains permanently open to the body-cavity, and is situated nearly opposite the extreme front end of the Wolffian body.

Between the 80th and 100th hour of incubation the ducts of the permanent kidneys begin to make their appearance. Near its posterior extremity each Wolffian duct becomes expanded, and from the dorsal side of this portion a diverticulum is constricted off, the blind end of which points forwards. This is the duct of the permanent kidneys, and around its end the kidneys are found. It is usually stated that the tubules of the permanent kidneys arise as outgrowths from the duct, but this requires to be worked over again.

The condition of the urinogenital system in birds immediately after the formation of the permanent kidneys is strikingly similar to its permanent condition in adult Selachians. There is the Müllerian duct in both opening in front into the body-cavity and behind into the cloaca. In both the kidneys consist of two parts—an anterior and posterior—which have been called respectively Wolffian bodies and permanent kidneys in birds and Leydig's glands and the kidneys in Selachians.

The duct of the permanent kidney, which at first opens into that of the Wolffian body, subsequently becomes further split off from the Wolffian duct, and opens independently into the cloaca.

[Pg 164] The subsequent changes of these parts are different in the two sexes.

In the female the Müllerian ducts[46] persist and become the oviducts. Their anterior ends remain open to the body-cavity. The changes in their lower ends in the various orders of Sauropsida and Mammalia are too well known to require repetition here. The Wolffian body and duct atrophy: there are left however in many cases slight remnants of the anterior extremity of the body forming the parovarium of the bird, and also frequently remnants of the posterior portion of the gland as well as of the duct. The permanent kidney and its duct remain unaltered.

In the male the Müllerian duct becomes almost completely obliterated. The Wolffian duct persists and forms the vas deferens, and the anterior so-called sexual portion of the Wolffian body also persists in an altered form. Its tubules unite with the seminiferous tubules, and also form the epididymis. Unimportant remnants of the posterior part of the Wolffian body also persist, but are without function. In both sexes the so-called permanent kidneys form the sole portion of the primitive uriniferous system which persists in the adult.

In considering the relations between the modes of development of the urinogenital organs of the bird and of the Selachians, the first important point to notice is, that whereas in the Selachians the segmental duct of the kidneys is first developed and subsequently becomes split into the Müllerian and Wolffian ducts; in the bird these two ducts develop independently. This difference in development would be accurately described by saying that in birds the segmental duct of the kidneys develops as in Selachians, but that the Müllerian duct develops independently of it.

Since in Selachians the Wolffian duct is equivalent to the segmental duct of the kidneys with the Müllerian removed from it, when in birds the Müllerian duct develops independently of the segmental kidney duct, the latter becomes the same as the Wolffian duct.

[Pg 165] The second mode of stating the difference in development in the two cases represents the embryological facts of the bird far better than the other method.

It explains why the Wolffian duct appears earlier than the Müllerian and not at the same time, as one might expect according to the other way of stating the case. If the Wolffian duct is equivalent to the segmental duct of Selachians, it must necessarily be the first duct to develop; and not improbably the development of the Müllerian duct would in birds be expected to occur at the time corresponding to that at which the primitive duct in Selachians became split into two ducts.

It probably also explains the similarity in the mode of development of the Wolffian duct in birds and the primitive duct of the kidneys in Selachians.

This way of stating the case is also in accordance with theoretical conclusions. As the egg-bearing function of the Müllerian duct became more and more confirmed we might expect that the adult condition would impress itself more and more upon the embryonic development, till finally the Müllerian duct ceased to be at any period connected with the kidneys, and the history of its origin ceased to be traceable in its development. This seems to have actually occurred in the higher vertebrates, so that the only persisting connection between the Müllerian duct and the urinary system is the brief but important junction of the two at their lower ends on the sixth or seventh day. This junction justly surprised Waldeyer (Eierstock u. Ei, p. 129), but receives a complete and satisfactory explanation on the hypothesis given above.

The original development of the segmental tubes is in the bird solely retained in the tubules of the Wolffian body arising independently of the Wolffian duct, and I have hitherto failed to find that there is a distinct division of the Wolffian bodies into segments corresponding with the vertebral segments.

I have compared the permanent kidneys to the lower portion of the kidneys of Selachians. The identity of the anatomical condition of the adult Selachian and embryonic bird which has been already pointed out speaks strongly in favour of this view; and when we further consider that the duct of [Pg 166] the permanent kidneys is developed in nearly the same way as the supposed homologous duct in Selachians, the suggested identity gains further support. The only difficulty is the fact that in Selachians the tubules of the part of the kidneys under comparison develop as segmental involutions in point of time anteriorly to their duct, while in birds they develop in a manner not hitherto certainly made out but apparently in point of time posteriorly to their duct. But when the immense modifications in development which the whole of the gland of the excretory organ has undergone in the bird are considered, I do not think that the fact I have mentioned can be brought forward as a serious difficulty.[TN5]

The further points of comparison between the Selachian and the bird are very simple. The Müllerian duct in its later stages behaves in the higher vertebrates precisely as in the lower. It becomes in fact the oviduct in the female and atrophies in the male. The behaviour of the Wolffian duct is also exactly that of the duct which I have called the Wolffian duct in Ichthyopsida, and in the tubules of the Wolffian body uniting with the tubuli seminiferi we have represented the junction of the segmental tubes with the testis in Selachians and Amphibians. It is probably this junction of two independent organs which led Waldeyer to the erroneous view that the tubuli seminiferi were developed from the tubules of the Wolffian body.

With the bird I conclude the history of the origin of the urinogenital system of vertebrates. I have attempted, and I hope succeeded, in tracing out by the aid of comparative anatomy and embryology the steps by which a series of independent and simple segmental organs like those of Annelids have become converted into the complicated series of glands and ducts which constitute the urinogenital system of the higher vertebrates. There are no doubt some points which require further elucidation amongst the Ganoid and Osseous fishes. The most important points which appear to me still to need further research, both embryological and anatomical, are the abdominal pores of fishes, the generative ducts of Ganoids, especially Lepidosteus, and the generative ducts of Osseous fishes.

[Pg 167] The only further point which requires discussion is the embryonic layer from which these organs are derived.

I have shewn beyond a doubt (loc. cit.) that in Selachians these organs are formed from the mesoblast. The unanimous testimony of all the recent investigators of Amphibians leads to the same conclusion. In birds, on the other hand, various investigators have attempted to prove that these organs are derived from the epiblast. The proof they give is the following: the epiblast and mesoblast appear fused in the region of the axis cord. From this some investigators have been led to the conclusion that the whole of the mesoblast is derived from the upper of the two primitive embryonic layers. To these it may be replied that, even granting their view to be correct, it is no proof of the derivation of the urinogenital organs from the epiblast, since it is not till the complete formation of the three layers that any one of them can be said to exist. Others look upon the fusion of the two layers as a proof of the passage of cells from the epiblast into the mesoblast. An assumption in itself, which however is followed by the further assumption that it is from these epiblast cells that the urinogenital system is derived! Whatever may have been the primitive origin of the system, its mesoblastic origin in vertebrates cannot in my opinion be denied.

Kowalewsky (Embryo. Stud. an Vermen u. Arthropoda, Mem. Akad. St Petersbourg, 1871) finds that the segmental tubes of Annelids develop from the mesoblast. We must therefore look upon the mesoblastic origin of the excretory system as having an antiquity greater even than that of vertebrates.

[26] From the Journal of Anatomy and Physiology, Vol. X. 1875.

[27] The more important of these are:—

Semper—Ueber die Stammverwandtschaft der Wirbelthiere u. Anneliden. Centralblatt f. Med. Wiss. 1874, No. 35.

Semper—Segmentalorgane bei ausgewachsenen Haien. Centralblatt f. Med. Wiss. 1874, No. 52.

Semper—Das Urogenitalsystem der höheren Wirbelthiere. Centralblatt f. Med. Wiss. 1874, No. 59.

Semper—Stammesverwandtschaft d. Wirbelthiere u. Wirbellosen. Arbeiten aus Zool. Zootom. Inst. Würzburg. II Band.

Semper—Bildung u. Wachstum der Keimdrüsen bei den Plagiostomen. Centralblatt f. Med. Wiss. 1875, No. 12.

Semper—Entw. d. Wolf. u. Müll. Gang. Centralblatt f. Med. Wiss. 1875, No. 29.

Alex. Schultz—Phylogenie d. Wirbelthiere. Centralblatt f. Med. Wiss. 1874, No. 51.

Spengel—Wimpertrichtern i. d. Amphibienniere. Centralblatt f. Med. Wiss. 1875, No. 23.

Meyer—Anat. des Urogenitalsystems der Selachier u. Amphibien. Sitzb. Naturfor. Gesellschaft. Leipzig, 30 April, 1875.

F. M. Balfour—Preliminary Account of development of Elasmobranch fishes. Quart. Journ. of Micro. Science, Oct. 1874. (This edition, Paper V. p. 60 et seq.)

W. Müller—Persistenz der Urniere bei Myxine glutinosa. Jenaische Zeitschrift, 1873.

W. Müller—Urogenitalsystem d. Amphioxus u. d. Cyclostomen. Jenaische Zeitschrift, 1875.

Alex. Götte—Entwicklungsgeschichte der Unke (Bombinator igneus).

[28] These organs were discovered independently by Professor Semper and myself. Professor Semper's preliminary account appeared prior to my own which was published (with illustrations) in the Quarterly Journal of Mic. Science. Owing to my being in South America, I did not know of Professor Semper's investigations till several months after the publication of my paper.

[29] These outgrowths are at first solid in both Pristiurus, Scyllium and Torpedo, but in Torpedo attain a considerable length before a lumen appears in them.

[30] This duct is often called either Müller's duct, the oviduct, or the duct of the primitive kidneys 'Urnierengang.' None of these terms are very suitable. A justification of the name I have given it will appear from the facts given in the later parts of this paper. In my previous paper I have always called it oviduct, a name which is very inappropriate.

[31] This splitting was first of all discovered and an account of it published by Semper (Centralblatt f. Med. Wiss. 1875, No. 29). I had independently made it out for the female a few weeks before the publication of Semper's account—but have not yet made observations about the point for the male.

My own previous account of the origin of the Wolffian duct (Quart. Journ. of Micros. Science, Oct. 1874, and this edition, Paper V.), is completely false, and was due to my not having had access to a complete series of my sections when I wrote the paper.

[32] Sitzun.[TN4] der Naturfor. Gesellschaft, Leipzig, 30 April, 1875.

[33] We owe to Professor Semper the discovery of the arrangement of the seminal ducts. Centralblatt f. Med. Wiss. 1875, No. 12.

[34] This upper portion of the kidneys is called Leydig's gland by Semper. It would be better to call it the Wolffian body, for I shall attempt to shew that it is homologous with the gland so named in Sauropsida and Mammalia.

[35] Further study of my sections has shewn me that the initial independence of these organs is even more complete than might be gathered from the description in my paper (loc. cit.). I now find, as I before conjectured, that they at first correspond exactly with the muscle-plates, there being one for each muscle-plate. This can be seen in the fresh embryos, but longitudinal sections shew it in an absolutely demonstrable manner.

[36] I am unfortunately only acquainted with Dr Rosenberg's paper from an abstract.

[37] For this specimen I am indebted to Dr Günther.

[38] According to Müller (Myxinoiden, 1845) there is in Myxine an abdominal pore with two short canals leading into it, and Vogt and Pappenheim (An. Sci. Nat. Part IV. Vol. XI.) state that in Petromyzon there are two such pores, each connected with a short canal.

[39] My own rough examination of preserved specimens was hardly sufficient to enable me to determine for certain the presence or absence of these pores. Mr Bridge, of Trinity College, has, however, since then commenced a series of investigations on this point, and informs me that these pores are certainly absent in Scyllium as well as in other genera.

[40] The description of the attachment of the vas deferens to the testis in the Carp given by Vogt and Pappenheim (Ann. Scien. Nat. 1859) does not agree with what I found in the Perch (Perca fluvialis). The walls of the duct are in the Perch continuous with the investment of the testis, and the gland of the testis occupies, as it were, the greater part of the duct; there is, however, a distinct cavity corresponding to what Vogt and P. call the duct, near the border of attachment of the testis into which the seminal tubules open. I could find at the posterior end of the testis no central cavity which could be distinguished from the cavity of this duct.

[41] This is mentioned by Müller (Ganoid fishes, Berlin Akad. 1844), Hyrtl (loc. cit.), and Günther (loc. cit.), and through the courtesy of Dr Günther I have had an opportunity of confirming the fact of the presence of the abdominal pores on two specimens of Lepidosteus in the British Museum.

[42] My account of the development of these parts in Amphibians is derived for the most part from Götte, Die Entwicklungsgeschichte der Unke.

[43] It is called Kopfniere (head-kidney), or Urniere (primitive kidney), by German authors. Leydig correctly looks upon it as together with the permanent kidney constituting the Urniere of Amphibians. The term Urniere is one which has arisen in my opinion from a misconception; but certainly the Kopfniere has no greater right to the appellation than the remainder of the kidney.

[44] In Bombinator igneus, Von Wittich stated that the embryonic condition was retained. Leydig, Anatom. d. Amphib. u. Reptilien, shewed that this is not the case, but that in the male the Müllerian duct is very small, though distinct.

[45] This account of the origin of the Wolffian body differs from that given by Waldeyer, and by Dr Foster and myself (Elements of Embryology, Foster and Balfour), but I have been led to alter my view from an inspection of Mr Sedgwick's preparations, and I hope to shew that theoretical considerations lead to the expectation that the Wolffian body would develop independently of the duct.

[46] The right oviduct atrophies in birds, and the left alone persists in the adult.

[Pg 168]

VIII. On the Development of the Spinal Nerves in Elasmobranch Fishes[47].

With Plates 22 and 23.

In the course of an inquiry into the development of Elasmobranch Fishes, my attention has recently been specially directed to the first appearance and early stages of the spinal nerves, and I have been led to results which differ so materially from those of former investigators, that I venture at once to lay them before the Society. I have employed in my investigations embryos of Scyllium canicula, Scyllium stellare, Pristiurus, and Torpedo. The embryos of the latter animal, especially those hardened in osmic acid, have proved by far the most favourable for my purpose, though, as will be seen from the sequel, I have been able to confirm the majority of my conclusions on embryos of all the above-mentioned genera.

A great part of my work was done at the Zoological Station founded by Dr Dohrn at Naples; and I have to thank both Dr Dohrn and Dr Eisig for the uniformly obliging manner in which they have met my requirements for investigation. I have more recently been able to fill up a number of lacunæ in my observations by the study of embryos bred in the Brighton Aquarium; for these I am indebted to the liberality of Mr Lee and the directors of that institution.

The first appearance of the Spinal Nerves in Pristiurus.

In a Pristiurus-embryo, at the time when two visceral clefts become visible from the exterior (though there are as yet [Pg 169] no openings from without into the throat), a transverse section through the dorsal region exhibits the following features (Pl. 22, fig. A):—

The external epiblast is formed of a single row of flattened elongated cells. Vertically above the neural canal the cells of this layer are more columnar, and form the rudiment of the primitively continuous dorsal fin.

The neural canal (nc) is elliptical in section, and its walls are composed of oval cells two or three deep. The wall at the two sides is slightly thicker than at the ventral and dorsal ends, and the cells at the two ends are also smaller than elsewhere. A typical cell from the side walls of the canal is about 1/1900 inch in its longest diameter. The outlines of the cells are for the most part distinctly marked in the specimens hardened in either chromic or picric acid, but more difficult to see in those prepared with osmic acid; their protoplasm is clear, and in the interior of each is an oval nucleus very large in proportion to the size of its cell. The long diameter of a typical nucleus is about 1/3000 inch, or about two-thirds of that of the cell.

The nuclei are granular, and very often contain several especially large and deeply stained granules; in other cases only one such is present, which may then be called a nucleolus.

In sections there may be seen round the exterior of the neural tube a distinct hyaline membrane: this becomes stained of a brown colour with osmic acid, and purple or red with hæmatoxylin or carmine respectively. Whether it is to be looked upon as a distinct membrane differentiated from the outermost portion of the protoplasm of the cells, or as a layer of albumen coagulated by the reagents applied, I am unable to decide for certain. It makes its appearance at a very early period, long before that now being considered; and similar membranes are present around other organs as well as the neural tube. The membrane is at this stage perfectly continuous round the whole exterior of the neural tube as well on the dorsal surface as on the ventral.

The section figured, whose features I am describing, belongs to the middle of the dorsal region. Anteriorly to this point the spinal cord becomes more elliptical in section, and the spinal canal more lanceolate; posteriorly, on the other hand, the spinal [Pg 170] canal and tube become more nearly circular in section. Immediately beneath the neural tube is situated the notochord (ch). It exhibits at this stage a central area rich in protoplasm, and a peripheral layer very poor in protoplasm; externally it is invested by a distinct cuticular membrane.

Beneath the notochord is a peculiar rod of cells, constricted from the top of the alimentary canal[48]. On each side and below this are the two aortæ, just commencing to be formed, and ventral to these is the alimentary canal.

On each side of the body two muscle-plates are situated; their upper ends reach about one-third of the way up the sides of the neural tube. The two layers which together constitute the muscle-plates are at this stage perfectly continuous with the somatic and splanchnic layers of the mesoblast, and the space between the two layers is continuous with the body-cavity. In addition to the muscle-plates and their ventral continuations, there are no other mesoblast-cells to be seen. The absence of all mesoblastic cells dorsal to the superior extremities of the muscles is deserving of special notice.

Very shortly after this period and, as a rule, before a third visceral cleft has become visible, the first traces of the spinal nerves make their appearance.

First Stage.—The spinal nerves do not appear at the same time along the whole length of the spinal canal, but are formed first of all in the neck and subsequently at successive points posterior to this.

Their mode of formation will be most easily understood by referring to Pl. 22, figs. B I, B II, B III, which are representations of three sections taken from the same embryo. B I is from the region of the heart; B II belongs to a part of the body posterior to this, and B III to a still posterior region.

In most points the sections scarcely differ from Pl. 22, fig. A, which, indeed, might very well be a posterior section of the embryo to which these three sections belong.

The chief point, in addition to the formation of the spinal nerves, which shews the greater age of the embryo from which the sections were taken is the complete formation of the aortæ.

[Pg 171] The upper ends of the muscle-plates have grown no further round the neural canal than in fig. A, and no scattered mesoblastic connective-tissue cells are visible.

In fig. A the dorsal surface of the neural canal was as completely rounded off as the ventral surface; but in fig. B III this has ceased to be the case. The cells at the dorsal surface of the neural canal have become rounder and smaller and begun to proliferate, and the uniform outline of the neural canal has here become broken (fig. B III, pr). The peculiar membrane completely surrounding the canal in fig. A now terminates just below the point where the proliferation of cells is taking place.

The prominence of cells which springs in this way from the top of the neural canal is the commencing rudiment of a pair of spinal nerves. In fig. B II, a section anterior to fig. B III, this formation has advanced much further (fig. B II, pr). From the extreme top of the neural canal there have now grown out two club-shaped masses of cells, one on each side; they are perfectly continuous with the cells which form the extreme top of the neural canal, and necessarily also are in contact with each other dorsally. Each grows outwards in contact with the walls of the neural canal; but, except at the point where they take their origin, they are not continuous with its walls, and are perfectly well separated by a sharp line from them.

In fig. B I, though the club-shaped processes still retain their attachment to the summit of the neural canal, they have become much longer and more conspicuous.

Specimens hardened in both chromic acid (Pl. 22, fig. C) and picric acid give similar appearances as to the formation of these bodies.

In those hardened in osmic acid, though the mutual relations of the masses of cells are very clear, yet it is difficult to distinguish the outlines of the individual cells.

In the chromic acid specimens (fig. C) the cells of these rudiments appear rounded, and each of them contains a large nucleus.

I have been unable to prepare longitudinal sections of this stage, either horizontal or vertical, to shew satisfactorily the extreme summit of the spinal cord; but I would call attention [Pg 172] to the fact that the cells forming the proximal portion of the outgrowth are seen in every transverse section at this stage, and therefore exist the whole way along, whereas the distal portion is seen only in every third or fourth section, according to the thickness of the sections. It may be concluded from this that there appears a continuous outgrowth from the spinal canal, from which discontinuous processes grow out.

In specimens of a very much later period (Pl. 23, fig. I) the proximal portions of the outgrowth are unquestionably continuous with each other, though their actual junctions with the spinal cord are very limited in extent. The fact of this continuity at a later period is strongly in favour of the view that the posterior branches of the spinal nerves arise from the first as a continuous outgrowth of the spinal cord, from which a series of distal processes take their origin. I have, however, failed to demonstrate this point absolutely. The processes, which we may call the nerve-rudiments, are, as appears from the later stages, equal in number to the muscle-plates.

It may be pointed out, as must have been gathered from the description above, that the nerve-rudiments have at this stage but one point of attachment to the spinal cord, and that this one corresponds with the dorsal or posterior root of the adult nerve.

The rudiments are, in fact, those of the posterior root only.

The next or second stage in the formation of these structures to which I would call attention occurs at about the time when three to five visceral clefts are present. The disappearance from the notochord in the anterior extremity of the body of a special central area rich in protoplasm serves as an excellent guide to the commencement of this epoch.

Its investigation is beset with far greater difficulties than the previous one. This is owing partly to the fact that a number of connective-tissue cells, which are only with great difficulty to be distinguished from the cells which compose the spinal nerves, make their appearance around the latter, and partly to the fact that the attachment of the spinal nerves to the neural canal becomes much smaller, and therefore more difficult to study.

Fortunately, however, in Torpedo these peculiar features [Pg 173] are not present to nearly the same extent as in Pristiurus and Scyllium.

The connective-tissue cells, though they appear earlier in Torpedo than in the two other genera, are much less densely packed, and the large attachment of the nerves to the neural canal is retained for a longer period.

Under these circumstances I consider it better, before proceeding with this stage, to give a description of the occurrences in Torpedo, and after that to return to the history of the nerves in the genera Pristiurus and Scyllium.

The development of the Spinal Nerves in Torpedo.

The youngest Torpedo-embryo in which I have found traces of the spinal nerves belongs to the earliest part of what I called the second stage.

The segmental duct[49] is just appearing, but the cells of the notochord have not become completely vacuolated. The rudiments of the spinal nerves extend half of the way towards the ventral side of the spinal cord; they grow out in a most distinct manner from the dorsal surface of the spinal cord (Pl. 22, fig. D a, pr); but the nerve-rudiments of the two sides are no longer continuous with each other at the dorsal median line, as in the earlier Pristiurus-embryos. The cells forming the proximal portion of the rudiment have the same elongated form as the cells of the spinal cord, but the remaining cells are more circular.

From the summit of the muscle-plates (mp) an outgrowth of connective tissue has made its appearance (c), which eventually fills up the space between the dorsal surface of the cord and the external epiblast. There is not the slightest difficulty in distinguishing the connective-tissue cells from the nerve-rudiment. I believe that in this embryo the origin of the nerves from the neural canal was a continuous one, though naturally the peripheral ends of the nerve-rudiments were separate from each other.

The most interesting feature of the stage is the commencing formation of the anterior roots. Each of these arises (Pl. 22, [Pg 174] fig. D a, ar) as a small but distinct outgrowth from the epiblast of the spinal cord, near the ventral corner of which it appears as a conical projection. Even from the very first it has an indistinct form of termination and a fibrous appearance, while the protoplasm of which it is composed becomes very attenuated towards its termination.

The points of origin of the anterior roots from the spinal cord are separated from each other by considerable intervals. In this fact, and also in the nerves of the two sides never being united with each other in the ventral median line, the anterior roots exhibit a marked contrast to the posterior.

There exists, then, in Torpedo-embryos by the end of this stage distinct rudiments of both the anterior and posterior roots of the spinal nerves. These rudiments are at first quite independent of and disconnected with each other, and both take their rise as outgrowths of the epiblast of the neural canal.

The next Torpedo-embryo (Pl. 22, fig. D b), though taken from the same female, is somewhat older than the one last described. The cells of the notochord are considerably vacuolated; but the segmental duct is still without a lumen. The posterior nerve-rudiments are elongated, pear-shaped bodies of considerable size, and, growing in a ventral direction, have reached a point nearly opposite the base of the neural canal. They still remain attached to the top of the neural canal, though the connexion has in each case become a pedicle so narrow that it can only be observed with great difficulty.

It is fairly certain that by this stage each posterior nerve-rudiment has its own separate and independent junction with the spinal cord; their dorsal extremities are nevertheless probably connected with each other by a continuous commissure.

The cells composing the rudiments are still round, and have, in fact, undergone no important modifications since the last stage.

The important feature of the section figured (fig. D b), and one which it shares with the other sections of the same embryo, is the appearance of connective-tissue cells around the nerve-rudiment. These cells arise from two sources; one of these is supplied by the vertebral rudiments, which at the end of [Pg 175] the last stage (Pl. 22, fig. C, vr) become split off from the inner layer of the muscle-plates. The vertebral rudiments have in fact commenced to grow up on each side of the neural canal, in order to form the mass of cells out of which the neural arches are subsequently developed.

The dorsal extremities of the muscle-plates form the second source of these connective-tissue cells. These latter cells lie dorsal and external to the nerve-rudiments.

The presence of this connective tissue, in addition to the nerve-rudiments, removes the possibility of erroneous interpretations in the previous stages of the Pristiurus-embryo.

It might be urged that the two masses which I have called nerve-rudiments are nothing else than mesoblastic connective tissue commencing to develop around the neural canal, and that the appearance of attachment to the neural canal which they present is due to bad preparation or imperfect observation. The sections of both this and the last Torpedo-embryo which I have been describing clearly prove that this is not the case. We have, in fact, in the same sections the developing connective tissue as well as the nerve-rudiments, and at a time when the latter still retains its primitive attachment to the neural canal. The anterior root (fig. D b, ar) is still a distinct conical prominence, but somewhat larger than in the previously described embryo; it is composed of several cells, and the cells of the spinal cord in its neighbourhood converge towards its point of origin.

In a Torpedo-embryo (Pl. 22, fig. D c) somewhat older than the one last described, though again derived from the oviduct of the same female, both the anterior and the posterior rudiments have made considerable steps in development.

In sections taken from the hinder part of the body I found that the posterior rudiments nearly agreed in size with those in fig. D b.

It is, however, still less easy than there to trace the junction of the posterior rudiments with the spinal cord, and the upper ends of the rudiments of the two sides do not nearly meet.

In a considerable series of sections I failed to find any case [Pg 176] in which I could be absolutely certain that a junction between the nerve and the spinal cord was effected; and it is possible that in course of the change of position which this junction undergoes there may be for a short period a break of continuity between the nerve and the cord. This, however, I do not think probable. But if it takes place at all, it takes place before the nerve becomes functionally active, and so cannot be looked upon as possessing any physiological significance.

The rudiment of the posterior nerve in the hinder portion of the body is still approximately homogeneous, and no distinction of parts can be found in it.

In the same region of the body the anterior rudiment retains nearly the same condition as in the previous stage, though it has somewhat increased in size.

In the sections taken from the anterior part of the same embryo the posterior rudiment has both grown in size and also commenced to undergo histological changes by which it has become divided into a root, a ganglion, and a nerve.

The root (fig. D c, pr) consists of small round cells which lie close to the spinal cord, and ends dorsally in a rounded extremity.

The ganglion (g) consists of larger and more elongated cells, and forms an oval mass enclosed on the outside by the downward continuation of the root, having its inner side nearly in contact with the spinal cord.

From its ventral end is continued the nerve, which is of considerable length, and has a course approximately parallel to that of the muscle-plate. It forms a continuation of the root rather than of the ganglion.

Further details in reference to the histology of the nerve-rudiment at this stage are given later in this paper, in the description of Pristiurus-embryos, of which I have a more complete series of sections than of the Torpedo-embryos.

When compared with the nerve-rudiment in the posterior part of the same embryo, the nerve-rudiment last described is, in the first place, considerably larger, and has secondly undergone changes, so that it is possible to recognize in it parts which can be histologically distinguished as nerve and ganglion.

The developmental changes which have taken place in the [Pg 177] anterior root are not less important than those in the posterior. The anterior root now forms a very conspicuous cellular prominence growing out from the ventral corner of the spinal cord (fig. D c, ar). It has a straight course from the spinal cord to the muscle-plate, and there shews a tendency to turn downwards at an open angle: this, however, is not represented in the specimen figured. The cells of which it is composed each contain a large oval nucleus, and are not unlike the cells which form the posterior rudiment. The anterior and posterior nerves are still quite unconnected with each other; and in those sections in which the anterior root is present the posterior root of the same side is either completely absent or only a small part is to be seen. The cells of the spinal cord exhibit a slight tendency to converge towards the origin of the anterior nerve-root.

In the spinal cord itself the epithelium of the central canal is commencing to become distinguished from the grey matter, but no trace of the white matter is visible.

I have succeeded in making longitudinal vertical sections of this stage, which prove that the ends of the posterior roots adjoining the junction with the cord are all connected with each other (Pl. 22, fig. D d).

If the figure representing a transverse section of the embryo (fig. D c) be examined, or better still the figure of a section of the slightly older Scyllium-embryo (Pl. 23, fig. HI or II), the posterior root will be seen to end dorsally in a rounded extremity, and the junction with the spinal cord to be effected, not by the extremity of the nerve, but by a part of it at some little distance from this.

It is from these upper ends of the rudiments beyond the junction with the spinal cord that I believe the commissures to spring which connect together the posterior roots.

My sections shewing this for the stage under consideration are not quite as satisfactory as is desirable; nevertheless they are sufficiently good to remove all doubt as to the presence of these commissures.

A figure of one of these sections is represented (Pl. 22, fig. D d). In this figure pr points to the posterior roots and x to the commissures uniting them.

[Pg 178] In a stage somewhat subsequent to this I have succeeded in making longitudinal sections, which exhibit these junctions with a clearness which leaves nothing to be desired.

It is there effected (Pl. 23, fig. L) in each case by a protoplasmic commissure with imbedded nuclei[50]. Near its dorsal extremity each posterior root dilates, and from the dilated portion is given off on each side the commissure uniting it with the adjoining roots.

Considering the clearness of this formation in this embryo, as well as in the embryo belonging to the stage under description, there cannot be much doubt that at the first formation of the posterior rudiments a continuous outgrowth arises from the spinal cord, and that only at a later period do the junctions of the roots with the cord become separated and distinct for each nerve.

I now return to the more complete series of Pristiurus-embryos, the development of whose spinal nerves I have been able to observe.

Second Stage of the Spinal Nerves in Pristiurus.

In the youngest of these (Pl. 22, fig. E) the notochord has undergone but very slight changes, but the segmental duct has made its appearance, and is as much developed as in the Torpedo-embryo from which fig. D b was taken.

(The embryo from which fig. E a was derived had three visceral clefts.)

There have not as yet appeared any connective-tissue cells dorsal to the top of the muscle-plates, so that the posterior nerve-rudiments are still quite free and distinct.

The cells composing them are smaller than the cells of the neural canal; they are round and nucleated; and, indeed, in their histological constitution the nerve-rudiments exhibit no important deviations from the previous stage, and they have hardly increased in size. In their mode of attachment to the neural tube an important change has, however, already commenced to be visible.

In the previous stage the two nerve-rudiments met above the [Pg 179] summit of the spinal cord and were broadly attached to it there; now their points of attachment have glided a short distance down the sides of the spinal cord[51].

The two nerve-rudiments have therefore ceased to meet above the summit of the canal; and in addition to this they appear in section to narrow very much before becoming united with its walls, so that their junctions with these appear in a transverse section to be effected by at most one or two cells, and are, comparatively speaking, very difficult to observe.

In an embryo but slightly older than that represented in Fig. E a the first rudiment of the anterior root becomes visible. This appears, precisely as in Torpedo, in the form of a small projection from the ventral corner of the spinal cord (fig. E b, ar).

The second step in this stage (Pl. 22, fig. F) is comparable, as far as the connective-tissue is concerned, with the section of Torpedo (Pl. 22, fig. D d). The notochord (the histological details of whose structure are not inserted in this figure) is rather more developed, and the segmental duct, as was the case with the corresponding Torpedo-embryo, has become hollow at its anterior extremity.

The embryo from which the section was taken possessed five visceral clefts, but no trace of external gills.

In the section represented, though from a posterior part of the body, the dorsal nerve-rudiments have become considerably larger than in the last embryo; they now extend beyond the base of the neural canal. They are surrounded to a great extent by mesoblastic tissue, which, as in the case of the Torpedo, takes its origin from two sources, (1) from the commencing vertebral bodies, (2) from the summits of the muscle-plates.

It is in many cases very difficult, especially with chromic-acid specimens, to determine with certainty the limits of the rudiments of the posterior root.

[Pg 180] In the best specimens a distinct bordering line can be seen, and it is, as a rule, possible to state the characters by which the cells of the nerve-rudiments and vertebral bodies differ. The more important of these are the following:—(1) The cells of the nerve-rudiment are distinctly smaller than those of the vertebral rudiment; (2) the cells of the nerve-rudiment are elongated, and have their long axis arranged parallel to the long axis of the nerve-rudiment, while the cells surrounding them are much more nearly circular.

The cells of the nerve-rudiment measure about 1/1600 × 1/4500 to 1/1600 × 1/3200 inch, those of the vertebral rudiment 1/1600 × 1/1900 inch. The greater difficulty experienced in distinguishing the nerve-rudiment from the connective-tissue in Pristiurus than in Torpedo arises from the fact that the connective-tissue is much looser and less condensed in the latter than in the former.

The connective-tissue cells which have grown out from the muscle-plates form a continuous arch over the dorsal surface of the neural tube (vide Pl. 22, fig. F): and in some specimens it is difficult to see whether the arch is formed by the rudiment of the posterior root or by connective-tissue. It is, however, quite easy with the best specimens to satisfy one's self that it is from the connective-tissue, and not the nerve-rudiment, that the dorsal investment of the neural canal is derived.

As in the previous case, the upper ends of each pair of posterior nerve-rudiments are quite separate from one another, and appear in sections to be united by a very narrow root to the walls of the neural canal at the position indicated in fig. F[52].

The cells forming the nerve-rudiments have undergone slight modifications; they are for the most part more distinctly elongated than in the earlier stage, and appear slightly smaller in comparison with the cells of the neural canal.

They possess as yet no distinctive characters of nerve-cells. They stain more deeply with osmic acid than the cells around them, but with hæmatoxylin there is but a very slight difference in intensity between their colouring and that of the neighbouring connective-tissue cells.

The anterior roots have grown considerably in length, but [Pg 181] their observation is involved in the same difficulties with chromic-acid specimens as that of the posterior rudiments.

There is a further difficulty in observing the anterior roots, which arises from the commencing formation of white matter in the cord. This is present in all the anterior sections of the embryo from which fig. F is taken. When the white matter is formed the cells constituting the junction of the anterior nerve-root with the spinal cord undergo the same changes as the cells which are being converted into the white matter of the cord, and become converted into nerve-fibres; these do not stain with hæmatoxylin, and thus an apparent space is left between the nerve-root and the spinal cord. This space by careful examination may be seen to be filled up with fibres. In osmic acid sections, although even in these the white matter is stained less deeply than the other tissues, it is a matter of comparative ease to observe the junction between the anterior nerve root and the spinal cord.

I have been successful in preparing satisfactory longitudinal sections of embryos somewhat older than that shewn in fig. F, and they bring to light several important points in reference to the development of the spinal nerves. Three of these sections are represented in Pl. 22, figs. G1, G2, and G3.

The sections are approximately horizontal and longitudinal. G1 is the most dorsal of the three; it is not quite horizontal though nearly longitudinal. The section passes exactly through the point of attachment of the posterior roots to the walls of the neural canal.

The posterior rudiments appear as slight prominences of rounded cells projecting from the wall of the neural canal. From transverse sections the attachment of the nerves to the wall of the neural canal is proved to be very narrow, and from these sections it appears to be of some length in the direction of the long axis of the embryo. A combination of the sections taken in the two directions leads to the conclusion that the nerves at this stage thin out like a wedge before joining the spinal cord.

The independent junctions of the posterior rudiments with the spinal cord at this stage are very clearly shewn, though the rudiments are probably united with each other just dorsal to their junction with the spinal cord.

[Pg 182] The nerves correspond in number with the muscle-plates, and each arises from the spinal cord, nearly opposite the middle line of the corresponding muscle-plates (figs. G1 and G2).

Each nerve-rudiment is surrounded by connective-tissue cells, and is separated from its neighbours by a considerable interval.

At its origin each nerve-rudiment lies opposite the median portion of a muscle-plate (figs. G1 and G2); but, owing to the muscle-plate acquiring an oblique direction, at the level of the dorsal surface of the notochord it appears in horizontal sections more nearly opposite the interval between two muscle-plates (figs. G2 and G3).

In horizontal sections I find masses of cells which make their appearance on a level with the ventral surface of the spinal cord. I believe I have in some sections successfully traced these into the spinal cord, and I have little doubt that they are the anterior roots of the spinal nerves; they are opposite the median line of the muscle-plates, and do not appear to join the posterior roots (vide fig. G3, ar).

At the end of this period or second stage the main characters of the spinal nerves in Pristiurus are the following:—

(1) The posterior nerve-rudiments form somewhat wedge-shaped masses of tissue attached dorsally to the spinal cord.

(2) The cells of which they are composed are typical undifferentiated embryonic cells, which can hardly be distinguished from the connective-tissue cells around them.

(3) The nerves of each pair no longer meet above the summit of the spinal canal, but are independently attached to its sides.

(4) Their dorsal extremities are probably united by commissures.

(5) The anterior roots have appeared; they form small conical projections from the ventral corner of the spinal cord, but have no connexion with the posterior rudiments.

The Third Stage of the Spinal Nerves in Pristiurus.

With the third stage the first distinct histological differentiations of the nerve-rudiments commence. Owing to the [Pg 183] changes both in the nerves themselves and in the connective-tissue around them, which becomes less compact and its cells stellate, the difficulty of distinguishing the nerves from the surrounding cells vanishes; and the difficulties of investigation in the later stages are confined to the modes of attachment of the nerves to the neural canal, and the histological changes which take place in the rudiments themselves.

The stage may be considered to commence at the period when the external gills first make their appearance as small buds from the walls of the visceral clefts. Already, in the earliest rudiments of the posterior root of this period now figured, a number of distinct parts are visible (Pl. 23, fig. H I).

Surrounding nearly the whole structure there is present a delicate investment similar to that which I mentioned as surrounding the neural canal and other organs; it is quite structureless, but becomes coloured with all staining reagents. I must again leave open the question whether it is to be looked upon as a layer of coagulated protoplasm or as a more definite structure. This investment completely surrounds the proximal portion of the posterior root, but vanishes near its distal extremity.

The nerve-rudiment itself may be divided into three distinct portions:—(1) the proximal portion, in which is situated the pedicle of attachment to the wall of the neural canal; (2) an enlarged portion, which may conveniently, from its future fate, be called the ganglion; (3) a distal portion beyond this. The proximal portion presents a fairly uniform diameter, and ends dorsally in a rounded expansion; it is attached remarkably enough, not by its extremity, but by its side, to the spinal cord. The dorsal extremities of the posterior nerves are therefore free; as was before mentioned, they probably serve as the starting-point of the longitudinal commissures between the posterior roots.

The spinal cord at this stage is still made up of fairly uniform cells, which do not differ in any important particulars from the cells which composed it during the last stage. The outer portion of the most peripheral layer of cells has already begun to be converted into the white matter.

The delicate investment spoken of before still surrounds the [Pg 184] whole spinal cord, except at the points of junction of the cord with the nerve-rudiments. Externally to this investment, and separated from it for the most part by a considerable interval, a mesoblastic sheath (Pl. 23, fig. H I, i) for the spinal cord is beginning to be formed.

The attachment of the nerve-rudiments to the spinal cord, on account of its smallness, is [TN6] still very difficult to observe. In many specimens where the nerve is visible a small prominence may be seen rising up from the spinal cord at a point corresponding to x (Pl. 23, fig. H I). It is, however, rare to see this prominence and the nerve continuous with each other: as a rule they are separated by a slight space, and frequently one of the cells of the mesoblastic investment of the spinal cord is interposed between the two. In some especially favourable specimens, similar to the one figured, there can be seen a distinct cellular prominence (fig. H I, x) from the spinal cord, which becomes continuous with a small prominence on the lateral border of the nerve-rudiment near its free extremity. The absence of a junction between the two in a majority of sections is only what might be expected, considering how minute the junction is.

Owing to the presence of the commissure connecting the posterior roots, some part of a nerve is present in every section.

The proximal extremity of the nerve-rudiment itself is composed of cells, which, by their smaller size and a more circular form, are easily distinguished from cells forming the ganglionic portion of the nerve.

The ganglionic portion of the nerve, by its externally swollen configuration, is at once recognizable in all the sections in which the nerve is complete. The delicate investment before mentioned is continuous around it. The cells forming it are larger and more elongated than the cells forming the upper portion of the nerve-rudiment: each of them possesses a large and distinct nucleus.

The remainder of the nerve rudiment forms the commencement of the true nerve. It can in this stage be traced only for a very small distance, and gradually fades away, in such a manner that its absolute termination is very difficult to observe.

The connective-tissue cells which surround the nerve-rudiment [Pg 185] are far looser than in the last stage, and are commencing to throw out processes and become branched.

The anterior root-nerve has grown very considerable since the last stage. It projects from the same region of the cord as before, but on approaching the muscle-plate takes a sudden bend downwards (fig. H II, ar).

I have failed to prove that the anterior and posterior roots are at this stage united.

Fourth Stage.

In an embryo but slightly more advanced than the one last described, important steps have been made in the development of the nerve-rudiment. The spinal cord itself now possesses a covering of white matter; this is thickest at the ventral portion of the cord, and extends to the region of the posterior root of the spinal nerve.

The junction of the posterior root with the spinal cord is easier to observe than in the last stage.

It is still effected by means of unaltered cells, though the cells which form the projection from the cord to the nerve are commencing to undergo changes similar to those of the cells which are being converted into white matter.

In the rudiment of the posterior root itself there are still three distinct parts, though their arrangement has undergone some alteration and their distinctness has become more marked (Pl. 23, fig. I I).

The root of the nerve (fig. I I, pr) consists, as before, of nearly circular cells, each containing a nucleus, very large in proportion to the size of the cell. The cells have a diameter of about 1/3000 of an inch. This mass forms not only the junction between the ganglion and the spinal canal, but is also continued into a layer investing the outer side of the ganglion and continuous with the nerve beyond the ganglion.

The cells which compose the ganglion (fig. I I, sp.g) are easily distinguished from those of the root. Each cell is elongated with an oval nucleus, large in proportion to the cell; and its protoplasm appears to be continued into an angular, not to say fibrous process, sometimes at one and more rarely at [Pg 186] both ends. The processes of the cells are at this stage very difficult to observe: figs. I a, I b, I c represent three cells provided with them and placed in the positions they occupied in the ganglion.

The relatively very small amount of protoplasm in comparison to the nucleus is fairly represented in these figures, though not in the drawing of the ganglion as a whole. In the centre of each nucleus is a nucleolus.

Fig. I b, in which the process points towards the root of the nerve, I regard as a commencing nerve-fibre: its more elongated shape seems to imply this. In the next stage special bundles of nerve-fibres become very conspicuous in the ganglion. The long diameter of an average ganglion-cell is about 1/1600 of an inch. The whole ganglion forms an oval mass, well separated both from the nerve-root and the nerve, and is not markedly continuous with either. On its outer side lies the downward process of the nerve-root before mentioned.

The nerve itself is still, as in the last case, composed of cells which are larger and more elongated than either the cells of the root or the ganglion.

The condition of the anterior root at this stage is hardly altered from what it was; it is composed of very small cells, which with hæmatoxylin stain more deeply than any other cell of the section. A figure of it is given in I II.

Horizontal longitudinal sections of this stage are both easy to make and very instructive. On Pl. 23, fig. K I is represented a horizontal section through a plane near the dorsal surface of the spinal cord: each posterior root is seen in this section to lie nearly opposite the anterior extremity of a muscle-plate.

In a more ventral plane (fig. K II) this relation is altered, and the posterior roots lie opposite the hinder parts of the muscle-plates.

The nerves themselves are invested by the hyaline membrane spoken of above; and surrounding this again there is present a delicate mesoblastic investment of spindle-shaped cells.

Longitudinal sections also throw light upon the constitution of the anterior nerve roots (vide fig. K II, ar). In the two segments on the left-hand side in this figure the anterior roots [Pg 187] are cut through as they are proceeding, in a more or less horizontal course, from the spinal cord to the muscle-plates.

Where the section (which is not quite horizontal) passes through the plane of the notochord, as on the right-hand side, the anterior roots are cut transversely. Each root, in fact, changes its direction, and takes a downward course.

The anterior roots are situated nearly opposite the middle of the muscle-plates: their section is much smaller than that of the posterior roots, and with hæmatoxylin they stain more deeply than any of the other cells in the preparation.

The anterior roots, so far as I have been able to observe, do not at this stage unite with the posterior; but on this point I do not speak with any confidence.

The period now arrived at forms a convenient break in the development of the spinal nerves; and I hope to treat the remainder of the subject, especially the changes in the ganglion, the development of the ganglion-cells, and of the nerve-fibres, in a subsequent paper.

I will only add that, not long after the stage last described, the posterior root unites with the anterior root at a considerable distance below the cord: this is shewn in Pl. 23, fig. L. Still later the portion of the root between the ganglion and the spinal cord becomes converted into nerve-fibres, and the ganglion becomes still further removed from the cord, while at the same time it appears distinctly divided into two parts.

As regards the development of the cranial nerves, I have made a few observations, which, though confessedly incomplete, I would desire to mention here, because, imperfect as they are, they seem to shew that in Elasmobranch Fishes the cranial nerves resemble the spinal nerves in arising as outgrowths from the central nervous system.

I have given a figure of the development of a posterior root of a cranial nerve in fig. M I. The section is taken from the same embryo as figs. B I, B II, and B III.

It passes through the anterior portion of a thickening of the external epiblast, which eventually becomes involuted as the auditory vesicle.

The posterior root of a nerve (VII) is seen growing out from the summit of the hind brain in precisely the same manner that [Pg 188] the posterior roots of the spinal nerves grow out from the spinal cord: it is the rudiment of the seventh or facial nerve. The section behind this (fig. M II), still in the region of the ear, has no trace of a nerve, and thus serves to shew the early discontinuity of the posterior nerve-rudiments which arise from the brain.

I have as yet failed to detect any cranial anterior roots like those of the spinal nerves[53]. The similarity in development between the cranial and spinal nerves is especially interesting, as forming an important addition to the evidence which at present exists that the cranial nerves are only to be looked on as spinal nerves, especially modified in connexion with the changes which the anterior extremity of the body has undergone in existing vertebrates.

* * * * *

My results may be summarized as follows:—

Along the extreme dorsal summit of the spinal cord there arises on each side a continuous outgrowth.

From each outgrowth processes corresponding in number to the muscle-plates grow downwards. These are the posterior nerve-rudiments.

The outgrowths, at first attached to the spinal cord throughout their whole length, soon cease to be so, and remain in connexion with it in certain spots only, which form the junctions of the posterior roots with the spinal cord.

The original outgrowth on each side remains as a bridge, uniting together the dorsal extremities of all the posterior rudiments. The points of junction of the posterior roots with the spinal cord are at first situated at the extreme dorsal summit of the latter, but eventually travel down, and are finally placed on the sides of the cord.

After these events the posterior nerve-rudiments grow rapidly in size, and become differentiated into a root (by which they are attached to the spinal canal), a ganglion, and a nerve.

The anterior roots, like the posterior, are outgrowths from the spinal cord; but the outgrowths to form them are from the [Pg 189] first discontinuous, and the points from which they originally spring remain as those by which they are permanently attached to the spinal cord, and do not, as in the case of the posterior roots, undergo a change of position. The anterior roots arise, not vertically below, but opposite the intervals between the posterior roots.

The anterior roots are at first quite separate from the posterior roots; but soon after the differentiation of the posterior rudiment into a root, ganglion, and nerve, a junction is effected between each posterior nerve and the corresponding anterior root. The junction is from the first at some little distance from the ganglion.

* * * * *

Investigators have hitherto described the spinal nerves as formed from part of the mesoblast of the protovertebræ. His alone, so far as I know, takes a different view.

His's[54] observations lead him to the conclusion that the posterior roots are developed as ingrowths from the external epiblast into the space between the protovertebræ and the neural canal. These subsequently become constricted off, unite with the neural canal and form spinal nerves.

These statements, which have not been since confirmed, diverge nearly to the same extent from my own results as does the ordinary account of the development of these parts.

Hensen (Virchow's Archiv, Vol. XXXI. 1864) also looks upon the spinal nerves as developed from the epiblast, but not as a direct result of his own observations[55].

Without attempting, for the present at least, to explain this divergence, I venture to think that the facts which I have just described have distinct bearings upon one or two important problems.

One point of general anatomy upon which they throw considerable light is the primitive origin of nerves.

So long as it was admitted that the spinal and cerebral nerves [Pg 190] developed in the embryo independently of the central nervous system, their mode of origin always presented to my mind considerable difficulties.

It never appeared clear how it was possible for a state of things to have arisen in which the central nervous system, as well as the peripheral terminations of nerves, whether motor or sensory, were formed independently of each other, while between them a third structure was developed which, growing in both directions (towards the centre and towards the periphery), ultimately brought the two into connexion.

That such a condition could be a primitive[TN7] one seemed scarcely possible.

Still more remarkable did it appear, on the supposition that the primitive mode of formation of these parts was represented in the developmental history of vertebrates, that we should find similar structural elements in the central and in the peripheral nervous systems.

The central nervous system arises from the epiblast, and yet contains precisely similar nerve-cells and nerve-fibres to the peripheral nervous system, which, if derived, as is usually stated, from the mesoblast, was necessarily supposed to have a completely different origin from the central nervous system.

Both of these difficulties are to a great extent removed by the facts of the development of these parts in Elasmobranchii.

If it be admitted that the spinal roots develop as outgrowths from the central nervous system in Elasmobranch Fishes, the question arises, how far can it be supposed to be possible that in other vertebrates the spinal roots and ganglia develop independently of the spinal cord, and only subsequently become united with it.

I have already insisted that this cannot be the primary condition; and though I am of opinion that the origin of the nerves in higher vertebrates ought to be worked over again, yet I do not think it impossible that, by a secondary adaptation, the nerve-roots might develop in the mesoblast[56].

[Pg 191] The presence of longitudinal commissures connecting the central ends of all the posterior roots is very peculiar. The commissures may possibly be looked on as outlying portions of the cord, rather than as parts of the nerves.

I have not up to this time followed their history beyond a somewhat early period in embryonic life, and am therefore unacquainted with their fate in the adult.

As far as I am aware, no trace of similar structures has been met with in other vertebrates.

The commissures have a very strong resemblance to those by which in Elasmobranch Fishes the glossopharyngeal nerve and the branches of the pneumogastric are united in an early embryonic stage[57].

I think it not impossible that the commissures in the two cases represent the same structures. If this is the case, it would seem that the junction of a number of nerves to form the pneumogastric is not a secondary state, but the remnant of a primary one, in which all the spinal nerves were united, as they embryonically are in Elasmobranchii.

One point brought out in my investigations appears to me to have bearings upon the origin of the central canal of the Vertebrate nervous system, and in consequence upon the origin of the Vertebrate group itself.

The point I allude to is the posterior nerve-rudiments making their first appearance at the extreme dorsal summit of the spinal cord.

The transverse section of the ventral nervous cord of an ordinary segmented worm consists of two symmetrical halves placed side by side.

If by a mechanical folding the two lateral halves of the nervous cord became bent towards each other, while into the groove formed between the two the external skin became pushed, we should have an approximation to the Vertebrate spinal cord. Such a folding might take place to give extra rigidity to the body in the absence of a vertebral column.

If this folding were then completed in such a way that the groove, lined by external skin and situated between the [Pg 192] two lateral columns of the nervous system, became converted into a canal, above and below which the two columns of the nervous system united, we should have in the transformed nervous cord an organ strongly resembling the spinal cord of Vertebrates.

This resemblance would even extend beyond mere external form. Let the ventral nervous cord of the common earthworm, Lumbricus agricola, be used for comparison[58], a transverse section of which is represented by Leydig[59] and Claparède. In this we find that on the ventral surface (the Annelidan ventral surface) of the nervous cord the ganglion-cells (grey matter) (k) are situated, and on the dorsal side the nerve-fibres or white matter (h). If the folding that I have supposed were to take place, the grey and white matters would have very nearly the relative situations which they have in the Vertebrate spinal cord.

The grey matter would be situated in the interior and surround the epithelium of the central canal, and the white matter would nearly surround the grey and form the anterior white commissure. The nerves would then arise, not from the sides of the nervous cord as in existing Vertebrates, but from its extreme ventral summit.

One of the most striking features which I have brought to light with reference to the development of the posterior roots, is the fact of their growing out from the extreme dorsal summit of the neural canal—a position analogous to the ventral summit of the Annelidan nervous cord. Thus the posterior roots of the nerves in Elasmobranchii arise in the exact manner which might have been anticipated were the spinal cord due to such a folding as I have suggested. The argument from the nerves becomes the stronger, from the great peculiarity in the position of the outgrowth, a feature which would be most perplexing without some such explanation as I have proposed. The central epithelium of the neural canal according to this view represents the external skin; and its ciliation is to be explained as a remnant of the ciliation of the external skin now found amongst many of the lower Annelids.

[Pg 193] I have, however, employed the comparison of the Vertebrate and Annelidan nervous cords, not so much to prove a genetic relation between the two as to shew the à priori possibility of the formation of a spinal canal and the à posteriori evidence we have of the Vertebrate spinal canal having been formed in the way indicated.

I have not made use of what is really the strongest argument for my view, viz. that the embryonic mode of formation of the spinal canal, by a folding in of the external epiblast, is the very method by which I have supposed the spinal canal to have been formed in the ancestors of Vertebrates.

My object has been to suggest a meaning for the peculiar primitive position of the posterior roots, rather than to attempt to explain in full the origin of the spinal canal.

EXPLANATION OF THE PLATES[60].

Plate 22.

Fig. A. Section through the dorsal region of an embryo of Scyllium stellare, with the rudiments of two visceral clefts. The section illustrates the general features at a period anterior to the appearance of the posterior nerve-roots.

nc. neural canal. mp. muscle-plate. ch. notochord. x. subnotochordal rod. ao. rudiment of dorsal aorta. so. somatopleure. sp. splanchnopleure. al. alimentary tract. All the parts of the section except the spinal cord are drawn somewhat diagrammatically.

Figs. B I, B II, B III. Three sections of a Pristiurus-embryo. B I is through the heart, B II through the anterior part of the dorsal region, and B III through a point slightly behind this. Drawn with a camera. (Zeiss CC ocul. 2.)

In B III there is visible a slight proliferation of cells from the dorsal summit of the neural canal.

In B II this proliferation definitely constitutes two club-shaped masses of cells (pr), both attached to the dorsal summit of the neural canal. The masses are the rudiments of the posterior nerve-roots.

In B I the rudiments of the posterior roots are of considerable length.

[Pg 194] pr. rudiment of posterior roots. nc. neural canal. mp. muscle-plate. ch. notochord. x. subnotochordal rod. ao. dorsal aorta. so. somatopleure. sp. splanchnopleure. al. alimentary canal. ht. heart.

Fig. C. Section from a Pristiurus-embryo, slightly older than B. Camera. (Zeiss CC ocul. 2.) The embryo from which this figure was taken was slightly distorted in the process of removal from the blastoderm.

vr. rudiment of vertebral body. Other reference letters as in previous figures.

Fig. D a. Section through the dorsal region of a Torpedo-embryo with three visceral clefts. (Zeiss CC ocul. 2.) The section shews the formation of the dorsal nerve-rudiments (pr) and of a ventral anterior nerve-rudiment (ar), which at this early stage is not distinctly cellular.

ar. rudiment of an anterior nerve-root. y. cells left behind on the separation of the external skin from the spinal cord. c. connective-tissue cells springing from the summit of the muscle-plates. Other reference letters as above.

Fig. D b. Section from dorsal region of a Torpedo-embryo somewhat older than D a. Camera. (Zeiss CC ocul. 2.) The posterior nerve-rudiment is considerably longer than in fig. Da, and its pedicle of attachment to the spinal cord is thinner. The anterior nerve-rudiment, of which only the edge is present in the section, is distinctly cellular.

m. mesoblast growing up from vertebral rudiment. sd. segmental duct.

Fig. D c. Section from a still older Torpedo-embryo. Camera. (Zeiss CC ocul. 2.) The connective-tissue cells are omitted. The rudiment of the ganglion (g) on the posterior root has appeared. The rudiment of the posterior nerve is much longer than before, and its junction with the spinal cord is difficult to detect. The anterior root is now an elongated cellular structure.

g. ganglion.

Fig. D d. Longitudinal and vertical section through a Torpedo-embryo of the same age as D c.

The section shews the commissures (x) uniting the posterior roots.

Fig. E a. Section of a Pristiurus-embryo belonging to the second stage. Camera. (Zeiss CC ocul. 2.) The section shews the constriction of the pedicle which attaches the posterior nerve-rudiments to the spinal cord.

pr. rudiment of posterior nerve-root. nc. neural canal. mp. muscle-plate. vr. vertebral rudiment. sd. segmental duct. ch. notochord. so. somatopleure. sp. splanchnopleure. ao. aorta. al. alimentary canal.

Fig. E b. Section of a Pristiurus-embryo slightly older than Ea. Camera. (Zeiss CC ocul. 2.) The section shews the formation of the anterior nerve-root (ar).

ar. rudiment of the anterior nerve-root.

Fig. F. Section of a Pristiurus-embryo with the rudiments of five visceral clefts. Camera. (Zeiss CC ocul. 2.)

The rudiment of the posterior root is seen surrounded by connective-tissue, from which it cannot easily be distinguished. The artist has not been very successful in rendering this figure.

[Pg 195] Figs. G1, G2, G3. Three longitudinal and horizontal sections of an embryo somewhat older than F. The embryo from which these sections were taken was hardened in osmic acid, but the sections have been represented without tinting. G1 is most dorsal of the three sections. Camera. (Zeiss CC ocul. 1.)

nc. neural canal. sp.c. spinal cord. pr. rudiment of posterior root. ar. rudiment of anterior root. mp. muscle-plate. c. connective-tissue cells. ch. notochord.

Plate 23.

Fig. H I. Section through the dorsal region of a Pristiurus-embryo in which the rudimentary external gills are present as very small knobs. Camera. (Zeiss CC ocul. 2.)

The section shews the commencing differentiation of the posterior nerve-rudiment into root (pr), ganglion (sp.g), and nerve (n), and also the attachment of the nerve-root to the spinal cord (x). The variations in the size and shape of the cells in the different parts of the nerve-rudiment are completely lost in the figure.

pr. posterior nerve-root. sp.g. ganglion of posterior root. n. nerve of posterior root. x. attachment of posterior root to spinal cord. w. white matter of spinal cord. i. mesoblastic investment to the spinal cord.

Fig. H II. Section through the same embryo as H I. (Zeiss CC ocul. 1.)

The section contains an anterior root, which takes its origin at a point opposite the interval between two posterior roots.

The white matter has not been very satisfactorily represented by the artist.

Figs. I I, I II. Two sections of a Pristiurus-embryo somewhat older than H. Camera. (Zeiss CC ocul. 1.)

The connective-tissue cells are omitted.

Figs. I a, I b, I c. Three isolated cells from the ganglion of one of the posterior roots of the same embryo.

Figs. K I, K II. Two horizontal longitudinal sections through an embryo in which the external gills have just appeared. K I is the most dorsal of the two sections. Camera. (Zeiss CC ocul. 1.)

The sections shew the relative positions of the anterior and posterior roots at different levels.

pr. posterior nerve-rudiment. ar. anterior nerve-rudiment. sp.c. spinal cord. n.c. neural canal. mp. muscle-plate. mp´. first-formed muscles.

Fig. L. Longitudinal and vertical section through the trunk of a Scyllium-embryo after the external gills have attained their full development. Camera. (Zeiss CC ocul. 1.)

The embryo was hardened in a mixture of chromic acid and osmic acid.

The section shews the commissures which dorsally unite the posterior roots, and also the junction of the anterior and posterior roots. The commissures are unfortunately not represented in the figure with great accuracy; their outlines are in nature perfectly regular, and not, as in the figure, notched at the junctions of the cells composing them. Their cells are apparently more or less completely fused, and certainly not nearly so clearly marked as in the figure. The commissures stain very deeply with the mixture of osmic and chromic acid, and form one of the most conspicuous [Pg 196] features in successful longitudinal sections of embryos so hardened. In sections hardened with chromic acid only they cannot be seen with the same facility.

sp.c. spinal cord. gr. grey matter. w. white matter. ar. anterior root. pr. posterior root. x. commissure uniting the posterior roots.

Figs. M I, M II. Two sections through the head of the same embryo as fig. B. M I, the foremost of the two, passes through the anterior part of the thickening of epiblast, which becomes involuted as the auditory vesicle. It contains the rudiment of the seventh nerve, VII. Camera. (Zeiss CC ocul. 2.)

VII. rudiment of seventh nerve. au. thickening of external epiblast, which becomes involuted as the auditory vesicle. n.c. neural canal. ch. notochord. pp. body-cavity in the head. so. somatopleure. sp. splanchnopleure. al. throat exhibiting an outgrowth to form the first visceral cleft.

[47] [From the Philosophical Transactions of the Royal Society of London, Vol. CLXVI. Pt. 1. Received October 5, Read December 16, 1875.]

[48] Vide Balfour, Preliminary account of the Development of Elasmobranch Fishes, Quart. Journ. of Microsc. Science, Oct. 1874, p. 33. [This edition, p. 96.]

[49] Vide Balfour, Origin and History of Urinogenital Organs of Vertebrates, Journal of Anatomy and Physiology, Oct. 1875. [This edition, No. VII.]

[50] This commissure is not satisfactorily represented in the figure. Vide Explanation of Plate 23.

[51] [May 18, 1876.—Observations I have recently made upon the development of the cranial nerves incline me to adopt an explanation of the change which takes place in the point of attachment of the spinal nerves to the cord differing from that enunciated in the text. I look upon this change as being apparent rather than real, and as due to a growth of the roof of the neural canal in the median dorsal line, which tends to separate the roots of the two sides more and more, and cause them to assume a more ventral position.]

[52] The artist has not been very successful in rendering this figure.

[53] [May 18, 1876.—Subsequent observations have led me to the conclusion that no anterior nerve-roots are to be found in the brain.]

[54] Erste Anlage des Wirbelthier-Leibes.

[55] [May 18, 1876.—Since the above was written Hensen has succeeded in shewing that in mammals the rudiments of the posterior roots arise in a manner closely resembling that described in the present paper; and I have myself, within the last few days, made observations which incline me to believe that the same holds good for the chick. My observations are as yet very incomplete.]

[56] [May 18, 1876.—Hensen's observations, as well as those recently made by myself on the chick, render it almost certain that the nerves in all Vertebrates spring from the spinal cord.]

[57] Balfour, A Preliminary Account of the Development of Elasmobranch Fishes, Q. J. Micros. Sc. 1874, plate XV. fig. 14, v.g. [This edition, Pl. 4, fig. 14, vg].

[58] The nervous cords of other Annelids resemble that of Lumbricus in the relations of the ganglion-cells of the nerve-fibres.

[59] Tafeln zur vergleichenden Anatomie, Taf. iii. fig. 8.

[60] The figures on these Plates give a fair general idea of the appearance presented by the developing spinal nerves; but the finer details of the original drawings have in several cases become lost in the process of copying.

The figures which are tinted represent sections of embryos hardened in osmic acid; those without colour sections of embryos hardened in chromic acid.

[Pg 197]

IX. On the Spinal Nerves of Amphioxus[61].

During a short visit to Naples in January last, I was enabled, through the kindness of Dr Dohrn, to make some observations on the spinal nerves of Amphioxus. These were commenced solely with the view of confirming the statements of Stieda on the anatomy of the spinal nerves, which, if correct, appeared to me to be of interest in connection with the observations I had made that, in Elasmobranchii, the anterior and posterior roots arise alternately and not in the same vertical plane. I have been led to conclusions on many points entirely opposed to those of Stieda, but, before recording these, I shall proceed briefly to state his results, and to examine how far they have been corroborated by subsequent observers.

Stieda[62], from an examination of sections and isolated spinal cords, has been led to the conclusion that, in Amphioxus, the nerves of the opposite sides arise alternately, except in the most anterior part of the body, where they arise opposite each other. He also states that the nerves of the same side issue alternately from the dorsal and ventral corners of the spinal cord. He regards two of these roots (dorsal and ventral) on the same side as together equivalent to a single spinal nerve of higher vertebrates formed by the coalescence of a dorsal and ventral root.

Langerhans[63] apparently agrees with Stieda as to the facts about the alternation of dorsal and ventral roots, but differs [Pg 198] from him as to the conclusions to be drawn from those facts. He does not, for two reasons, believe that two nerves of Amphioxus can be equivalent to a single nerve in higher vertebrates: (1) Because he finds no connecting branch between two succeeding nerves, and no trace of an anastomosis. (2) Because he finds that each nerve in Amphioxus supplies a complete myotome, and he considers it inadmissible to regard the nerves, which in Amphioxus together supply two myotomes, as equivalent to those which in higher vertebrates supply a single myotome only.

Although the agreement as to facts between Langerhans and Stieda is apparently a complete one, yet a critical examination of the statements of these two authors proves that their results, on one important point at least, are absolutely contradictory. Stieda, Pl. III. fig. 19, represents a longitudinal and horizontal section through the spinal cord which exhibits the nerves arising alternately on the two sides, and represents each myotome supplied by one nerve. In his explanation of the figure he expressly states that the nerves of one plane only (i.e. only those with dorsal or only those with ventral roots) are represented; so that if all the nerves which issue from the spinal cord had been represented double the number figured must have been present. But since each myotome is supplied by one nerve in the figure, if all the nerves present were represented, each myotome would be supplied by two nerves.

Since Langerhans most emphatically states that only one nerve is present for each myotome, it necessarily follows that he or Stieda has made an important error; and it is not too much to say that this error is more than sufficient to counterbalance the value of Langerhans' evidence as a confirmation of Stieda's statements.

I commenced my investigations by completely isolating the nervous system of Amphioxus by maceration in nitric acid according to the method recommended by Langerhans[64]. On examining specimens so obtained it appeared that, for the greater length of the cord, the nerves arose alternately on the [Pg 199] two sides, as was first stated by Owsjannikow, and subsequently by Stieda and Langerhans; but to my surprise not a trace could be seen of a difference of level in the origin of the nerves of the same side.

The more carefully the specimens were examined from all points of view, the more certainly was the conclusion forced upon me, that nerves issuing from the ventral corner of the spinal cord, as described by Stieda, had no existence.

Not satisfied by this examination, I also tested the point by means of sections. I carefully made transverse sections of a successfully hardened Amphioxus, through the whole length of the body. There was no difficulty in seeing the dorsal roots in every third section or so, but not a trace of a ventral root was to be seen. There can, I think, be no doubt, that, had ventral roots been present, they must, in some cases at least, have been visible in my sections.

In dealing with questions of this kind it is no doubt difficult to prove a negative; but, since the two methods of investigation employed by me both lead to the same result, I am able to state with considerable confidence that my observations lend no support to the view that the alternate spinal nerves of Amphioxus have their roots attached to the ventral corner of the spinal cord.

How a mistake on this point arose it is not easy to say. All who have worked with Amphioxus must be aware how difficult it is to conserve the animal in a satisfactory state for making sections. The spinal cord, especially, is apt to be distorted in shape, and one of its ventral corners is frequently produced into a horn-like projection terminating in close contact with the sheath. In such cases the connective tissue fibres of the sheath frequently present the appearance of a nerve-like prolongation of the cord; and for such they might be mistaken if the sections were examined in a superficial manner. It is not, however, easy to believe that, with well conserved specimens, a mistake could be made on this point by so careful and able an investigator as Stieda, especially considering that the histological structure of the spinal nerves is very different from that of the fibrous prolongations of the sheath of the spinal cord.

[Pg 200] It only remains for me to suppose that the specimens which Stieda had at his disposal, were so shrunk as to render the origin of the nerves very difficult to determine.

The arrangement of the nerves of Amphioxus, according to my own observations, is as follows.

The anterior end of the central nervous system presents on its left and dorsal side a small pointed projection, into which is prolonged a diverticulum from the dilated anterior ventricle of the brain. This may perhaps be called the olfactory nerve, though clearly of a different character to the other nerves. It was first accurately described by Langerhans[65].

Vertically below the olfactory nerve there arise two nerves, which issue at the same level from the ventral side of the anterior extremity of the central nervous system. These form the first pair of nerves, and are the only pair which arise from the ventral portion of the cerebro-spinal cord. The two nerves, which form the second pair, arise also opposite each other but from the dorsal side of the cord. The first and second pair of nerves have both been accurately drawn and described by Langerhans: they, together with the olfactory nerve, can easily be seen in nervous systems which have been isolated by maceration.

In the case of the third pair of nerves, the nerve on the right-hand side is situated not quite opposite but slightly behind that on the left. The right nerve of the fourth pair is situated still more behind the left, and, in the case of the fifth pair, the nerve to the right is situated so far behind the left nerve that it occupies a position half-way between the left nerves of the fifth and sixth pairs. In all succeeding nerves the same arrangement holds good, so that they exactly alternate on two sides.

Such is the arrangement carefully determined by me from one specimen. It is possible that it may not be absolutely constant, but the following general statement almost certainly holds good.

All the nerves of Amphioxus, except the first pair, have their roots inserted in the dorsal part of the cord. In the case of [Pg 201] the first two pairs the nerves of the two sides arise opposite each other; in the next few pairs, the nerves on the right-hand side gradually shift backwards: the remaining nerves spring alternately from the two sides of the cord.

For each myotome there is a single nerve, which enters, as in the case of other fishes, the intermuscular septum. This point may easily be determined by means of longitudinal sections, or less easily from an examination of macerated specimens. I agree with Langerhans in denying the existence of ganglia on the roots of the nerves.

[61] From the Journal of Anatomy and Physiology, Vol. X. 1876.

[62] Mém. Acad. Pétersbourg, Vol. XIX.

[63] Archiv f. mikr. Anatomie, Vol. XII.

[64] Loc. cit.

[65] Loc. cit.

[Pg 202]
[Pg 203]

X.

A MONOGRAPH

ON THE

DEVELOPMENT OF
ELASMOBRANCH FISHES.

Published 1878.

[Pg 204]
[Pg 205]

PREFACE.

The present Monograph is a reprint of a series of papers published in the Journal of Anatomy and Physiology during the years 1876, 1877 and 1878. The successive parts were struck off as they appeared, so that the earlier pages of the work were in print fully two years ago. I trust the reader will find in this fact a sufficient excuse for a certain want of coherence, which is I fear observable, as well as for the omission of references to several recent publications. The first and second chapters would not have appeared in their present form had I been acquainted, at the time of writing them, with the researches which have since been published, on the behaviour of the germinal vesicle and on the division of nuclei. I may also call attention to the valuable papers of Prof. His[66] on the formation of the layers in Elasmobranchii, and of Prof. Kowalevsky[67] on the development of Amphioxus, to both of which I would certainly have referred, had it been possible for me to do so.

Professor His deals mainly with the subjects treated of in Chapter III., and gives a description very similar to my own of the early stages of development. His interpretations of the observed changes are, however, very different from those at which I have arrived. Although this is not the place for a discussion of Prof. His's views, I may perhaps state that, in spite of the arguments he has brought forward in support of his position, I am still inclined to maintain the accuracy of my original account. The very striking paper on Amphioxus by Kowalevsky (the substance of which I understand to have been published in Russia at an earlier period) contains a confirmation of the views expressed in chapter VI. on the development [Pg 206] of the mesoblast, and must be regarded as affording a conclusive demonstration, that in the case of Vertebrata the mesoblast has primitively the form of a pair of diverticula from the walls of the archenteron.

* * * * *

The present Memoir, while differing essentially in scope and object from the two important treatises by Professors His[68] and Götte[69], which have recently appeared in Germany, has this much in common with them, that it deals monographically with the development of a single type: but here the resemblance ends. Both of these authors seek to establish, by a careful investigation of the development of a single species, the general plan of development of Vertebrates in general, if not of the whole animal kingdom. Both reject the theory of descent, as propounded by Mr Darwin, and offer completely fresh explanations of the phenomena of Embryology. Accepting, as I do, the principle of natural selection, I have had before me, in writing the Monograph, no such ambitious aim as the establishment of a completely new system of Morphology. My object will have been fully attained if I have succeeded in adding a few stones to the edifice, the foundations of which were laid by Mr Darwin in his work on the Origin of Species.

I may perhaps call attention to one or two special points in this work which seem to give promise of further results. The chapter on the Development of the Spinal and Cranial Nerves contains a modification of the previously accepted views on this subject, which may perhaps lead to a more satisfactory conception of the origin of nerves than has before been possible, and a more accurate account of the origin of the muscle-plates and vertebral column. The attempt to employ the embryological relations of the cephalic prolongations of the body-cavity, and of the cranial nerves, in the solution of the difficult problems of the Morphology of the head, may prove of use in the line of study so successfully cultivated by our great English Anatomist, Professor Huxley. Lastly, I venture to hope that my conclusions in reference to the relations of the sympathetic system and the suprarenal body, and to the development of the mesoblast, [Pg 207] the notochord, the limbs, the heart, the venous system, and the excretory organs, are not unworthy of the attention of Morphologists.

* * * * *

The masterly manner in which the systematic position of Elasmobranchii is discussed by Professor Gegenbaur, in the introduction to his Monograph on the Cranial Skeleton of the group, relieves me from the necessity of entering upon this complicated question. It is sufficient for my purpose that the Elasmobranch Fishes be regarded as forming one of the most primitive groups among Vertebrates, a view which finds ample confirmation in the importance of the results to which Prof. Gegenbaur and his pupils have been led in this branch of their investigations.

* * * * *

Though I trust that the necessary references to previous contributions in the same department of enquiry have not been omitted, the 'literature of the subject' will nevertheless be found to occupy a far smaller share of space than is usual in works of a similar character. This is an intentional protest on my part against, what appears to me, the unreasonable amount of space so frequently occupied in this way. The pages devoted to the 'previous literature' only weary the reader, who is not wise enough to skip them, and involve a great and useless expenditure of time on the part of any writer, who is capable of something better than the compilation of abstracts.

* * * * *

In conclusion, my best thanks are due to Drs Dohrn and Eisig for the uniformly kind manner in which they have forwarded my researches both at the Zoological Station in Naples, and after my return to England; and also to Mr Henry Lee and to the Manager and Directors of the Brighton Aquarium, who have always been ready to respond to my numerous demands on their liberality.

To my friend and former teacher Dr Michael Foster I tender my sincerest thanks for the never-failing advice and assistance which he has given throughout the whole course of the work.

[66] Zeitschrift f. Anat. u. Entwicklungsgeschichte, Bd. II.

[67] Archiv f. Micr. Anat. Bd. XIII.

[68] Erste Anlage des Wirbelthierleibes.

[69] Entwicklungsgeschichte der Unke.

[Pg 208]
[Pg 209]

[Pg 213]

CHAPTER I.

The Ripe Ovarian Ovum.

The ripe ovum is nearly spherical, and, after the removal of its capsule, is found to be unprovided with any form of protecting membrane.

My investigations on the histology of the ripe ovarian ovum have been made with the ova of the Gray Skate (Raja batis) only, and owing to a deficiency of material are somewhat imperfect.

The bulk of the ovum is composed of yolk spherules, imbedded in a protoplasmic matrix. Dr Alexander Schultz[70], who has studied with great care the constitution of the yolk, finds, near the centre of the ovum, a kernel of small yolk spherules, which is succeeded by a zone of spherules which gradually increase in size as they approach the surface. But, near the surface, he finds a layer in which they again diminish in size and exhibit numerous transitional forms on the way to molecular yolk granules. These Dr Schultz regards as in a retrogressive condition.

Another interesting feature about the yolk is the presence in it of a protoplasmic network. Dr Schultz has completely confirmed, and on some points enlarged, my previous observations on this subject[71]. Dr Schultz's confirmation is the more important, since he appears to be unacquainted with my previous investigations. In my paper (loc. cit.), after giving a description of the network I make the following statement as to its distribution.

[Pg 214] A specimen of this kind is represented in Plate 14, fig. 2, ny, where the meshes of the network are seen to be finer immediately around the nuclei, and coarser in the intervals. The specimen further shews, in the clearest manner, that this network is not divided into areas, each representing a cell and each containing a nucleus. I do not know to what extent this network extends into the yolk. I have never yet seen the limits of it, though it is very common to see the coarsest yolk-granules lying in its meshes. Some of these are shewn in Plate 14, fig. 2, y.k. [This edition, p. 65.]

Dr Schultz, by employing special methods of hardening and cutting sections of the whole egg, has been able to shew that this network extends, in the form of fine radial lines, from the centre to the circumference; and he rightly states, that it exhibits no cell-like structures. I have detected this network extending throughout the whole yolk in young eggs, but have failed to see it with the distinctness which Dr Schultz attributes to it in the ripe ovum. Since it is my intention to enter fully both into the structure and meaning of this network in my account of a later stage, I say no more about it here.

At one pole of the ripe ovum a slight examination demonstrates the presence of a small circular spot, sharply distinguished from the remainder of the yolk by its lighter colour. Around this spot is an area which is also of a lighter colour than the yolk, and the outer border of which gradually shades into the normal tint of the yolk. If a section be made through this part (vide Pl. 6, fig. 1) the circular spot will be found to be the germinal vesicle, and the area around it a disc of yolk containing smaller spherules than the surrounding parts. The germinal vesicle possessed the same structure in both the ripe eggs examined by me; and, in both, it was situated quite on the external surface of the yolk.

In one of my specimens it was flat above, but convex below; in the other and, on the whole, the better preserved of the two, it had the somewhat quadrangular but rather irregular section represented in Pl. 6, fig. 1. It consisted of a thickish membrane and its primitive contents. The membrane surrounded the upper part of the contents and exhibited numerous folds and creases (vide fig. 1). As it extended downwards it became thinner, and completely disappeared at some little distance from the lower end of the contents. These, therefore, rested below on the yolk. At its circumference the membrane of the disc was [Pg 215] produced into a kind of fold, forming a rim which rested on the surface of the yolk.

In neither of my specimens is the cavity in the upper part of the membrane filled by the contents; and the upper part of the membrane is so folded and creased that sections through almost any portion of it pass through the folds. The regularity of the surface of the yolk is not broken by the germinal vesicle, and the yolk around exhibits not the slightest signs of displacement. In the germinal vesicle figured the contents are somewhat irregular in shape; but in my other specimen they form a regular mass concave above and convex below. In both cases they rest on the yolk, and the floor of the yolk is exactly moulded to suit the surface of the contents of the germinal vesicle. The contents have a granular aspect, but differ in constitution from the surrounding yolk. Each germinal vesicle measured about one-fiftieth of an inch in diameter.

It does not appear to me possible to suppose that the peculiar appearances which I have drawn and described are to be looked upon as artificial products either of the chromic acid, in which the ova were hardened, or of the instrument with which sections of them were made. It is hardly conceivable that chromic acid could cause a rupture of the membrane and the ejection of the contents of the vesicle. At the same time the uniformity of the appearances in the different sections, the regularity of the whole outline of the egg, and the absence of any signs of disturbance in the yolk, render it impossible to believe that the structures described are due to faults of manipulation during or before the cutting of the sections.

We can only therefore conclude that they represent the real state of the germinal vesicle at this period. No doubt they alone do not supply a sufficient basis for any firm conclusions as to the fate of the germinal vesicle. Still, if they cannot sustain, they unquestionably support certain views. The natural interpretation of them is that the membrane of the germinal vesicle is in the act of commencing to atrophy, preparatory to being extruded from the egg, while the contents of the germinal vesicle are about to be absorbed.

In favour of the extrusion of the membrane rather than its absorption are the following features:

[Pg 216] (1) The thickness of its upper surface. (2) The extension of its edge over the yolk. (3) Its position external to the yolk.

In favour of the view that the contents will be left behind and absorbed when the membrane is pushed out, are the following features of my sections:

(1) The rupture of the membrane of the germinal vesicle on its lower surface. (2) The position of the contents almost completely below the membrane of the vesicle and surrounded by yolk.

In connection with this subject, Oellacher's valuable observations upon the behaviour of the germinal vesicle in Osseous Fishes and in Birds at once suggest themselves[72]. Oellacher sums up his results upon the behaviour of the germinal vesicle in Osseous Fishes in the following way (p. 12):

The germinal vesicle of the Trout's egg, at a period when the egg is very nearly ripe, lies near the surface of the germinal disc which is aggregated together in a hollow of the yolk.... After this a hole appears in the membrane of the germinal vesicle, which opens into the space between the egg-membrane and the germinal disc. The hole widens more and more, and the membrane frees itself little by little from the contents of the germinal vesicle, which remain behind in the form of a ball on the floor of the cavity formed in this way. The cavity becomes flatter and flatter and the contents are pushed up further and further from the germinal disc. When the hollow, in which lie the contents of the original germinal vesicle, completely vanishes, the covering membrane becomes inverted ... and the membrane is spread out on the convex surface of the germinal disc as a circular, investing structure. It is clear that by the removal of the membrane the contents of the germinal vesicle become lost.

These very definite statements of Oellacher tell strongly against my interpretation of the appearance presented by the germinal vesicle of the ripe Skate's egg. Oellacher's account is so precise, and his drawings so fully bear out his interpretations, that it is very difficult to see where any error can have crept in.

On the other hand, with the exception of those which Oellacher has made, there cannot be said to be any satisfactory observations demonstrating the extrusion of the germinal vesicle from the ovum. Oellacher has observed this definitely for the Trout, but his observations upon the same point in the Bird would quite as well bear the interpretation that the membrane alone became pushed out, as that this occurred to the germinal vesicle, contents and all.

[Pg 217] While, then, there are on the one hand Oellacher's observations on a single animal, hitherto unconfirmed, there are on the other very definite observations tending to shew that the germinal vesicle has in many cases an altogether different fate. Götte[73], not to mention other observers before him, has in the case of Batrachian's eggs traced out with great precision the gradual atrophy of the germinal vesicle, and its final absorption into the matter of the ovum.

Götte distinguishes three stages in the degeneration of the germinal vesicle of Bombinator's egg. In the first stage the germinal vesicle has begun to travel up towards the surface of the egg. It retains nearly its primitive condition, but its contents have become more opaque and have partly withdrawn themselves from the thin membrane. The germinal spots are still circular, but in some cases have increased in size. The most important feature of this stage is the smaller size of the germinal vesicle than that of the cavity of the yolk in which it lies, a condition which appears to demonstrate the commencing atrophy of the vesicle.

In the next stage the cavity containing the germinal vesicle has vanished without leaving a trace. The germinal vesicle itself has assumed a lens-like form, and its borders are irregular and pressed in here and there by yolk. Of the membrane of the germinal vesicle, and of the germinal spots, only scanty remnants are to be seen, many of which lie in the immediately adjoining yolk.

In the last stage no further trace of a distinct germinal vesicle is present. In its place is a mass of very finely granular matter, which is without a distinct border and graduates into the surrounding yolk and is to be looked on as a remnant of the germinal vesicle.

This careful investigation of Götte proves beyond a doubt that in Batrachians neither the membrane, nor the contents of the germinal vesicle, are extruded from the egg.

In Mammalia, Van Beneden[74] finds that the germinal vesicle becomes invisible, though he does not consider that it absolutely ceases to exist. He has not traced the steps of the process with the same care as Götte, but it is difficult to believe that an [Pg 218] extrusion of the vesicle in the way described by Oellacher would have escaped his notice.

Passing from Vertebrates to Invertebrates, we find that almost every careful investigator has observed the disappearance, apparent or otherwise, of the germinal vesicle, but that very few have watched with care the steps of the process.

The so-called Richtungskörper has been supposed to be the extruded remnant of the germinal vesicle. This view has been especially adopted and supported by Oellacher (loc. cit.), and Flemming[75].

The latter author regards the constant presence of this body, and the facility with which it can be stained, as proofs of its connection with the germinal vesicle, which has, however, according to his observations, disappeared before the appearance of the Richtungskörper.

Kleinenberg[76], to whom we are indebted for the most precise observations we possess on the disappearance of the germinal vesicle, gives the following account of it, pp. 41 and 42.

"We left the germinal vesicle as a vesicle with a distinct doubly contoured membrane, and equally distributed granular contents, in which the germinal spot had appeared.... The germinal vesicle reaches 0.06 mm. in diameter, and at the same time its contents undergo a separation. The greater part withdraws itself from the membrane and collects as a dense mass around the germinal spot, while closely adjoining the membrane there remains only a very thin but unbroken lining of the plasmoid material. The intermediate space is filled with a clear fluid, but the layer which lines the membrane retains its connection with the mass around the germinal vesicle by means of numerous fine threads which traverse the space filled with fluid.... At about the time when the formation of the pseudocells in the egg is completed the germinal spot undergoes a retrogressive metamorphosis, it loses its circular outline and it now appears as if coagulated; then it breaks up into small fragments, and I am fairly confident that these become dissolved. The germinal vesicle ... becomes, on the egg assuming a spherical form, drawn into an eccentric position towards the pole of the egg directed outwards, where it lies close to the surface and only covered by a very thin layer of plasma. In this situation its degeneration now begins, and ends in its complete disappearance. The granular contents become more and more fluid; at the same time part of them pass out through the membrane. This, which so far was firmly stretched, next collapses to a somewhat egg-like sac, whose wall is thickened and in places folded.

[Pg 219]The inner mass which up to this time has remained compact now breaks up into separate highly refractive bodies, of spherical or angular form and of very different sizes; between them, here and there, are scattered drops of a fluid fat.... I am very much inclined to regard the solid bodies in question as fat or as that peculiar modification of albuminoid bodies which we recognise as the certain forerunner of the formation of fat in so many pathologically altered tissues; and therefore to refer the disappearance of the germinal vesicle to a fatty degeneration. On one occasion I believe that I observed an opening in the membrane at this stage; if this is a normal condition it would be possible to believe that its solid contents passed out and were taken up in the surrounding plasma. What becomes of the membrane I am unable to say; in any case the germinal vesicle has vanished to the very last trace before impregnation occurs.

Kleinenberg clearly finds that the germinal vesicle disappears completely before the appearance of the Richtungskörper, in which he states a pseudocell or yolk-sphere is usually found.

The connection between the Richtungskörper and the germinal vesicle is not a result of strict observation, and there can be no question that the evidence in the case of invertebrates tends to prove that the germinal vesicle in no case disappears owing to its extrusion from the egg, but that if part of it is extruded from the egg as Richtungskörper this occurs when its constituents can no longer be distinguished from the remainder of the yolk. This is clearly the case in Hydra, where, as stated above, one of the pseudocells or yolk-spheres is usually found imbedded in the Richtungskörper.

My observations on the Skate tend to shew that, in its case, the membrane of the germinal vesicle is extruded from the egg, though they do not certainly prove this. That conclusion is however supported by the observations of Schenk[77]. He found in the impregnated, but not yet segmented, germinal disc a cavity which, as he suggests, might well have been occupied by the germinal vesicle. It is not unreasonable to suppose that the membrane, being composed of formed matter and able only to take a passive share in vital functions, could, without thereby influencing the constitution of the ovum, be ejected.

If we suppose, and this is not contradicted by observation, that the Richtungskörper is either only the metamorphosed membrane of the germinal vesicle with parts of the yolk, or part of the yolk alone, and assume that in Oellacher's observations [Pg 220] only the membrane and not the contents were extruded from the egg, it would be possible to frame a consistent account of the behaviour of the germinal vesicle throughout the animal kingdom, which may be stated in the following way.

The germinal vesicle usually before, but sometimes immediately after impregnation undergoes atrophy and its contents become indistinguishable from the remainder of the egg. In those cases in which its membrane is very thick and resistent, e.g. Osseous and Elasmobranch Fishes, Birds, etc., this may be incapable of complete resorption, and be extruded bodily from the egg. In the case of most ova, it is completely absorbed, though at a subsequent period it may be extruded from the egg as the Richtungskörper. In all cases the contents of the germinal vesicle remain in the ovum.

In some cases the germinal vesicle is stated to persist and to undergo division during the process of segmentation; but the observations on this point stand in need of confirmation.

My investigations shew that the germinal vesicle atrophies in the Skate before impregnation, and in this respect accord with very many recent observations. Of these the following may be mentioned.

(1) Oellacher (Bird, Osseous Fish). (2) Götte (Bombinator igneus). (3) Kupffer (Ascidia canina). (4) Strasburger (Phallusia mamillata). (5) Kleinenberg (Hydra). (6) Metschnikoff (Geryonia, Polyzenia leucostyla, Epibulia aurantiaca, and other Hydrozoa).

This list is sufficient to shew that the disappearance of the germinal vesicle before impregnation is very common, and I am unacquainted with any observations tending to shew that its disappearance is due to impregnation.

In some cases, e.g. Asterocanthion[78], the germinal vesicle vanishes after the spermatozoa have begun to surround the egg; but I do not know that its disappearance in these cases has been shewn to be due to impregnation. To do so it would be necessary to prove that in ripe eggs let loose from the ovary, but not fertilized, the germinal vesicle did not undergo the same changes as in the case of fertilized eggs; and this, as far as I [Pg 221] know, has not been done. After the disappearance of the germinal vesicle, and before the first act of division, a fresh nucleus frequently appears [—vide—Auerbach (Ascaris nigrovenosa), Fol (Geryonia), Kupffer (Ascidia canina), Strasburger (Phallusia mamillata), Flemming (Anodon), Götte (Bombinator igneus)], which is generally stated to vanish before the appearance of the first furrow; but in some cases (Kupffer and Götte, and as studied with especial care, Strasburger) it is stated to divide. Upon the second nucleus, or upon its relation to the germinal vesicle, I have no observations; but it appears to me of great importance to determine whether this fresh nucleus arises absolutely de novo, or is formed out of the matter of the germinal vesicle.

The germinal vesicle is situated in a bed of finely divided yolk-particles. These graduate insensibly into the coarser yolk-spherules around them, though the band of passage between the coarse and the finer yolk-particles is rather narrow. The mass of fine yolk-granules may be called the germinal disc. It is not to be looked upon as diverging in any essential particular from the remainder of the yolk, for the difference between the two is one of degree only. It contains in fact a larger bulk of active protoplasm, as compared with yolk-granules, than does the remainder of the ovum. The existence of this agreement in kind has been already strongly insisted on in my preliminary paper; and Schultz (loc. cit.) has arrived at an entirely similar conclusion, from his own independent observations.

One interesting feature about the germinal disc at this period is its size.

My observations upon it have been made with the eggs of the Skate (Raja) alone; but I think that it is not probable that its size in the Skate is greater than in Scyllium or Pristiurus. If its size is the same in all these genera, then the germinal disc of the unimpregnated ovum is very much greater than that portion of the ovum which undergoes segmentation, and which is usually spoken of as the germinal disc in impregnated ova.

I have no further observation on the ripe ovarian ovum; and my next observations concern an ovum in which two furrows have already appeared.

[70] Archiv für Micro. Anat. Vol. XI. 1875.

[71] Quart. Journ. Micro. Science, Oct. 1874. [This edition, No. V.]

[72] Archiv für Micr. Anat. Vol. VIII. p. 1.

[73] Entwicklungsgeschichte der Unke.

[74] Recherches sur la Composition et la Signification de l'Œuf.

[75] Studien in der Entwicklungsgeschichte der Najaden, Sitz. d. k. Akad. Wien, Bd. LXXI. 1875.

[76] Hydra. Leipzig, 1872.

[77] Die Eier von Raja quadrimaculata, Sitz. der k. Akad. Wien, Bd. LXVIII.

[78] Agassiz, Embryology of the Star-Fish.

[Pg 222]

CHAPTER II.

The Segmentation.

I have not been fortunate enough to obtain an absolutely complete series of eggs during segmentation.

In the cases of Pristiurus and Scyllium only have I had any considerable number of eggs in this condition, though one or two eggs of Raja in which the process was not completed have come into my hands.

In the youngest impregnated Pristiurus eggs, which I have obtained, the germinal disc was already divided into four segments.

The external appearance of the blastoderm, which remains nearly constant during segmentation, has been already well described by Leydig[79].

The yolk has a pale greenish tinge which, on exposure to the air, acquires a yellower hue. The true germinal disc appears as a circular spot of a bright orange colour, and is, according to Leydig's measurements, 1½m. in diameter. Its colour renders it very conspicuous, a feature which is further increased by its being surrounded by a narrow dark line (Pl. 6, fig. 2), the indication of a shallow groove. Surrounding this line is a concentric space which is lighter in colour than the remainder of the yolk, but whose outer border passes by insensible gradations into the yolk. As was mentioned in my preliminary paper (loc. cit.), and as Leydig (loc. cit.) had before noticed, the germinal disc is always situated at the pole of the yolk which is near the rounded end of the Pristiurus egg. It occupies a corresponding position in the eggs of both species of Scyllium (stellare and canicula) near the narrower end of the egg to which the shorter pair of strings is attached. The germinal disc in the youngest egg [Pg 223] examined, exhibited two furrows which crossed each other at right angles in the centre of the disc, but neither of which reached its edge. These furrows accordingly divided the disc into four segments, completely separated from each other at the centre of the disc, but united near its circumference.

I made sections, though not very satisfactorily, of this germinal disc. The sections shewed that the disc was composed of a protoplasmic basis, in which were imbedded innumerable minute spherical yolk-globules so closely packed as to constitute nearly the whole mass of the germinal disc.

In passing from the coarsest yolk-spheres to the fine spherules of the germinal disc, three bands of different-sized yolk-particles have to be traversed. These bands graduate into one another and are without sharp lines of demarcation. The outer of the three is composed of the largest-sized yolk-spherules which constitute the greater part of the ovum. The middle band forms a concentric layer around the germinal disc, and is composed of yolk-spheres considerably smaller than those outside it. Where it cuts the surface it forms the zone of lighter colour immediately surrounding the germinal disc. The innermost band is formed by the germinal disc itself and is composed of spherules of the smallest size. These features are shewn in Pl. 6, fig. 6, which is the section of a germinal disc with twenty-one segments; in it however the outermost band of spherules is not present.

From this description it is clear, as has already been mentioned in the description of the ripe unimpregnated ovum, that the germinal disc is not to be looked upon as a body entirely distinct from the remainder of the ovum, but merely as a part of the ovum in which the protoplasm is more concentrated and the yolk-spherules smaller than elsewhere. Sections shew that the furrows visible on the surface end below, as indeed they do on the surface, before they reach the external limit of the finely granular matter of the germinal disc. There are therefore at this stage no distinct segments: the otherwise intact germinal disc is merely grooved by two furrows.

I failed to observe any nuclei in the germinal disc just described, but it by no means follows that they were not present.

[Pg 224] In the next youngest of the eggs[80] examined the germinal disc was already divided into twenty-one segments. When viewed from the surface (Pl. 6, fig. 3), the segments appeared divided into two distinct groups—an inner group of eleven smaller segments, and an outer group of segments surrounding the former. The segments of both the inner and the outer group were very irregular in shape and varied considerably in size. The amount of irregularity is far from constant and many germinal discs are more regular than the one figured.

In this case the situation of the germinal disc and its relations to the yolk were precisely the same as in the earlier stage.

In sections of this germinal disc (Pl. 6, fig. 6), the groove which separates it from the yolk is well marked on one side, but hardly visible at the other extremity of the section.

Passing from the external features of this stage to those which are displayed by sections, the striking point to be noticed is the persisting continuity of the segments, marked out on the surface, with the floor of the germinal disc.

The furrows which are visible on the surface merely form a pattern, but do not isolate a series of distinct segments. They do not even extend to the limit of the finely granular matter of the germinal disc.

The section represented, Pl. 6, fig. 6, bears out the statements about the segments as seen on the surface. There are three smaller segments in the middle of the section, and two larger at the two ends. These latter are continuous with the coarser yolk-spheres surrounding the germinal disc and are not separated from them by a segmentation furrow.

In a slightly older embryo than the one figured I met with a few completely isolated segments at the surface. These segments were formed by the apparent bifurcation of furrows as they neared the surface of the germinal disc. The segments thus produced are triangular in form. They probably owe their origin to the meeting of two oblique furrows. The last-formed of these furrows apparently ceases to be prolonged after meeting the first-formed furrow. I have not in any case [Pg 225] observed an example of two furrows crossing one another at this stage.

The furrows themselves for the most part are by no means simple slits with parallel sides. They exhibit a beaded structure, shewn imperfectly in Pl. 6, fig. 6, but better in Pl. 6, fig. 6a, which is executed on a larger scale. They present intervals of dilatations where the protoplasms of the segments on the two sides of the furrow are widely separated, alternating with intervals where the protoplasms of the two segments are almost in contact and are only separated from one another by a very narrow space.

A closer study of the germinal disc at this period shews that the cavities which cause the beaded structure of the furrows are not only present along the lines of the furrows but are also found scattered generally through the germinal disc, though far more thickly in the neighbourhood of the furrows. Their appearance is that of vacuoles, and with these they are probably to be compared. There can be little question that in the living germinal disc they are filled with fluid. In some cases, they are collected in very large numbers in the region of a furrow. Such a case as this is shewn in Pl. 6, fig. 6b. In numerous other cases they occur, roughly speaking, alternately on each side of a furrow. Some furrows, though not many, are entirely destitute of these structures. The character of their distribution renders it impossible to overlook the fact that these vacuole-like bodies have important relations with the formation of the segmentation furrows.

Lining the two sides of the segmentation furrows there is present in sections a layer which stains deeply with colouring reagents; and the surface of the blastoderm is stained in the same manner. In neither case is it permissible to suppose that any membrane-like structure is present. In many cases a similar very delicate, but deeply-stained line, invests the vacuolar cavities, but the fluid filling these remains quite unstained. When distinct segments are formed, each of these is surrounded by a similarly stained line.

The yolk-spherules are so numerous, and render even the thinnest section so opaque, that I have failed to make satisfactory observations on the behaviour of the nucleus. I find [Pg 226] nuclei in many of the segments, though it is very difficult even to see them, and only in very favourable specimens can their structure be studied. In some cases, two of them lie one on each side of a furrow; and in one case at the extreme end of a furrow I could see two peculiar aggregations of yolk-spherules united by a band through which the furrow, had it been continued, would have passed. The connection (if any exists) between this appearance and the formation of the fresh nuclei in the segments, I have been unable to elucidate.

The peculiar appearances attending the formation of fresh nuclei in connection with cell-division, which have recently been described by so many observers, have hitherto escaped my observation at this stage of the segmentation, though I shall describe them in a later stage. A nucleus of this stage is shewn on Pl. 6, fig. 6c. It is lobate in form and is divided by lines into areas in each of which a deeply-stained granule is situated.

The succeeding stages of segmentation present from the surface no fresh features of great interest. The somewhat irregular (Pl. 6, figs. 4 and 5) circular line, which divides the peripheral larger from the central smaller segments, remains for a long time conspicuous. It appears to be the representative of the horizontal furrow which, in the Batrachian ovum, separates the smaller pigmented spheres from the larger spheres of the lower pole of the egg.

As the segments become smaller and smaller, the distinction between the peripheral and the central segments becomes less and less marked; but it has not disappeared by the time that the segments become too small to be seen with the simple lens. When the spheres become smaller than in the germinal disc represented on Pl. 6, fig. 5, the features of segmentation can be more easily and more satisfactorily studied by means of sections.

To the features presented in sections, both of the latter and of the earlier blastoderms, I now return. A section of one of the earlier germinal discs, of about the age of the one represented on Pl. 6, fig. 4, is shewn in Pl. 6, fig. 7.

It is clear at a glance that we are now dealing with true segments completely circumscribed on all sides. The peripheral [Pg 227] segments are, as a rule, larger than the more central ones, though in this respect there is considerable irregularity. The segments are becoming smaller by repeated division; but, in addition to this mode of increase, there is now going on outside the germinal disc a segmentation of the yolk, by which fresh segments are being formed from the yolk and added to those which already exist in the germinal disc. One or two such segments are seen in the act of being formed (Pl. 6, fig. 7, f); and it is to be noticed that the furrows which will eventually mark out the segments, do so at first in a partial manner only, and do not circumscribe the whole circumference of the segment in the act of being formed. These fresh furrows are thus repetitions on a small scale of the earliest segmentation furrows.

It deserves to be noticed that the portion of the germinal disc which has already undergone segmentation, is still surrounded by a broad band of small-sized yolk-spherules. It appears to me probable that owing to changes taking place in the spherules of the yolk, which result in the formation of fresh spherules of a small size, this band undergoes a continuous renovation.

The uppermost row of segmentation spheres is now commencing to be distinguished from the remainder as a separate layer which becomes progressively more distinct as segmentation proceeds.

The largest segments in this section measure about the 1/100th of an inch in diameter, and the smallest about 1/300th of an inch.

The nuclei at this stage present points of rather a special interest. In the first place, though visible in many, and certainly present in all the segments[81], they are not confined to these: they are also to be seen, in small numbers, in the band of fine spherules which surrounds the already segmented part of the germinal disc. Those found outside the germinal disc are not confined to the spots where fresh segments are appearing, [Pg 228] but are also to be seen in places where there are no traces of fresh segments.

This fact, especially when taken in connection with the formation of fresh segments outside the germinal disc and with other facts which I shall mention hereafter, is of great morphological interest as bearing upon the nature and homologies of the food-yolk. It also throws light upon the behaviour and mode of increase of the nuclei. All the nuclei, both those of the segments and those of the yolk, have the peculiar structure I described in the last stage.

In specimens of this stage I have been able to observe certain points which have an important bearing upon the behaviour of the nucleus during cell-division.

Three figures, illustrating the behaviour of the nucleus, as I have seen it in sections of blastoderms hardened in chromic acid, are shewn in Pl. 6, figs. 7a, 7b and 7c.

In the place of the nucleus is to be seen a sharply defined figure (Fig. 7a) stained in the same way as the nucleus or more deeply. It has the shape of two cones placed base to base. From the apex of each cone there diverge towards the base a series of excessively fine striæ. At the junction between the two cones is an irregular linear series of small deeply stained granules which form an apparent break between the two. The line of this break is continued very indistinctly beyond the edge of the figure on each side.

From the apex of each cone there diverge outwards into the protoplasm of the cell a series of indistinct markings. They are rendered obscure by the presence of yolk-spherules, which completely surround the body just described, but which are not arranged with any reference to these markings. These latter striæ, diverging from the apex of the cone, are more distinctly seen when the apex points to the observer (Fig. 7b), than when a side of the cone is in view.

The striæ diverging outwards from the apices of the cones must be carefully distinguished from the striæ of the cones themselves. The cones are bodies quite as distinctly differentiated from the protoplasm of the cell as nuclei, while the striæ which diverge from their apices are merely structures in the general protoplasm of the cell.

[Pg 229] In some cells, which contain these bodies, no trace of a commencing line of division is visible. In other cases (Fig. 7c), such a line of division does appear and passes through the junction of the two cones. In one case of this kind I fancied I could see (and have represented) a coloured circular body in each cone. I do not feel any confidence that these two bodies are constantly present; and even where visible they are very indistinct.

Instead of an ordinary nucleus a very indistinctly marked vesicular body sometimes appears in a segment; but whether it is to be looked on as a nucleus not satisfactorily stained, or as a nucleus in the act of being formed, I cannot decide.

With reference to the situation of the cone-like bodies I have described I have made an observation which appears to me to be of some interest. I find that bodies of this kind are found in the yolk completely outside the germinal disc. I have made this observation, in at least two cases which admitted of no doubt (vide Fig. 7, nx´).

We have therefore the remarkable fact, that whatever connection these bodies may have with cell-division, they can occur in cases where this is altogether out of the question and where an increase in the number of nuclei can be their only product.

These are the main facts which I have been able to determine with reference to the nuclei of this stage; but it will conduce to clearness if I now finish what I have to say upon this subject.

At a still later stage of segmentation the same peculiar bodies are to be seen as during the stage just described, but they are rarer; and, in addition to them, other bodies are to be seen of a character intermediate between ordinary nuclei and the former bodies.

Three such are represented in Pl. 6, figs. 8a, 8b, 8c. In all of these there can be traced out the two cones, which are however very irregular. The striation of the cones is still present, but is not nearly so clear as it was in the earlier stage.

In addition to this, there are numerous deeply stained granules scattered about the two figures which resemble exactly the granules of typical nuclei.

[Pg 230] All these bodies occupy the place of an ordinary nucleus, they stain like an ordinary nucleus and are as sharply defined as an ordinary nucleus.

There is present around some of these, especially those situated in the yolk, the network of lines of the yolk described by me in a preliminary paper[82], and I feel satisfied that there is in some cases an actual connection between the network and the nuclei. This network I shall describe more fully hereafter.

Further points about these figures and the nuclei of this stage I should like to have been able to observe more completely than I have done, but they are so small that with the highest powers I possess (Zeiss, Immersion No. 2 = 1/15 in.) their complete and satisfactory investigation is not possible.

Most of the true nuclei of the cells of the germinal disc are regularly rounded; those however of the yolk are frequently irregular in shape and often provided with knob-like processes. The gradations are so complete between typical nuclei and bodies like that shewn (Pl. 6, fig. 8c) that it is impossible to refuse the name of nucleus to the latter.

In many cases two nuclei are present in one cell.

In later stages knob-like nuclei of various sizes are scattered in very great numbers in the yolk around the blastoderm (vide Pl. 7). In some cases it appears to me that several of these are in close juxtaposition, as if they had been produced by the division of one primitive nucleus. I do not feel absolutely confident that this is the case, owing to the fact that in the investigation of a knobbed body there is great difficulty in ascertaining that the knobs, which appear separate in one plane, are not in reality united in another.

I have, in spite of careful search, hitherto failed to find amongst these later nuclei cone-like figures, similar to those I found in the yolk during segmentation. This is the more remarkable since in the early stages of segmentation, when very few nuclei are present in the yolk, the cone-like figures are not uncommon; whereas, in the latter stages of development when the nuclei of the yolk are very common and obviously increasing rapidly, such figures are not to be met with.

[Pg 231] In no case have I been able to see a distinct membrane round any of the nuclei.

I have hitherto attempted to describe the appearances bearing on the behaviour of the nuclei in as objective a manner as possible.

My observations are not as complete as could be desired; but, taken in conjunction with those of other investigators, they appear to me to point towards certain definite conclusions with reference to the behaviour of the nucleus in cell-division.

The most important of these conclusions may be stated as follows. In the act of cell-division the nuclei of the resulting cells are formed from the nucleus of the primitive cell.

This may occur:—

(1) By the complete solution of the old nucleus within the protoplasm of the mother cell and the subsequent reaggregation of its matter to form the nuclei of the freshly formed daughter cells,

(2) By the simple division of the nucleus,

(3) Or by a process intermediate between these two where part of the old nucleus passes into the general protoplasm and part remains always distinguishable and divides; the fresh nucleus being in this case formed from the divided parts as well as from the dissolved parts of the old nucleus.

Included in this third process it is permissible to suppose that we may have a series of all possible gradations between the extreme processes 1 and 2. If it be admitted, and the evidence we have is certainly in favour of it, that in some cases, both in animal and vegetable cells, the nucleus itself divides during cell division, and in others the nucleus completely vanishes during the cell-division, it is more reasonable to suspect the existence of some connection between the two processes, than to suppose that they are entirely different in kind. Such a connection is given by the hypothesis I have just proposed.

The evidence for this view, derived both from my own observations and those of other investigators, may be put as follows.

The absolute division of the nucleus has been stated to occur in animal cells, but the number of instances where the [Pg 232] evidence is quite conclusive are not very numerous. Recently F. E. Schultze[83] appears to have observed it in the case of an Amœba in an altogether satisfactory manner. The instance is quoted by Flemming[84]. Schultze saw the nucleus assume a dumb-bell shape, divide, and the two halves collect themselves together. The whole process occupied a minute and a half and was shortly followed by the division of the Amœba, which occupied eight minutes. Amongst vegetable cells the division of the nucleus seems to be still rarer than with animal cells. Sachs[85] admits the division of the nucleus in the case of the parenchyma cells of certain Dicotyledons (Sambucus, Helianthus, Lysimachia, Polygonum, Silene) on the authority of Hanstein.

The division of the nucleus during cell-division, though seemingly not very common, must therefore be considered as a thoroughly well authenticated occurrence.

The frequent disappearance of the nucleus during cell-division is now so thoroughly recognised, both for animal and vegetable cells, as to require no further mention.

In many cases the partial or complete disappearance of the nucleus is accompanied by the formation of two peculiar star-like figures. Appearances of the kind have been described by Fol[86], Flemming[87], Auerbach[88] and possibly also Oellacher[89] as well as other observers.

These figures[90] are possibly due to the streaming out of the [Pg 233] protoplasm of the nucleus into that of the cell[91]. The appearance of striation may on this hypothesis be explained as due to the presence of granules in the protoplasm. When the streaming out of the protoplasm of a nucleus into that of a cell takes place, any large granule which cannot be moved by the stream will leave behind it a slack area where there is no movement of the fluid. Any granules which are carried into this area will remain there, and by the continuation of a process of this kind a row of granules may be formed, and a series of such rows would produce an appearance of striation. In many cases, e.g. Anodon, vide Flemming[92], even the larger yolk-spherules are arranged in this fashion.

On the supposition that the striation of these figures is due to the outflow from the nucleus, the appearances presented in Elasmobranchii admit of the following explanation.

The central body consisting of two cones (figs. 7a, 7c) is almost without question the remnant of the primitive nucleus. This is shewn by its occupying the same position as the primitive nucleus, staining in the same way, and by there being a series of insensible gradations between it and a typical nucleus. The contents must be supposed to be streaming out from the two apices of the cones, as appears from the striæ in the body converging on each side towards the apex, and then diverging again from it. In my specimens the yolk-spherules are not arranged with any reference to the radiating striation.

It is very likely that in the cases of the disappearance of the nucleus, its protoplasm streams out in two directions, towards the two parts of the cell which will eventually become separated from each other; and probably, after the division, the matter of the old nucleus is again collected to form two fresh nuclei.

In some cases of cell-division a remnant of the old nucleus is stated to be visible after the fresh nuclei have appeared. These cases, of which I have not seen full accounts, are perhaps analogous to what occasionally happens with the germinal [Pg 234] vesicle of an ovum. The whole of the contents of the germinal vesicle become at its disappearance mingled with the protoplasm of the ovum, but the resistant membrane remains and is eventually ejected from the egg, vide p. 215 et seq. If the remnant of the old nucleus in the cases described is nothing more than its membrane, no difficulty is offered to the view that the constituents of the old nucleus may help to form the new ones.

In many cases the total bulk of the new nuclei is greater than that of the old one; in such instances part of the protoplasm of the cell necessarily has a share in forming the new nuclei.

Although, in instances where the nucleus vanishes, an absolute demonstration of the formation of the fresh nuclei from the matter of the old one is not possible; yet, if cases of the division of the old nucleus to form the new ones be admitted to exist, the derivation in the first process of the fresh nuclei from the old ones must be postulated in order to maintain a continuity between the two processes of formation; and, as I have attempted to shew, all the circumstantial evidence is in favour of it.

Admitting the existence of the two extreme processes of nuclear formation, I wish to shew that my results in Elasmobranchii tend to demonstrate the existence of intermediate steps between them. The first figures I described of two opposed cones, appear to me almost certainly to represent nuclei in the act of dissolution; but though a portion of the nucleus may stream out into the yolk, I think it impossible that the whole of it does[93].

I described these bodies in two states. An earlier one, in which the two cones were separated by an irregular row of deeply stained granules; and a later one in which a furrow had already appeared dividing the cones as well as the cell. In neither of these conditions could I see any signs of the body vanishing completely. It was as clearly defined and as deeply stained as an ordinary nucleus, and in its later condition the signs of the streaming out of material from its pointed extremities were less marked than in the earlier stage.

[Pg 235] All these facts, to my mind, point to the view that these cone-like bodies do not disappear, but form the basis for the new nuclei. Possibly the body visible in each cone in the later stage, was the commencement of this new nucleus. Götte[94] has figured structures somewhat similar to these bodies, but I hardly understand either his figure or his account sufficiently clearly to be able to pronounce upon the identity of the two. In case they are identical, Götte gives a very different explanation of them from my own[95].

A second of my results, which points to a series of intermediate steps between division and solution of the nucleus, is the distribution in time of the peculiar cone-like bodies. These are present in fair abundance at an early period of segmentation, when there are but few nuclei either in the blastoderm or the yolk. But at later periods, when there are both more nuclei, especially in the yolk, and they are also increasing in numbers more rapidly than before, no bodies of this kind are to be seen. This fact becomes the more striking from the lobate appearance of the later nuclei of the yolk, an appearance which exactly suits the hypothesis of the rapid budding off of fresh nuclei.

The observations of R. Hertwig[96] on the gemmation of Podophrya gemmipara, support my interpretation of the knobbed condition of the nuclei. Hertwig finds (p. 47) that

The horse-shoe shaped nucleus grows out into numerous anastomosing projections. Over the free ends of the projections little knobs appear on the surface of the body, into which the lengthening ends of the processes of the nucleus grow up. Here they bend themselves into a horse-shoe form. The newly-formed nucleus then separates from the original nucleus, and afterwards the bud containing it from the body.

From the peculiar arrangement of the net-work of lines of the yolk around these knobbed nuclei, it is reasonable to conclude that interchange of material between the protoplasm of [Pg 236] the yolk and the nuclei is still taking place, even during the later periods.

These facts about the distribution in time of the cone-like bodies afford a strong presumptive evidence of a change in the manner of nuclear increase.

The last argument I propose urging on this head is derived from the bodies (Pl. 6, fig. 8a, b, c) which I have described as intermediate between the true cone-like bodies and typical nuclei. They appear to afford evidence of less and less of the matter of the nucleus streaming out into the yolk and of a large proportion of it becoming divided.

The conclusion to be derived from all these facts is that for Elasmobranchii in the earlier stages of segmentation, and during the formation of fresh segments, a partial solution of the old nucleus takes place, but all its constituents serve for the reconstruction of the fresh nuclei.

In later periods of development a still smaller part of the nucleus becomes dissolved, and the rest divides; but the two fresh nuclei are still derived from the two sources. After the close of segmentation the fresh nuclei are formed by a simple division of the older ones.

The appearance of the cone-like bodies in the yolk outside the germinal disc is a point of some interest. It demonstrates in a conclusive manner that whatever influence (if any) the nucleus may have in ordinary cases of cell division, yet it may undergo changes of a precisely similar character to those which it experiences during cell division, without exerting any influence on the surrounding protoplasm[97]. If the lobate nuclei are also nuclei undergoing division, we have in the egg of an Elasmobranch examples of all the known forms of nuclear increase unaccompanied by cell division.

The next stage in the segmentation does not present so many features of interest as the last one. The segments are [Pg 237] now so small, as to be barely visible from the surface with a simple lens. A section of an embryo of this stage is represented in Pl. 6, fig. 8. The section, which is drawn on the same scale as the section belonging to the last stage, serves to shew the relative size of the segments in the two cases.

The epiblast is now more distinct than it was. The segments composing it are markedly smaller than the remainder of the cells of the germinal disc, but possess nuclei of an absolutely larger size than do the other cells. They are irregular in shape, with a slight tendency to be columnar. An average segment of this layer measures about 1/700 inch.

The cells of the lower layer are more polygonal than those of the epiblast, and are decidedly larger. An average specimen of the larger cells of the lower layer measures about 1/400 in. in diameter, and is therefore considerably smaller than one of the smallest cells of the last stage. The formation of fresh segments from the yolk still continues with fair rapidity, but nearly comes to an end shortly after this.

Of the nuclei of the lower layer cells, there is not much to add to what has already been said. Not infrequently two nuclei may be observed in a single cell.

The nuclei in the yolk which surrounds the germinal disc are more numerous than in the earlier periods, and are now to be met with in fair numbers in every section (fig. 8, n´).

These are the main features which characterise the present stage, they are in all essential points similar to those of the last stage, and the two germinal discs hardly differ except in the size of the segments of which they are composed.

In the last stage which I consider as belonging to the segmentation, the cells of the whole blastoderm have become smaller (Pl. 6, fig. 9).

The epiblast (ep) now consists of a very marked layer of columnar cells. It is, as far as I have been able to observe, never more than one cell deep. The cells of the lower layer are of an approximately uniform size, though a few of those at the circumference of the blastoderm considerably exceed the remainder in the bulk.

There are two fresh features of importance in germinal discs of this age.

[Pg 238] Instead of being but indistinctly separated from the surrounding yolk, the blastoderm has now very clearly defined limits.

This is an especially marked feature of preparations made with osmic acid. In these there may frequently be seen a deeply stained doubly contoured line, which forms the limit of the yolk, where it surrounds the germinal disc. Lines of this kind are often to be seen on the surface of the yolk, or even of the blastoderm, but are probably to be regarded as products of reagents, rather than as organised structures. The outline of the germinal disc is well rounded, though it is occasionally broken, from the presence of a larger cell in the act of being formed from the yolk.

It is not probable that any great importance is to be attached to the comparative distinctness of the outline of the germinal disc at this stage, which is in a great measure due to a cessation in the formation of fresh cells in the surrounding yolk, and in part to the small and comparatively uniform size of the cells of the germinal disc.

The formation of fresh cells from the yolk nearly comes to an end during this period, but it still continues on a small scale.

The number of the nuclei around the germinal disc has increased.

Another feature of interest which first becomes apparent during this stage is the asymmetry of the germinal disc. If a section were made through the germinal disc, as it lay in situ in the egg capsule, parallel or nearly so to the long axis of the capsule, one end of the section would be found to be much thicker than the other. There would in fact be a far larger collection of cells at one extremity of the germinal disc than at the other. The end at which this collection of cells is formed points towards the end of the egg capsule opposite to that near which the yolk is situated. This collection of cells is the first trace of the embryo; and with its appearance the segmentation may be supposed to terminate.

The section I have represented, though not quite parallel to the long axis of the egg, is sufficiently nearly so to shew the greater mass of cells at the embryonic end of the germinal disc.

[Pg 239] This very early appearance of a distinction in the germinal disc between the extremity at which the embryo appears and the non-embryonic part of the disc, besides its inherent interest, has a further importance from the fact that in Osseous Fishes a similar occurrence takes place. Oellacher[98] and Götte[99] both agree as to the very early period at which a thickening of one extremity of the blastoderm in Osseous Fishes is formed, which serves to indicate the position at which the embryo will appear. There are many details of development in which Osseous Fish and Elasmobranchii agree, which, although if taken individually are without any great importance, yet serve to shew how long even insignificant features in development may be retained.

* * * * *

The segmentation of the Elasmobranch egg presents in most of its features great regularity, and exhibits in its mode of occurrence the closest resemblance to that in other meroblastic vertebrate ova.

There is, nevertheless, one point with reference to which a slight irregularity may be observed. In almost all eggs segmentation commences by, what for convenience may be called, a vertical furrow which is followed by a second vertical furrow at right angles to the first. The third furrow however is a horizontal one, and cuts the other two at right angles. This method of segmentation must be looked on as the normal one, in almost all the important groups of the animal kingdom, both for the so-called holoblastic and meroblastic eggs, and the gradations intermediate between the two. The Frog amongst vertebrates exhibits a most typical instance of this form of segmentation.

In Elasmobranchii the first two furrows are formed in a perfectly normal manner, but though I have not observed the actual formation of the next furrow, yet from the later stages, which I have observed, I conclude that it is parallel to one of the first formed furrows; and it is fairly certain that, not till a considerably later period, is a furrow homologous with the horizontal furrow of the Batrachian egg formed. This furrow appears to be represented in the Elasmobranch segmentation [Pg 240] by the irregular circumscription of a body of central smaller spheres from a ring of peripheral larger ones (vide Pl. 6, figs. 3, 4 and 5).

In the Bird the representative of the horizontal furrow appears relatively much earlier. It is formed when there are eight segments marked out on the surface of the germinal disc[100]. From Oellacher's[101] account of the segmentation in the fowl[102] it seems certain, as might be anticipated, that this furrow is nearly parallel to the surface of the disc, so that it cuts the earlier formed vertical furrows and causes the segments of the germinal disc to be completely circumscribed below as well as at the surface. In the Elasmobranch egg this is not the case; so that, even after the smaller central segments have become separated from the outer ring of larger ones, none of the segments of the disc are completely circumscribed, and only appear to be so in surface views (vide Pl. 6, fig. 6). Segmentation in the Elasmobranch egg differs in the following particulars from that in the Bird's egg:

(1) The equivalent of the horizontal furrow of the Batrachian egg appears much later than in the Bird.

(2) When it has appeared it travels inwards much more slowly.

As a result of these differences, the segments of the germinal disc of the Birds' eggs are much earlier circumscribed on all sides than those of the Elasmobranch egg.

As might be expected, the segmentation of the Elasmobranch egg resembles in many points that of Osseous Fishes (vide Oellacher[103] and Klein[104]). It may be noticed, that with Osseous as with Elasmobranch Fishes, the furrow corresponding with the horizontal furrow of the Amphibian's egg does not appear at as early a period as is normal. The third furrow of an Osseous Fish egg is parallel to one of the first formed pair.

In Oellacher's[105] figures, Pl. 23, figs. 19-21, peculiar beadings [Pg 241] of the sides of the earlier formed furrows are distinctly shewn. No mention of these is made in the text, but they are unquestionably similar to those I have described in the Elasmobranch furrows. In the case of Elasmobranchii I pointed out that not only were the sides of the furrow beaded, but that there appeared in the protoplasm, close to the furrows, peculiar vacuole-like cavities, precisely similar to the cavities which were the cause of the beadings of the furrows.

The presence of these seems to shew that the molecular cohesion of the protoplasm becomes, as compared with other parts, much diminished in the region where a furrow is about to appear, so that before the protoplasm finally gives way along a particular line to form a furrow, its cohesion is broken at numerous points in this region, and thus a series of vacuole-like spaces is formed.

If this is the true explanation of the formation of these spaces, their presence gives considerable support to the views of Dr Kleinenberg upon the causes of segmentation, so clearly and precisely stated in his monograph upon Hydra; and is opposed to any view which regards the forces which come into play during segmentation as resident in the nucleus.

I have not observed the peculiar threads of protoplasm which Oellacher[106] describes as crossing the commencing segmentation furrows. I have also failed to discover any signs of a concentration of the yolk-spherules, round one or two centres, in the segmentation spheres, similar to that observed by Oellacher in the segmenting eggs of Osseous Fish. The appearances observed by him are probably connected with the behaviour of the nucleus during segmentation, and are related to the curious bodies I have already described.

With reference to the nuclei which Oellacher[107] has described as occurring in the eggs of Osseous Fish during segmentation, there can, I think, be little doubt that they are identical with the peculiar nuclei in the Elasmobranch eggs.

He[108] says:

In an unsegmented germ there occurred at a certain point in the section ... a small aggregation of round bodies. I do not feel satisfied whether these aggregations represent one or more nuclei.

[Pg 242] Fig. 29 shews such aggregation; by focusing at its optical section eleven unequally large rounded bodies measuring from 0.004 - 0.009 mm. may be distinguished. They lay as if in a multilocular gap in the germ mass, which however they did not quite fill. In each of these bodies there appeared another but far smaller body. These aggregations were distinguished from the germ by an especially beautiful intense violet gold chloride colouration of their elements. The smaller elements contained in the larger were still more intensely coloured than the larger.

He further states that these aggregations equal the segments in number, and that the small bodies within the elements are not always to be seen with the same distinctness.

Oellacher's description as well as his figures of these bodies leaves no doubt in my mind that they are exactly similar bodies to those which I have already spoken of as nuclei, and the characteristic features of which I have shortly mentioned, and shall describe more fully at a later stage. A moderately full description of them is to be found in my preliminary paper[109].

Their division into a series of separate areas each with a deeply-stained body, as well as the staining of the whole of them, exactly corresponds to what I have found. That each is a single nucleus is quite certain, though their knobbed form might occasionally lead to the view of their being divided. This knobbed condition, observed by Oellacher as well as myself, certainly supports the view, that they are in the act of budding off fresh nuclei. Oellacher conceives, that the areas into which these nuclei are divided represent a series of separate bodies—this according to my observations is not the case. Nuclei of the same form have already been described in Nephelis, and are probably not very rare. They pass by insensible gradations into ordinary nuclei with numerous granules.

One marked feature of the segmentation of the Elasmobranch egg is the continuous advance of the process of segmentation into the yolk and the assimilation of this into the germ by the direct formation of fresh segments out of it. Into the significance of this feature I intend to enter fully hereafter; but it is interesting to notice that Oellacher's descriptions point to a similar feature in the segmentation of Osseous Fish. This however consists chiefly in the formation of fresh segments [Pg 243] from the lower parts of the germinal disc which in Osseous Fish is more distinctly marked off from the food-yolk than in Elasmobranchii.

I conclude my description of the segmentation by a short account of what other investigators have written about its features in these fishes. One of the earliest descriptions of this process was given by Leydig[110]. To his description of the germinal disc, I have already done full justice.

In the first stage of segmentation which he observed 20-30 segments were already visible on the surface. In each of these he recognized a nucleus but no nucleolus.

He rightly states that the segments have no membrane, and describes the yolk-spherules which fill them.

The next investigator is Gerbe[111]. I have unfortunately been unable to refer to this elaborate paper, but I gather from an abstract that M. Gerbe has given a careful description of the external features of segmentation.

Schenk[112] has also made important investigations on the subject. He considers that the ovum is invested with a very delicate membrane. This membrane I have failed to find a trace of, and agree with Leydig[113] in denying its existence. Schenk further found that after impregnation, but before segmentation, the germinal disc divided itself into two layers, an upper and a lower. Between the two a cavity made its appearance which Schenk looks upon as the segmentation cavity. Segmentation commences in the upper of the two layers, but Schenk does not give a precise account of the fate of the lower. I have had no opportunity of investigating the impregnated ovum before the commencement of segmentation, but my observations upon the early stages of this process render it clear that no division of the germinal disc exists subsequently [Pg 244] to the commencement of segmentation, and that the cavity discovered by Schenk can have no connection whatever with the segmentation cavity. I am indeed inclined to look upon this cavity as an artificial product. I have myself met with somewhat similar appearances, after the completion of segmentation, which were caused by the non-penetration of my hardening reagent beyond a certain point.

Without attempting absolutely to explain the appearances described by Professor Schenk, I think that his observations ought to be repeated, either by himself or some other competent observer.

Several further facts are recorded by Professor Schenk in his interesting paper. He states that immediately after impregnation, the germinal disc presents towards the yolk a strongly convex surface, and that at a later period, but still before the commencement of segmentation, this becomes flattened out. He has further detected amœboid movements in the disc at the same period. As to the changes of the germinal disc during segmentation, his paper contains no facts of importance.

Next in point of time to the paper of Schenk, is my own preliminary account of the development of the Elasmobranch Fishes[114]. In this a large number of the facts here described in full are briefly alluded to.

The last author who has investigated the segmentation in Elasmobranchii, is Dr Alexander Schultz[115]. He merely states that he has observed the segmentation, and confirms Professor Schenk's statements about the amœboid movements of the germinal disc.

EXPLANATION OF PLATE 6.

Fig. 1. Section through the germinal disc of a ripe ovarian ovum of the Skate. gv. germinal vesicle.

Fig. 2. Surface-view of a germinal disc with two furrows.

Figs. 3, 4, 5. Surface-views of three germinal discs in different stages of segmentation.

[Pg 245] Fig. 6. Section through the germinal disc represented in fig 3. n. nucleus; x. edge of germinal disc. The engraver has not accurately copied my original drawings in respect to the structure of the segmentation furrows.

Figs. 6a and 6b. Two furrows of the same germinal disc more highly magnified.

Fig. 6c. A nucleus from the same germinal disc highly magnified.

Fig. 7. Section through a germinal disc of the same age as that represented in fig. 4. n. nucleus; nx. modified nucleus; nx´. modified nucleus of the yolk; f. furrow appearing in the yolk around the germinal disc.

Figs. 7a, 7b, 7c. Three segments with modified nuclei from the same germinal disc.

Fig. 8. Section through a somewhat older germinal disc. ep. epiblast; n´. nuclei of yolk.

Figs. 8a, 8b, 8c. Modified nuclei from the yolk from the same germinal disc.

Fig. 8d. Segment in the act of division from the same germinal disc.

Fig. 9. Section through a germinal disc in which the segmentation is completed. It shews the larger collection of cells at the embryonic end of the germinal disc than at the non-embryonic. ep. epiblast.

[79] Rochen und Haie.

[80] The germinal disc figured was from the egg of a Scyllium stellare and not Pristiurus, but I have also sections of a Pristiurus egg of the same age, which do not differ materially from the Scyllium sections.

[81] In the figure of this stage, I have inserted nuclei in all the segments. In the section from which the figure was taken, nuclei were not to be seen in many of the segments, but I have not a question that they were present in all of them. The difficulty of seeing them is, in part, due to the yolk-spherules and in part to the thinness of the section as compared with the diameter of a segmentation sphere.

[82] Loc. cit.

[83] Archiv f. Micr. Anat. XI. p. 592.

[84] Entwicklungsgeschicte der Najaden, LXXI. Bd. der Sitz. der k. Acad. Wien, 1875.

[85] Text-Book of Botany, English trans. p. 19.

[86] Entw. d. Geryonideneies. Jenaische Zeitschrift, Bd. VII.

[87] Loc. cit.

[88] Organologische Studien, Zweites Heft.

[89] Beiträge z. Entwicklungsgeschichte der Knochenfischen. Zeit. für Wiss. Zoologie. Bd. XXII. 1872.

[90] The memoirs of Auerbach and Strasburger (Zellbildung u. Zelltheilung) have unfortunately come into my hands too late for me to take advantage of them. Especially in the magnificent monograph of Strasburger I find drawings precisely resembling those from my specimens already in the hands of the engraver. Strasburger comes to the conclusion from his investigations that the modified nucleus always divides and never vanishes as is usually stated. If his views on this point are correct part of the hypothesis I have suggested above is rendered unnecessary. The striæ of the protoplasm, which in accordance with Auerbach's view I have considered as being due to a streaming out of the matter of the nucleus, he regards as resulting from a polarity of the particles in the cell and the attraction of the nucleus. My own investigations though, as far as they go, quite in accordance with those of Strasburger, do not supply any grounds for deciding on the meaning of these striæ; and in some respects they support Strasburger's views against those of other observers, since they demonstrate that in Elasmobranchii the modified nucleus does actually divide.

[91] This is the view which has been taken by Auerbach (Organologische Studien).

[92] Loc. cit.

[93] After Strasburger's observation it must be considered very doubtful whether the streaming out of the contents of the nucleus, in the manner implied in the text, really takes place.

[94] Entwicklungsgeschite der Unke, Pl. 1, fig. 18.

[95] As I before mentioned, Strasburger (Zellbildung u. Zelltheilung) has represented bodies precisely similar to those I have described, which appear during the segmentation in the egg of Phallusia mammillata as well as similar figures observed by Butschli in eggs of Cucullanus elegans and Blatta Germanica. The figures in this monograph are the only ones I have seen, which are identical with my own.

[96] Morphologisches Jahrbuch, Bd. 1. pp. 46, 47.

[97] Strasburger's (loc. cit.) arguments about the influence of the nucleus in cell division are not to my mind conclusive; though not without importance. It is difficult to reconcile his views with the facts of cell division observable during the Elasmobranch segmentation; but even if their truth be admitted they do not bring us much nearer to a satisfactory understanding of cell division, unless accompanied (and at present they are not so) by a rational explanation of the forces which produce the division of the nucleus.

[98] Zeitschrift für Wiss. Zoologie, Bd. XXIII. 1873.

[99] Archiv für Micr. Anat. Bd. IX. 1873.

[100] Vide Elements of Embryology, p. 23.

[101] Stricker's Studien, 1869, Pt. I, Pl. II. fig. 4.

[102] Unfortunately Professor Oellacher gives no account of the surface appearance of the germinal discs of which he describes the sections. It is therefore uncertain to what period his sections belong.

[103] Zeitschrift für Wiss. Zool. Bd. XXII. 1872.

[104] Monthly Microscopical Journal, March, 1872.

[105] Loc. cit.

[106] Loc. cit.

[107] Loc. cit.

[108] Loc. cit. pp. 410, 411, &c.

[109] Loc. cit. p. 415. [This Edition, p. 64.]

[110] Rochen u. Haie. It is here mentioned that Coste observed the segmentation in these fishes.

[111] Recherches sur la segmentation des products adventifs de l'œuf des Plagiostomes et particulièrement des Raies. Robin, Journal de l'Anatomie et de la Physiologie, p. 609, 1872.

[112] Die Eier von Raja quadrimaculata innerhalb der Eileiter. Sitz. der k. Akad. Wien. Vol. LXXIII. 1873.

[113] Loc. cit. My denial of the existence of this membrane naturally applies only to the egg after impregnation, and to the genera Scyllium and Pristiurus.

[114] Loc. cit.

[115] Die Embryonal Anlage der Selachier. Vorläufige Mittheilung, Centralblatt f. Med. Wiss. No. 33, 1875.

[Pg 246]

CHAPTER III.

Formation of the Layers.

In the last chapter the blastoderm was left as a solid lens-shaped mass of cells, thicker at one end than at the other, its uppermost row of cells forming a distinct layer. There very soon appears in it a cavity, the well-known segmentation cavity, or cavity of von Baer, which arises as a small space in the midst of the blastoderm, near its non-embryonic end (Pl. 7, fig. 1.).

This condition of the segmentation cavity, though already[116] described, has nevertheless been met with in one case only. The circumstance of my having so rarely met with this condition is the more striking because I have cut sections of a considerable number of blastoderms in the hope of encountering specimens similar to the one figured, and it can only be explained on one of the two following hypotheses. Either the stage is very transitory, and has therefore escaped my notice except in the one instance; or else the cavity present in this instance is not the true segmentation cavity, but merely some abnormal structure. That this latter explanation is a possible one, appears from the fact that such cavities do at times occur in other parts of the blastoderm. Dr Schultz[117] does not mention having found any stage of this kind.

The position of the cavity in question, and its general appearance, incline me to the view that it is the segmentation cavity[118]. If this is the true view of its nature the fact should be [Pg 247] noted that at first its floor is formed by the lower layer cells and not by the yolk, and that its roof is constituted by both the lower layer cells and the epiblast cells. The relations of the floor undergo considerable modifications in the course of development.

The other features of the blastoderm at this stage are very much those of the previous stage.

The embryonic swelling is very conspicuous. The cells of the blastoderm are still disposed in two layers: an upper one of slightly columnar cells one deep, which constitutes the epiblast, and a lower one consisting of the remaining cells of the blastoderm.

An average cell of the lower layer has a diameter of about 1/900 inch, but the cells at the periphery of the layer are in some cases considerably larger than the more central ones. All the cells of the blastoderm are still completely filled with yolk spherules. In the yolk outside the peculiar nuclei, before spoken of, are present in considerable numbers. They seem to have been mistaken by Dr Schultz[119] for cells: there can however be no question that they are true nuclei.

In the next stage the relations of the segmentation cavity undergo important modifications.

The cells which form its floor disappear almost completely from that position, and the floor becomes formed by the yolk.

The stage, during which the yolk serves as the floor of the segmentation cavity, extends over a considerable period of time, but during it I have been unable to detect any important change in the constitution of the blastoderm. It no doubt gradually extends over the yolk, but even this growth is not nearly so rapid as in the succeeding stage. Although therefore the stage I proceed to describe is of long continuance, a blastoderm at the beginning of it exhibits, both in its external and in its internal features, no important deviations from one at the end of it.

Viewed from the surface (Pl. 8, fig. A) the blastoderm [Pg 248] at this stage appears slightly oval, but the departure from the circular form is not very considerable. The long axis of the oval corresponds with what eventually becomes the long axis of the embryo. From the yolk the blastoderm is still well distinguished by its darker colour; and it is surrounded by a concentric ring of light-coloured yolk, the outer border of which shades insensibly into the normal yolk.

At the embryonic portion of the blastoderm is a slight swelling, clearly shewn in Plate 8, fig. A, which can easily be detected in fresh and in hardened embryos. This swelling is to be looked upon as a local exaggeration of a slightly raised rim present around the whole circumference of the blastoderm. The roof of the segmentation cavity (fig. A, s.c.) forms a second swelling; and in the fresh embryo this region appears of a darker colour than other parts of the blastoderm.

It is difficult to determine the exact shape of the blastoderm, on account of the traction exercised upon it in opening the egg; and no reliance can be placed on the forms assumed by hardened blastoderms. This remark also applies to the sections of blastoderms of this stage. There can be no doubt that the minor individual variations exhibited by almost every specimen are produced in the course of manipulations while the objects are fresh. These variations may affect even the relative length of a particular region and certainly the curvature of it. The roof of the segmentation cavity is especially apt to be raised into a dome-like form.

The main internal feature of this stage is the disappearance of the layer of cells which, during the first stage, formed the floor of the segmentation cavity. This disappearance is nevertheless not absolute, and it is doubtful whether there is any period in which the floor of the cavity is quite without cells.

Dr Schultz supposes[120] that the entire segmentation cavity is, in the living animal, filled with a number of loose cells. Though it is not in my power absolutely to deny this, the point being one which cannot be satisfactorily investigated in sections, yet no evidence has come under my notice which would lead to the conclusion that more cells are present in the segmentation cavity than are represented on Pl. 13, fig. 1, of [Pg 249] my preliminary paper[121], an illustration which is repeated on Pl. 7, fig. 2.

The number of cells on the floor of the cavity differs considerably in different cases, but these cases come under the category of individual variations, and are not to be looked upon as indications of different states of development.

In many cases especially large cells are to be seen on the floor of the cavity (Pl. 7, fig. 2, bd). In my preliminary paper[122] the view was expressed that these are probably cells formed around the nuclei of the yolk. This view I am inclined to abandon, and to substitute for it the suggestion made by Dr Schultz, that they are remnants of the larger segmentation cells which were to be seen in the previous stages.

Plate 7, figs. 2, 3, 4 (all sections of this stage) shew the different appearances presented by the floor of the segmentation cavity. In only one of these sections are there any large number of cells upon the floor; and in no case have cells been observed imbedded in the yolk forming this floor, as described by Dr Schultz[123], but in all cases the cells simply rested upon it.

Passing from the segmentation cavity to the blastoderm itself, the first feature to be noticed is the more decided differentiation of the epiblast. This now forms a distinct layer composed of a single row of columnar cells. These are slightly more columnar in the region of the embryonic swelling than elsewhere, and become less elongated at the edge of the blastoderm. In my specimens this layer was never more than one cell deep, but Dr Schultz[124] states that, in the Elasmobranch embryos investigated by him, the epiblast was composed of more than a single row of cells.

Each epiblast cell is filled with yolk-spherules and contains a nucleus. Very frequently the nuclei in the layer are arranged in a regular row (vide Pl. 7, fig. 3). In the later blastoderms of this stage there is a tendency in the cells to assume a wedge-like form with their thin ends pointing alternately in opposite [Pg 250] directions. This arrangement is, however, by no means strictly adhered to, and the regularity of it is exaggerated in Plate 7, fig. 4.

The nuclei of the epiblast cells have the same characters as those of the lower layer cells to be presently described, but their intimate structure can only be successfully studied in certain exceptionally favourable sections. In most cases the yolk-spherules around them render the finer details invisible.

There is at this stage no such obvious continuity as in the succeeding stage between the epiblast and the lower layer cells; and this statement holds good more especially with the best conserved specimens which have been hardened in osmic acid (Pl. 7, fig. 4). In these it is very easy to see that the epiblast simply thins out at the edge of the blastoderm without exhibiting the slightest tendency to become continuous with the lower layer cells[125].

The lower layer cells form a mass rather than a layer, and constitute the whole of the blastoderm not included in the epiblast. The shape of this mass in a longitudinal section may be gathered from an examination of Plate 7, figs. 3 and 4.

It presents an especially thick portion forming the bulk of the embryonic swelling, and frequently contains one or two cavities, which from their constancy I regard as normal and not as artificial products.

In addition to the mass forming the embryonic swelling there is seen in sections another mass of lower layer cells at the opposite extremity of the blastoderm, connected with the [Pg 251] former by a bridge of cells, which constitutes the roof of the segmentation cavity. The lower layer cells may thus be divided into three distinct parts:

(1) The embryo swelling.

(2) The thick rim of cells round the edge of the remainder of the blastoderm.

(3) The cells which form the roof of the segmentation cavity.

These three parts form a continuous whole, but in addition to these there exist the previously mentioned cells, which rest on the floor of the segmentation cavity.

With the exception of these latter, the lower layer is composed of cells having a fairly uniform size, and exhibits no trace of a division into two layers.

The cells are for the most part irregularly polygonal from mutual pressure; and in their shape and arrangement, exhibit a marked contrast to the epiblast cells. A few of the lower layer cells, highly magnified, are represented in Pl. 7, fig. 2a. An average cell measures about 1/800 to 1/900 of an inch, but some of the larger ones on the floor attain to the 1/475 of an inch.

Owing to my having had the good fortune to prepare some especially favourable specimens of this stage, it has been possible for me to make accurate observations both upon the nuclei of the cells of the blastoderm, and upon the nuclei of the yolk.

The nuclei of the blastoderm cells, both of the epiblast and lower layer, have a uniform structure. Those of the lower layer cells are about 1/1600 of an inch in diameter. Roughly speaking each consists of a spherical mass of clear protoplasm refracting more highly than the protoplasm of its cell. The nucleus appears in sections to be divided by deeply stained lines into a number of separate areas, and in each of these a deeply stained granule is placed. In some cases two or more of such granules may be seen in a single area. The whole of the nucleus stains with the colouring reagents more deeply than the protoplasm of the cells; but this is especially the case with the granules and lines.

Though usually spherical the nuclei not infrequently have a somewhat lobate form.

Very similar to these nuclei are the nuclei of the yolk.

[Pg 252] One of the most important differences between the two is that of size. The majority of the nuclei present in the yolk are as large or larger than an ordinary blastoderm cell; while many of them reach a size very much greater than this. The examples I have measured varied from 1/500 to 1/250 of an inch in diameter.

Though they are divided, like the nuclei of the blastoderm, with more or less distinctness into separate areas by a network of lines, their greater size frequently causes them to present an aspect somewhat different from the nuclei of the blastoderm. They are moreover much less regular in outline than these, and very many of them have lobate projections (Pl. 7, figs. 2a and 2c and 3), which vary from simple knobs to projections of such a size as to cause the nucleus to present an appearance of commencing constriction into halves. When there are several such projections the nucleus acquires a peculiar knobbed figure. With bodies of this form it becomes in many cases a matter of great difficulty to decide whether or no a particular series of knobs, which appear separate in one plane, are united in a lower plane, whether, in fact, there is present a single knobbed nucleus or a number of nuclei in close apposition. A nucleus in this condition is represented in Pl. 7, fig. 2b.

The existence of a protoplasmic network in the yolk has already been mentioned. This in favourable cases may be observed to be in special connection with the nuclei just described. Its meshes are finer in the vicinity of the nuclei, and its fibres in some cases almost appear to start from them (Pl. 7, fig. 12). For reasons which I am unable to explain the nuclei of the yolk and the surrounding meshwork present appearances which differ greatly according to the reagent employed. In most specimens hardened in osmic acid the protoplasm of the nuclei is apparently prolonged in the surrounding meshwork (Pl. 7, fig. 12). In other specimens hardened in osmic acid (Pl. 7, fig. 11), and in all hardened in chromic acid (Pl. 7, fig. 2a and 2c), the appearances are far clearer than in the previous case, and the protoplasmic meshwork merely surrounds the nuclei, without shewing any signs of becoming continuous with them.

There is also around each nucleus a narrow space in which the spherules of the yolk are either much smaller than elsewhere or completely absent, vide Pl. 7, fig. 2b.

[Pg 253] It has not been possible for me to satisfy myself as to the exact meaning of the lines dividing these nuclei into a number of distinct areas. My observations leave the question open as to whether they are to be looked upon as lines of division, or as protoplasmic lines such as have been described in nuclei by Flemming[126], Hertwig[127] and Van Beneden[128]. The latter view appears to me to be the more probable one.

Such are the chief structural features presented by these nuclei, which are present during the whole of the earlier periods of development and retain throughout the same appearance. There can be little doubt that their knobbed condition implies that they are undergoing a rapid division. The arguments for this view I have already insisted on, and, in spite of the observations of Dr Kleinenberg shewing that similar nuclei of Nephelis do not undergo division, the case for their doing so in the Elasmobranch eggs is to my mind a very strong one.

During this stage the distribution of these nuclei in the yolk becomes somewhat altered from that in the earlier stages. Although the nuclei are still scattered generally throughout the finer yolk-matter around the blastoderm, yet they are especially aggregated at one or two points. In the first place a special collection of them may be noticed immediately below the floor of the segmentation cavity. They here form a distinct row or even layer. If the presence of this layer is coupled with the fact that at this period cells are beginning to appear on the floor of the segmentation cavity, a strong argument is obtained for the supposition that around these nuclei cells are being produced, which pass into the blastoderm to form the floor. Of the actual formation of cells at this period I have not been able to obtain any satisfactory example, so that it remains a matter of deduction rather than of direct observation.

Another special aggregation of nuclei is generally present at the periphery of the blastoderm, and the same amount of doubt hangs over the fate of these as over that of the previously mentioned nuclei.

[Pg 254] The next stage is the most important in the whole history of the formation of the layers. Not only does it serve to shew, that the process by which the layers are formed in Elasmobranchii can easily be derived from a simple gastrula type like that of Amphioxus, but it also serves as the key by which other meroblastic types of development may be explained. At the very commencement of this stage the embryonic swelling becomes more conspicuously visible than it was. It now projects above the level of the yolk in the form of a rim. At one point, which eventually forms the termination of the axis of the embryo, this projection is at its greatest; while on either side of this it gradually diminishes and finally vanishes. This projection I propose calling, as in my preliminary paper[129], the embryonic rim.

The segmentation cavity can still be seen from the surface, and a marked increase in the size of the blastoderm may be noticed. During the stage last described, the growth was but very slight; hence the rather sudden and rapid growth which now takes place becomes striking.

Longitudinal sections at this stage, as at the earlier stages, are the most instructive. Such a section on the same scale as Pl. 7, fig. 4, is represented in Pl. 7, fig. 5. It passes parallel to the long axis of the embryo, through the point of greatest development of the embryonic ring.

The three fresh features of the most striking kind are (1) the complete envelopment of the segmentation cavity within the lower layer cells, (2) the formation of the embryonic rim, (3) the increase in distance between the posterior end of the blastoderm and the segmentation cavity. The segmentation cavity has by no means relatively increased in size. The roof has precisely its earlier constitution, being composed of an internal lining of lower layer cells and an external one of epiblast. The thin lining of lower layer cells is, in the course of mounting the sections, very apt to fall off; but I am absolutely satisfied that it is never absent.

The floor of the cavity has undergone an important change, being now formed by a layer of cells instead of by the yolk. A [Pg 255] precisely similar but more partial change in the constitution of the floor takes place in Osseous Fishes[130].

The mode in which the floor is formed is a question of some importance. The nuclei, which during the last stage formed a row beneath it, probably, as previously pointed out, take some share in its formation. An additional argument to those already brought forward in favour of this view may be derived from the fact that during this stage such a row of nuclei is no longer present.

This argument may be stated as follows:

Before the floor of cells for the segmentation cavity is formed a number of nuclei are present in a suitable situation to supply the cells for the floor; as soon as the floor of cells makes its appearance these nuclei are no longer to be seen. From this it may be concluded that their disappearance arises from their having become the nuclei of the cells which form the floor.

It appears to me most probable that there is a growth inwards from the whole peripheral wall of the cavity, and that this ingrowth, as well as the cells derived from the yolk, assist in forming the floor of the cavity. In Osseous Fish there appears to be no doubt that the floor is largely formed by an ingrowth of this kind.

A great increase is observable in the distance between the posterior end of the segmentation cavity and the edge of the blastoderm. This is due to the rapid growth of the latter combined with the stationary condition of the former. The growth of the blastoderm at this period is not uniform, but is more rapid in the non-embryonic than in the embryonic parts.

The main features of the epiblast remain the same as during the last stages. It is still composed of a very distinct layer one cell deep. Over the segmentation cavity, and over the whole embryonic end of the blastoderm, the cells are very thin, columnar, and, roughly speaking, wedge-shaped with the thin ends pointing alternately in different directions. For this reason, the nuclei form two rows; but both the rows are situated near the upper surface of the layer (vide Pl. 7, fig. 5). Towards the [Pg 256] posterior end of the blastoderm the cells are flatter and broader; and the layer terminates at the non-embryonic end of the blastoderm without exhibiting the slightest tendency to become continuous with the lower layer cells. At the embryonic end of the blastoderm the relations of the epiblast and lower layer cells are very different. At this part, throughout the whole extent of the embryonic rim, the epiblast is reflected and becomes continuous with the lower layer cells.

The lower layer cells form, for the most part, a uniform stratum in which no distinction into mesoblast and hypoblast is to be seen.

Both the lower layer cells and the epiblast cells are still filled with yolk-spherules.

The structures at the embryonic rim, and the changes which are there taking place, unquestionably form the chief features of interest at this stage.

The general relations of these parts are very fairly shewn in Pl. 7, fig. 5, which represents a section passing through the median line of the embryonic region. They are however more accurately represented in Pl. 7, fig. 5a, taken from the same embryo, but in a lateral part of the embryonic rim; or in Pl. 7, fig. 6, from a slightly older embryo. In all of these figures the epiblast cells are reflected at the edge of the embryonic rim, and become perfectly continuous with the hypoblast cells. A few of the cells, immediately beyond the line of this reflection, precisely resemble in character the typical epiblast cells; but the remainder exhibit a gradual transition into typical lower layer cells. Adjoining these transitional cells, or partly enclosed in the corner formed between them and the epiblast, are a few unaltered lower layer cells (m), which at this stage are not distinctly separated from the transitional cells. The transitional cells form the commencement of the hypoblast (hy); and the cells (m) between them and the epiblast form the commencement of the mesoblast. The gradual conversion of lower layer cells into columnar hypoblast cells, is a very clear and observable phenomenon in the best specimens. Where the embryonic rim projects most, a larger number of cells have assumed a columnar form. Where it projects less clearly, a smaller number have done so. But in all cases there may be observed a series of gradations between [Pg 257] the columnar cells and the typical rounded lower layer cells[131].

In the last described embryo, although the embryonic rim had attained to a considerable development, no trace of the medullary groove had made its appearance. In an embryo in the next stage of which I propose describing sections, this structure has become visible.

A surface view of a blastoderm of this age, with the embryo, is represented on Pl. 8, fig. B; and I shall, for the sake of convenience, in future speak of embryos of this age as belonging to period B.

The blastoderm is nearly circular. The embryonic rim is represented by a darker shading at the edge. At one point in this rim may be seen the embryo, consisting of a somewhat raised area with an axial groove (mg). The head end of the embryo is that which points towards the centre of the blastoderm, and its free peripheral extremity is at the edge of the blastoderm.

A longitudinal section of an embryo of the same age as the one figured[132] is represented on Pl. 7, fig. 7. The general growth has been very considerable, though as before explained, it is mainly confined to that part of the blastoderm where the embryonic rim is absent.

A fresh feature of great importance is the complete disappearance of the segmentation cavity, the place which was previously occupied by it being now filled up by an irregular network of cells. There can be little question that the obliteration of the segmentation cavity is in part due to the entrance into the blastoderm of fresh cells formed around the nuclei of the yolk. The formation of these is now taking place with great rapidity and can be very easily followed.

Since the segmentation cavity ceases to play any further part in the history of the blastoderm, it will be well shortly to review the main points in its history.

[Pg 258] Its earliest appearance is involved in some obscurity, though it probably arises as a simple cavity in the midst of the lower layer cells (Pl. 7, fig. 1). In its second phase the floor ceases to be formed of lower layer cells, and the place of these is taken by the yolk, on which however a few scattered cells still remain (Pl. 7, figs. 2, 3, 4). During the third period of its history, a distinct cellular floor is again formed for it, so that it comes a second time into the same relations with the blastoderm as at its earliest appearance. The floor of cells which it receives is in part due to a growth inwards from the periphery of the blastoderm, and in part to the formation of fresh cells from the yolk. Coincidently with the commencing differentiation of hypoblast and mesoblast the segmentation cavity grows smaller and vanishes.

One of the most important features of the segmentation cavity in the Elasmobranchii which I have studied, is the fact that throughout its whole existence its roof is formed of lower layer cells. There is not the smallest question that the segmentation cavity of these fishes is the homologue of that of Amphioxus, Batrachians, etc., yet in the case of all of these animals, the roof of the segmentation cavity is formed of epiblast only. How comes it then to be formed of lower layer cells in Elasmobranchii?

To this question an answer was attempted in my paper, Upon the Early Stages of the Development of Vertebrates[133]. It was there pointed out, that as the food material in the ovum increases, the bulk of the lower layer cells necessarily also increases; since these, as far as the blastoderm is concerned, are the chief recipients of food material. This causes the lower layer cells to encroach upon the segmentation cavity, and to close it in not only on the sides, but also above; from the same cause it results that the lower layer cells assume, from the first, a position around the spot where the future alimentary cavity will be formed, and that this cavity becomes formed by a simple split in the midst of the lower layer cells, and not by an involution.

All the most recent observations[134] on Osseous Fishes tend [Pg 259] to shew that in them, the roof of the segmentation cavity is formed alone of epiblast; but on account of the great difficulty which is experienced in distinguishing the layers in the blastoderms of these animals, I still hesitate to accept as conclusive the testimony on this point.

In the formation a second time of a cellular floor for the segmentation cavity in the third stage, the Elasmobranch embryo seems to resemble that of the Osseous Fish[135]. Upon this feature great stress is laid both by Dr Götte[136] and Prof. Haeckel[137]: but I am unable to agree with the interpretation of it offered by them. Both Dr Götte and Prof. Haeckel regard the formation of this floor as part of an involution to which the lower layer cells owe their origin, and consider the involution an equivalent to the alimentary involution of Batrachians, Amphioxus, &c. To this question I hope to return, but it may be pointed out that my observations prove that this view can only be true in a very modified sense; since the invagination by which hypoblast and alimentary canal are formed in Amphioxus is represented in Elasmobranchii by a structure quite separate from the ingrowth of cells to form the floor of the segmentation cavity.

The eventual obliteration of the segmentation cavity by cells derived from the yolk is to be regarded as an inherited remnant of the involution by which this obliteration was primitively effected. The passage upwards of cells from the yolk, may possibly be a real survival of the tendency of the hypoblast cells to grow inwards during the process of involution.

The last feature of the segmentation cavity which deserves notice is its excentric position. It is from the first situated in much closer proximity to the non-embryonic than to the embryonic end of the blastoderm. This peculiarity in position is also characteristic of the segmentation cavity of Osseous Fishes, as is shewn by the concordant observations of Oellacher[138] and Götte[139]. Its meaning becomes at once intelligible by referring to the diagrams in my paper[140] on the Early Stages in the Development of Vertebrates. It in fact arises from the asymmetrical character [Pg 260] of the primitive alimentary involution in all anamniotic vertebrates with the exception of Amphioxus.

Leaving the segmentation cavity I pass on to the other features of my sections.

There is still to be seen a considerable aggregation of cells at the non-embryonic end of the blastoderm. The position of this, and its relations with the portion of the blastoderm which at an earlier period contained the segmentation cavity, indicate that the growth of the blastoderm is not confined to its edge, but that it proceeds at all points causing the peripheral parts to glide over the yolk.

The main features of the cells of this blastoderm are the same as they were in the one last described. In the non-embryonic region the epiblast has thinned out, and is composed of a single row of cells, which, in the succeeding stages, become much flattened.

The lower layer cells over the greater part of their extent, have not undergone any histological changes of importance. Amongst them may frequently be seen a few exceptionally large cells, which without doubt have been derived directly from the yolk.

The embryonic rim is now a far more considerable structure than it was. Vide Pl. 7, fig. 7. Its elongation is mainly effected by the continuous conversion of rounded lower layer cells into columnar hypoblast cells at its central or anterior extremity.

This conversion of the lower layer cells into hypoblast cells is still easy to follow, and in every section cells intermediate between the two are to be seen. The nature of the changes which are taking place requires for its elucidation transverse as well as longitudinal sections. Transverse sections of a slightly older embryo than B are represented on Pl. 7, fig. 8a, 8b and 8c.

Of these sections a is the most peripheral or posterior, and c the most central or anterior. By a combination of transverse and longitudinal sections, and by an inspection of a surface view, it is rendered clear that, though the embryonic rim is a far more considerable structure in the region of the embryo than elsewhere (compare fig. 6 and fig. 7 and 7a), yet that this gain in size is not produced by an outgrowth of the embryo beyond [Pg 261] the rest of the germ, but by the conversion of the lower layer cells into hypoblast having been carried far further towards the centre of the germ in the axial line than in the lateral regions of the rim.

The most anterior of the series of transverse sections (Pl. 7, fig. 8c) I have represented, is especially instructive with reference to this point. Though the embryonic rim is cut through at the sides of the section, yet in these parts the rim consists of hardly more than a continuity between epiblast and lower layer cells, and the lower layer cells shew no trace of a division into mesoblast and hypoblast. In the axis of the embryo, however, the columnar hypoblast is quite distinct; and on it a small cap of mesoblast is seen on each side of the medullary groove. Had the embryonic rim resulted from a projecting growth of the blastoderm, such a condition could not have existed. It might have been possible to find the hypoblast formed at the sides of the section and not at the centre; but the reverse, as in these sections, could not have occurred. Indeed it is scarcely necessary to have recourse to sections to prove that the growth of the embryonic rim is towards the centre of the blastoderm. The inspection of a surface view of a blastoderm at this period demonstrates it beyond a doubt (Pl. 8, fig. B). The embryo, close to which the embryonic rim is alone largely developed, does not project outwards beyond the edge of the germ, but inwards towards its centre.

The space between the embryonic rim and the yolk (Pl. 7, fig. 7, al.) is the alimentary cavity. The roof of this is therefore primitively formed of hypoblast and the floor of yolk. The external opening of this space at the edge of the blastoderm is the exact morphological homologue of the anus of Rusconi, or blastopore of Amphioxus, the Amphibians, &c. The importance of the mode of growth in the embryonic rim depends upon the homology of the cavity between it and the yolk, with the alimentary cavity of Amphioxus and Amphibians. Since this homology exists, the direction of the growth of this cavity ought to be, as it in fact is, the same as in Amphioxus, etc., viz. towards the centre of the germ and original position of the segmentation cavity. Thus though a true invagination is not present as in the other cases, yet this is represented in Elasmobranchii by the [Pg 262] continuous conversion of lower layer cells into hypoblast along a line leading towards the centre of the blastoderm.

In the parts of the rim adjoining the embryo, the lower layer cells, on becoming continuous with the epiblast cells, assume a columnar form. At the sides of the rim this is not strictly the case, and the lower layer cells retain their rounded form, though quite continuous with the epiblast cells. One curious feature of the layer of epiblast in these lateral parts of the rim is the great thickness it acquires before being reflected and becoming continuous with the hypoblast (Pl. 7, fig. 8c). In the vicinity of the point of reflection there is often a rather large formation of cells around the nuclei of the yolk. The cells formed here no doubt pass into the blastoderm, and become converted into columnar hypoblast cells. In some cases the formation of these cells is very rapid, and they produce quite a projection on the under side of the hypoblast. Such a case is represented in Pl. 7, fig. 8b, n.al. The cells constituting this mass eventually become converted into the lateral and ventral walls of the alimentary canal.

The formation of the mesoblast has progressed rapidly. While many of the lower layer cells become columnar and form the hypoblast, others, between these and the epiblast, remain spherical. The latter do not at once become separated as a layer distinct from the hypoblast, and, at first, are only to be distinguished from them through their different character, vide Plate 7, figs. 6 and 7. They nevertheless constitute the commencing mesoblast.

Thus much of the mode of formation of the mesoblast can be easily made out in longitudinal sections, but transverse sections throw still further light upon it.

From these it may at once be seen that the mesoblast is not formed in one continuous sheet, but as two lateral masses, one on each side of the axial line of the embryo[141]. In my [Pg 263] preliminary account[142] it was stated that this was a condition of the mesoblast at a very early period, and that it was probably its condition from the beginning. Sections are now in my possession which satisfy me that, from the very first, the mesoblast arises as two distinct lateral masses, one on each side of the axial line.

In the embryo from which the sections Pl. 7, fig. 8a, 8b, 8c were taken, the mesoblast had, in most parts, not yet become separated from the hypoblast. It still formed with this a continuous layer, though the mesoblast cells were distinguishable by their shape from the hypoblast. In only one section (b) was any part of the mesoblast quite separated from the hypoblast.

In the hindermost part of the embryo the mesoblast is at its maximum, and forms, on each side, a continuous sheet extending from the median line to the periphery (fig. 8a). The rounder form of the mesoblast cells renders the line of junction between the layer constituted by them and the hypoblast fairly distinct; but towards the periphery, where the hypoblast cells have the same rounded form as the mesoblast, the fusion between the two layers is nearly complete.

In an anterior section the mesoblast is only present as a cap on both sides of the medullary groove, and as a mass of cells at the periphery of the section (fig. 8b); but no continuous layer of it is present. In the foremost of the three sections (fig. 8c) the mesoblast can scarcely be said to have become in any way separated from the hypoblast except at the summit of the medullary folds (m).

From these and similar sections it may be certainly concluded, that the mesoblast becomes first separated from the hypoblast as a distinct layer in the posterior region of the embryo, and only at a later period in the region of the head.

In an embryo but slightly more developed than B, the formation of the layer is quite completed in the region of the embryo. To this embryo I now pass on.

In the non-embryonic parts of the blastoderm no fresh features of interest have appeared. It still consists of two layers. The epiblast is composed of flattened cells, and the lower layer of a network of more rounded cells, elongated in a lateral [Pg 264] direction. The growth of the blastoderm has continued to be very rapid.

In the region of the embryo (Pl. 7, fig. 9) more important changes have occurred. The epiblast still remains as a single row of columnar cells. The hypoblast is no longer fused with the mesoblast, and forms a distinct dorsal wall for the alimentary cavity. Though along the axis of the embryo the hypoblast is composed of a single row of columnar cells, yet in the lateral part of the embryo its cells are less columnar and are one or two deep.

Owing to the manner in which the mesoblast became split off from the hypoblast, a continuity is maintained between the hypoblast and the lower layer cells of the blastoderm (Pl. 7, fig. 9), while the two plates of mesoblast are isolated and disconnected from any other masses of cells.

The alimentary cavity is best studied in transverse sections. (Vide Pl. 7, fig. 10a, 10b and 10c, three sections from the same embryo.) It is closed in above and at the sides by the hypoblast, and below by the yolk. In its anterior part a floor is commencing to be formed by a growth of cells from the walls of the two sides. The cells for this growth are formed around the nuclei of the yolk; a feature which recalls the fact that in Amphibians the ventral wall of the alimentary cavity is similarly formed in part from the so-called yolk cells.

We left the mesoblast as two masses not completely separated from the hypoblast. During this stage the separation between the two becomes complete, and there are formed two great lateral plates of mesoblast cells, one on each side of the medullary groove. Each of these corresponds to a united vertebral and lateral plate of the higher Vertebrates. The plates are thickest in the middle and posterior regions (Pl. 7, fig. 10a and 10b), but thin out and almost vanish in the region of the head. The longitudinal section of this stage represented in Pl. 7, fig. 9, passes through one of the lateral masses of mesoblast cells, and shews very distinctly its complete independence of all the other cells in the blastoderm.

From what has been stated with reference to the development of the mesoblast, it is clear that in Elasmobranchii this layer is derived from the same mass of cells as the hypoblast, [Pg 265] and receives none of its elements from the epiblast. In connection with its development, as two independent lateral masses, I may observe, as I have previously done[143], that in this respect it bears a close resemblance to mesoblast in Euaxes, as described by Kowalevsky[144]. This resemblance is of some interest, as bearing on a probable Annelid origin of Vertebrata. Kowalevsky has also shewn[145] that the mesoblast in Ascidians is similarly formed as two independent masses, one on each side of the middle line.

It ought, however, to be pointed out that a similar bilateral origin of the mesoblast had been recently met with in Lymnæus by Carl Rabl[146]. A fact which somewhat diminishes the genealogical value of this feature in the mesoblast in Elasmobranchii.

During the course of this stage the spherules of food-yolk immediately beneath the embryo are used up very rapidly. As a result of this the protoplasmic network, so often spoken of, comes very plainly into view. Considerable areas may sometimes be seen without any yolk-spherule whatever.

On Pl. 7, fig. 7a, and figs. 11 and 12, I have attempted to reproduce the various appearances presented by this network: and these figures give a better idea of it than any description. My observations tend to shew that it extends through the whole yolk, and serves to hold it together. It has not been possible for me to satisfy myself that it had any definite limits, but on the other hand, in many parts all my efforts to demonstrate its presence have failed. When the yolk-spherules are very thickly packed, it is difficult to make out for certain whether it is present or absent, and I have not succeeded in removing the yolk-spherules from the network in cases of this kind. In medium-sized ovarian eggs this network is very easily seen, and extends through the whole yolk. Part of such an egg is shewn in Pl. 7, [Pg 266] fig. 14. In full-sized ovarian eggs, according to Schultz[147], it forms, as was mentioned in the first chapter, radiating striæ, extending from the centre to the periphery of the egg. When examined with the highest powers, the lines of this network appear to be composed of immeasurably small granules arranged in a linear direction. These granules are more distinct in chromic acid specimens than in those hardened in osmic acid, but are to be seen in both. There can be little doubt that these granules are imbedded in a thread or thin layer of protoplasm.

I have already (p. 252) touched upon the relation of this network to the nuclei of the yolk[148].

During the stages which have just been described specially favourable views are frequently to be obtained of the formation of cells in the yolk and their entrance into the blastoderm. Two representations of these are given, in Pl. 7, fig. 7a, and fig. 13. In both of these distinctly circumscribed cells are to be seen in the yolk (c), and in all cases are situated near to the typical nuclei of the yolk. The cells in the yolk have such a relation to the surrounding parts, that it is quite certain that their presence is not due to artificial manipulation, and in some cases it is even difficult to decide whether or no a cell area is circumscribed round a nucleus (Pl. 7, fig. 13). Although it would be possible for cells in the living state to pass from the blastoderm into the yolk, yet the view that they have done so in the cases under consideration has not much to recommend it, if the following facts be taken into consideration. (1) That the cells [Pg 267] in the yolk are frequently larger than those in the blastoderm. (2) That there are present a very large number of nuclei in the yolk which precisely resemble the nuclei of the cells under discussion. (3) That in some cases (Pl. 7, fig. 13) cells are seen indistinctly circumscribed as if in the act of being formed.

Between the blastoderm and the yolk may frequently be seen a membrane-like structure, which becomes stained with hæmatoxylin, osmic acid etc. It appears to be a layer of coagulated albumen and not a distinct membrane.

Summary.

At the close of segmentation, the blastoderm forms a somewhat lens-shaped disc, thicker at one end than at the other; the thicker end being termed the embryonic end.

It is divided into two layers—an upper one, the epiblast, formed by a single row of columnar cells; and a lower one, consisting of the remaining cells of the blastoderm.

A cavity next appears in the lower layer cells, near the non-embryonic end of the blastoderm, but the cells soon disappear from the floor of this cavity which then comes to be constituted by yolk alone.

The epiblast in the next stage is reflected for a small arc at the embryonic end of the blastoderm, and becomes continuous with the lower layer cells; at the same time some of the lower layer cells of the embryonic end of the blastoderm assume a columnar form, and constitute the commencing hypoblast. The portion of the blastoderm, where epiblast and hypoblast are continuous, forms a projecting structure which I have called the embryonic rim. This rim increases rapidly by growing inwards more and more towards the centre of the blastoderm, through the continuous conversion of lower layer cells into columnar hypoblast.

While the embryonic rim is being formed, the segmentation cavity undergoes important changes. In the first place, it receives a floor of lower layer cells, partly from an ingrowth from the two sides, and partly from the formation of cells around the nuclei of the yolk.

[Pg 268] Shortly after the floor of cells has appeared, the whole segmentation cavity becomes obliterated.

When the embryonic rim has attained to some importance, the position of the embryo becomes marked out by the appearance of the medullary groove at its most projecting part. The embryo extends from the edge of the blastoderm inwards towards the centre.

At about the time of the formation of the medullary groove, the mesoblast becomes definitely constituted. It arises as two independent plates, one on each side of the medullary groove, and is entirely derived from lower layer cells.

The two plates of mesoblast are at first unconnected with any other cells of the blastoderm, and, on their formation, the hypoblast remains in connection with all the remaining lower layer cells. Between the embryonic rim and the yolk is a cavity,—the primitive alimentary cavity. Its roof is formed of hypoblast, and its floor of yolk. Its external opening is homologous with the anus of Rusconi, of Amphioxus and the Amphibians. The ventral wall of the alimentary cavity is eventually derived from cells formed in the yolk around the nuclei which are there present.

* * * * *

Since the important researches of Gegenbaur[149] upon the meroblastic vertebrate eggs, it has been generally admitted that the ovum of every vertebrate, however complicated may be its apparent constitution, is nevertheless to be regarded as a simple cell. This view is, indeed, opposed by His[150] and to a very modified extent by Waldeyer[151], and has recently been attacked from an entirely new standpoint by Götte[152]; but, to my mind, the objections of these authors do not upset the well founded conclusions of previous observations.

[Pg 269] As soon as the fact is recognised that both meroblastic and holoblastic eggs have the same fundamental constitution, the admission follows, naturally, though not necessarily, that the eggs belonging to these two classes differ solely in degree, not only as regards their constitution, but also as regards the manner in which they become respectively converted into the embryo. As might have been anticipated, this view has gained a wide acceptance.

Amongst the observations, which have given a strong objective support to this view, may be mentioned those of Professor Lankester upon the development of Cephalopoda[153], and of Dr Götte[154] upon the development of the Hen's egg. In Loligo Professor Lankester shewed that there appeared, in the part of the egg usually considered as food-yolk, a number of bodies, which eventually developed a nucleus and became cells, and that these cells entered into the blastoderm. These observations demonstrate that in the eggs of Loligo the so-called food-yolk is merely equivalent to a part of the egg which in other cases undergoes segmentation.

The observations of Dr Götte have a similar bearing. He made out that in the eggs of the Hen no sharp line is to be found separating the germinal disc from the yolk, and that, independently of the normal segmentation, a number of cells are derived from that part of the egg hitherto regarded as exclusively food-yolk. This view of the nature of the food-yolk was also advanced in my preliminary account of the development of Elasmobranchii[155], and it is now my intention to put forward the positive evidence in favour of this view, which is supplied from a knowledge of the phenomena of the development of the Elasmobranch ovum; and then to discuss how far the facts of the growth of the blastoderm in Elasmobranchii accord with the view that their large food-yolk is exactly equivalent to part of the ovum, which in Amphibians undergoes segmentation, rather than some fresh addition, which has no equivalent in the Amphibian or other holoblastic ovum.

Taking for granted that the ripe ovum is a single cell, the [Pg 270] question arises whether in the case of meroblastic ova the cell is not constituted of two parts completely separated from one another.

Is the meroblastic ovum, before or after impregnation, composed of a germinal disc in which all the protoplasm of the cell is aggregated, and of a food-yolk in which no protoplasm is present? or is the protoplasm present throughout, being simply more concentrated at the germinal pole than elsewhere? If the former alternative is accepted, we must suppose that the mass of food-yolk is a something added which is not present in holoblastic ova. If the latter alternative is accepted, it may then be maintained that holoblastic and meroblastic ova are constituted in the same way and differ only in the proportions of their constituents.

My own observations in conjunction with the specially interesting observations of Dr Schultz[156] justify the view which regards the protoplasm as present throughout the whole ovum, and not confined to the germinal disc. Our observations shew that a fine protoplasmic network, with ramifications extending throughout the whole yolk, is present both before and after impregnation.

The presence of this network is, in itself, only sufficient to prove that the yolk may be equivalent to part of a holoblastic ovum; to demonstrate that it is so requires something more, and this link in the chain of evidence is supplied by the nuclei of the yolk, which have been so often referred to.

These nuclei arise independently in the yolk, and become the nuclei of cells which enter the germ and the bodies of which are derived from the protoplasm of the yolk. Not only so, but the cells formed around these nuclei play the same part in the development of Elasmobranchii as do the largest so-called yolk cells in the development of Amphibians. Like the homologous cells in Amphibians, they mainly serve to form the ventral wall of the alimentary canal and the blood-corpuscles. The identity in the fate of the so-called yolk cells of Amphibians with the cells derived from the yolk in Elasmobranchii, must be considered as a proof of the homology of the yolk cells in the first case [Pg 271] with the yolk in the second; the difference between the yolk in the two cases arising from the fact that in the Elasmobranch ovum the yolk-spherules bear a larger proportion to the protoplasm than they do in the Amphibian ovum. As I have suggested elsewhere[157], the segmentation or non-segmentation of a particular part of the ovum depends solely upon the proportion borne by the protoplasm to the yolk particles; so that, when the latter exceed the former in a certain fixed proportion, segmentation is no longer possible; and, as this limit is approached, segmentation becomes slower, and the resulting segments larger and larger.

The question how far the facts in the developmental history of the various vertebrate blastoderms accord with the view of the nature of the yolk just propounded is one of considerable interest. An answer to it has already been attempted from a general point of view in my paper[158] entitled 'The Comparison of the early stages of development in Vertebrates'; but the subject may be conveniently treated here in a special manner for Elasmobranch embryos.

In the woodcut, fig. 1, A, B, C[159], are represented three diagrammatic longitudinal sections of an Elasmobranch embryo. A nearly corresponds with the longitudinal section represented on Pl. 7, fig. 4, and B with Pl. 7, fig. 7. In Pl. 7, fig. 7, the segmentation cavity has however completely disappeared, while it is still represented as present in the diagram of the same period. If these diagrams, or better still, the woodcuts fig. 2 A, B, C (which only differ from those of the Elasmobranch fish in the smaller amount of food-yolk), be compared with the corresponding ones of Bombinator, fig. 3, A, B, C, they will be found to be in fundamental agreement with them. First let fig. 1, A, or fig. 2, A, or Pl. 7, fig. 4, be compared with fig. 3, A. In all there is present a segmentation cavity situated not centrally but near the surface of the egg. The roof of the cavity is thin in all, being composed in the Amphibian of epiblast alone, and in[Pg 272] the Elasmobranch of epiblast and lower layer cells. The floor of the cavity is, in all, formed of so-called yolk (vide Pl. 7, fig. 4), which in all forms the main mass of the egg. In the Amphibian the yolk is segmented, and, though it is not segmented in the Elasmobranch, it contains in compensation the nuclei so often mentioned. In all, the sides of the segmentation cavity are formed by lower layer cells. In the Amphibian the sides are enclosed by smaller cells (in the diagram) which correspond exactly in function and position with the lower layer cells of the Elasmobranch blastoderm.

Fig. 1.

Diagrammatic longitudinal sections of an Elasmobranch embryo.

Epiblast without shading. Mesoblast black with clear outlines to the cells. Lower layer cells and hypoblast with simple shading.

ep. epiblast. m. mesoblast. al. alimentary cavity. sg. segmentation cavity. nc. neural canal. ch. notochord. x. point where epiblast and hypoblast become continuous at the posterior end of the embryo. n. nuclei of yolk.

A. Section of young blastoderm, with segmentation cavity in the middle of the lower layer cells.

B. Older blastoderm with embryo in which hypoblast and mesoblast are distinctly formed, and in which the alimentary slit has appeared. The segmentation cavity is still represented as being present, though by this stage it has in reality disappeared.

C. Older blastoderm with embryo in which neural canal has become formed, and is continuous posteriorly with alimentary canal. The notochord, though shaded like mesoblast, belongs properly to the hypoblast.

[Pg 273]

Fig. 2.

Diagrammatic longitudinal sections of embryo, which develops in the same manner as the Elasmobranch embryo, but in which the ovum contains far less food-yolk than is the case with the Elasmobranch ovum.

Epiblast without shading. Mesoblast black with clear outlines to the cells. Lower layer cells and hypoblast with simple shading.

ep. epiblast. m. mesoblast. hy. hypoblast. sg. segmentation cavity. al. alimentary cavity. nc. neural canal. hf. head fold. n. nuclei of the yolk.

The stages A, B and C are the same as in figure .[TN9]

[Pg 274]

Fig. 3.

Diagrammatic longitudinal sections of Bombinator igneus. Reproduced with modifications from Götte.

Epiblast without shading. Mesoblast black with clear outlines to the cells. Lower layer cells and hypoblast with simple shading.

ep. epiblast. l.l. lower layer cells. y. smaller lower layer cells at the sides of the segmentation cavity. m. mesoblast. hy. hypoblast. al. alimentary cavity. sg. segmentation cavity. nc. neural cavity. yk. yolk-cells.

A is the youngest stage in which the alimentary involution has not yet appeared. x is the point from which the involution will start to form the dorsal wall of the alimentary tract. The line on each side of the segmentation cavity, which separates the smaller lower layer cells from the epiblast cells, is not present in Götte's original figure. The two shadings employed in the diagram render it necessary to have some line, but at this stage it is in reality not possible to assert which cells belong to the epiblast and which to the lower layer.

B. In this stage the alimentary cavity has become formed, but the segmentation cavity is not yet obliterated.

x. point where epiblast and hypoblast become continuous.

C. The neural canal is already formed, and communicates posteriorly with the alimentary.

x. point where epiblast and hypoblast become continuous.

[Pg 275] The relation of the yolk to the blastoderm in the Elasmobranch embryo at this stage of development very well suits the view of its homology with the large cells of the Amphibian ovum. The only essential difference between the two ova arises from the roof of the segmentation cavity being in the Elasmobranch embryo formed of lower layer cells, which are absent in the Amphibian embryo. This difference no doubt depends upon the greater quantity of yolk particles present in the Elasmobranch ovum. These increase the bulk of the lower layer cells, which are thus compelled to creep up the sides of the segmentation cavity till they close it in above.

In the next stage for the Elasmobranch, fig. 1 and 2 B and Pl. 7, fig. 7, and for the Amphibian, fig. 3, B, the agreement between the two types is again very close. In both for a small portion (x) of the edge of the blastoderm the epiblast and hypoblast become continuous, while at all other parts the epiblast, accompanied by lower layer cells, grows round the yolk or round the large cells which correspond to it. The yolk cells of the Amphibian ovum form a comparatively small mass, and are therefore rapidly enveloped; while in the case of the Elasmobranch ovum, owing to the greater mass of the yolk, the same process occupies a long period. In both ova the portion of the blastoderm, where epiblast and hypoblast become continuous, forms the dorsal lip of an opening—the anus of Rusconi—which leads into the alimentary cavity. This cavity has the same relation in both ova. It is lined dorsally by lower layer cells, and ventrally by yolk or what corresponds with yolk; the ventral epithelium of the alimentary canal being in both cases eventually supplied by the yolk cells.

As in the earlier stage, so in the present one, the anatomical relations of the yolk to the blastoderm in the one case (Elasmobranch) are nearly identical with those of the yolk cells to the blastoderm in the other (Amphibian). The main features in which the two embryos differ, during the stage under consideration, arise from the same cause as the solitary point of difference during the preceding stage.

[Pg 276] In Amphibians, the alimentary cavity is formed coincidently with a true ingrowth of cells from the point where epiblast and hypoblast become continuous, and from this ingrowth the dorsal wall of the alimentary cavity is formed. The same ingrowth causes the obliteration of the segmentation cavity.

In the Elasmobranchii, owing to the larger bulk of the lower layer cells caused by the food-yolk, these have been compelled to arrange themselves in their final position during segmentation, and no room is left for a true invagination; but instead of this there is formed a simple split between the blastoderm and the yolk. The homology of this with the primitive invagination is nevertheless proved by the survival of a number of features belonging to the ancestral condition in which a true invagination was present. Amongst the more important of these are the following:—(1) The continuity of epiblast and hypoblast at the dorsal lip of the anus of Rusconi. (2) The continuous conversion of indifferent lower layer cells into hypoblast, which gradually extends backwards towards the segmentation cavity, and exactly represents the course of the invagination whereby in Amphibians the dorsal wall of the alimentary cavity is formed. (3) The obliteration of the segmentation cavity during the period when the pseudo-invagination is occurring.

The asymmetry of the gastrula or pseudo-gastrula in Cyclostomes, Amphibians, Elasmobranchii and, I believe, Osseous Fishes, is to be explained by the form of the vertebrate body. In Amphioxus, where the small amount of food-yolk present is distributed uniformly, there is no reason why the invagination and resulting gastrula should not be symmetrical. In other vertebrates, where more food-yolk is present, the shape and structure of the body render it necessary for the food-yolk to be stored away on the ventral side of the alimentary canal. This, combined with the unsymmetrical position of the anus, which primitively corresponds in position with the blastopore or anus of Rusconi, causes the asymmetry of the gastrula invagination, since it is not possible for the part of the ovum which will become the ventral wall of the alimentary canal, and which is loaded with food-yolk, to be invaginated in the same fashion as the dorsal wall. From the asymmetry, so caused, follow a large number of features in vertebrate development, [Pg 277] which have been worked out in some detail in my paper already quoted[160].

Prof. Haeckel, in a paper recently published[161], appears to imply that because I do not find absolute invagination in Elasmobranchii, I therefore look upon Elasmobranchii as militating against his Gastræa theory. I cannot help thinking that Prof. Haeckel must have somewhat misunderstood my meaning. The importance of the Gastræa theory has always appeared to me to consist not in the fact that an actual ingrowth of certain cells occurs—an ingrowth which might have many different meanings[162]—but in the fact that the types of early development of all animals can be easily derived from that of the typical gastrula. I am perfectly in accordance with Professor Haeckel in regarding the type of Elasmobranch development to be a simple derivative from that of the gastrula, although believing it to be without any true ingrowth or invagination of cells.

Professor Haeckel[163] in the paper just referred to published his view upon the mutual relationships of the various vertebrate blastoderms. In this paper, which appeared but shortly after my own[164] on the same subject, he has put forward views which differ from mine in several important details. Some of these bear upon the nature of food-yolk; and it appears to me that Professor Haeckel's scheme of development is incompatible with the view that the food-yolk in meroblastic eggs is the homologue of part of the hypoblast of the holoblastic eggs.

The following is Professor Haeckel's own statement of the scheme or type, which he regards as characteristic of meroblastic eggs, pp. 98 and 99.

Jetzt folgt der höchst wichtige und interessante Vorgang, den ich als Einstülpung der Blastula auffasse und der zur Bildung der Gastrula führt (Fig. 63, 64)[165]. Es schlägt sich nämlich der verdickte Saum der Keimscheibe, der Randwulst oder das Properistom, nach innen um und eine dünne Zellenschicht wächst als directe Fortsetzung desselben, wie ein immer [Pg 278] enger werdendes Diaphragma, in die Keimhöhle hinein. Diese Zellenschicht ist das entstehende Entoderm (Fig. 64 i, 74 i). Die Zellen, welche dieselbe zusammensetzen und aus dem innern Theile des Randwulstes hervorwachsen, sind viel grösser aber flacher als die Zellen der Keimhöhlendecke und zeigen ein dunkleres grobkörniges Protoplasma. Auf dem Boden der Keimhöhle, d. h. also auf der Eiweisskugel des Nahrungsdotters, liegen sie unmittelbar auf und rücken hier durch centripetale Wanderung gegen dessen Mitte vor, bis sie dieselbe zuletzt erreichen und nunmehr eine zusammenhängende einschichtige Zellenlage auf dem ganzen Keimhöhlenboden bilden. Diese ist die erste vollständige Anlage des Darmblatts, Entoderms oder Hypoblasts, und von nun an können wir, im Gegensatz dazu den gesammten übrigen Theil des Blastoderms, nämlich die mehrschichtige Wand der Keimhöhlendecke als Hautblatt, Exoderm oder Epiblast bezeichnen. Der verdickte Randwulst (Fig. 64 w, 74 w), in welchem beide primäre Keimblätter in einander übergehen, besteht in seinem oberen und äusseren Theile aus Exodermzellen, in seinem unteren und inneren Theile aus Entodermzellen.

In diesem Stadium entspricht unser Fischkeim einer Amphiblastula, welche mitten in der Invagination begriffen ist, und bei welcher die entstehende Urdarmhöhle eine grosse Dotterkugel aufgenommen hat. Die Invagination wird nunmehr dadurch vervollständigt und die Gastrulabildung dadurch abgeschlossen, dass die Keimhöhle verschwindet. Das wachsende Entoderm, dem die Dotterkugel innig anhängt, wölbt sich in die letztere hinein und nähert sich so dem Exoderm. Die klare Flüssigkeit in der Keimhöhle wird resorbirt und schliesslich legt sich die obere convexe Fläche des Entoderms an die untere concave des Exoderms eng an: die Gastrula des discoblastischen Eies oder die Discogastrula ist fertig (Fig. 65, 76; Meridiandurchschnitt Fig. 66, 75).

Die Discogastrula unsers Knochenfisches in diesem Stadium der vollen Ausbildung stellt nunmehr eine kreisrunde Kappe dar, welche wie ein gefüttertes Mützchen fast die ganze obere Hemisphäre der hyalinen Dotterkugel eng anliegend bedeckt (Fig. 65). Der Ueberzug des Mützchens entspricht dem Exoderm (e), sein Futter dem Entoderm (i). Ersteres besteht aus drei Schichten von kleineren Zellen, letzteres aus einer einzigen Schicht von grösseren Zellen. Die Exodermzellen (Fig. 77) messen 0.006 - 0.009 Mm., und haben ein klares, sehr feinkörniges Protoplasma. Die Entodermzellen (Fig. 78) messen 0.02 - 0.03 Mm. und ihr Protoplasma ist mehr grobkörnig und trüber. Letztere bilden auch den grössten Theil des Randwulstes, den wir nunmehr als Urmundrand der Gastrula, als Properistoma oder auch als Rusconi'schen After bezeichnen können. Der letztere umfasst die Dotterkugel, welche die ganze Urdarmhöhle ausfüllt und weit aus der dadurch verstopften Urmund-Oeffnung vorragt.

My objections to the view so lucidly explained in the passage just quoted, fall under two heads.

[Pg 279] (1) That the facts of development of the meroblastic eggs of vertebrates, are not in accordance with the views here advanced.

(2) That even if these views be accepted as representing the actual facts of development, the explanation offered of these facts would not be satisfactory.

* * * * *

Professor Haeckel's views are absolutely incompatible with the facts of Elasmobranch development, if my investigations are correct.

The grounds of the incompatibility may be summed up under the following heads:

(1) In Elasmobranchii the hypoblast cells occupy, even before the close of segmentation, the position which, on Professor Haeckel's view, they ought only eventually to take up after being involuted from the whole periphery of the blastoderm.

(2) There is no sign at any period of an invagination of the periphery of the blastoderm, and the only structure (the embryonic rim) which could be mistaken for such an invagination is confined to a very limited arc.

(3) The growth of cells to form the floor of the segmentation cavity, which ought to be part of this general invagination from the periphery, is mainly due to a formation of cells from the yolk.

It is this ingrowth of cells for the floor of the segmentation cavity which, I am inclined to think, Professor Haeckel has mistaken for a general invagination in the Osseous Fish he has investigated.

(4) Professor Haeckel fails to give an account of the asymmetry of the blastoderm; an asymmetry which is unquestionably also present in the blastoderm of most Osseous Fishes, though not noticed by Professor Haeckel in the investigations recorded in his paper.

The facts of development of Osseous Fishes, upon which Professor Haeckel rests his views, are too much disputed, for their [Pg 280] discussion in this place to be profitable[166]. The eggs of Osseous Fishes appear to me unsatisfactory objects for the study of this question, partly on account of all the cells of the blastoderm being so much alike, that it is a very difficult matter to distinguish between the various layers, and, partly, because there can be little question that the eggs of existing Osseous Fishes are very much modified, through having lost a great part of the food-yolk possessed by the eggs of their ancestors[167]. This disappearance of the food-yolk must, without doubt, have produced important changes in development, which would be especially marked in a pelagic egg, like that investigated by Professor Haeckel.

The Avian egg has been a still more disputed object than even the egg of the Osseous Fishes. The results of my own investigations on this subject do not accord with those of Dr Götte, or the views of Professor Haeckel[168].

Apart from disputed points of development, it appears to me that a comparative account of the development of the meroblastic [Pg 281] vertebrate ova ought to take into consideration the essential differences which exist between the Avian and Piscian blastoderms, in that the embryo is situated in the centre of the blastoderm in the first case and at the edge in the second[169].

This difference entails important modifications in development, and must necessarily affect the particular points under discussion. As a result of the different positions of the embryo in the two cases, there is present in Elasmobranchii and Osseous Fishes a true anus of Rusconi, or primitive opening into the alimentary canal, which is absent in Birds. Yet in neither Elasmobranchii[170] nor Osseous Fishes does the anus of Rusconi correspond in position with the point where the final closing in of the yolk takes place, but in them this point corresponds rather with the blastopore of Birds[171].

Owing also to the respective situations of the embryo in the [Pg 282] blastoderm, the alimentary and neural canals communicate posteriorly in Elasmobranchii and Osseous Fishes, but not in Birds. Of all these points Professor Haeckel makes no mention.

The support of his views which Prof. Haeckel attempts to gain from Götte's researches in Mammalia is completely cut away by the recent discoveries of Van Beneden[172] and Hensen[173].

It thus appears that Professor Haeckel's views but ill accord with the facts of vertebrate development; but even if they were to do so completely it would not in my opinion be easy to give a rational explanation of them.

Professor Haeckel states that no sharp and fast line can be drawn between the types of 'unequal' and 'discoidal' segmentation[174]. In the cases of unequal segmentation he admits, as is certainly the case, that the larger yolk cells (hypoblast) are simply enclosed by a growth of the epiblast around them; which is to be looked on as a modification of the typical gastrula invagination, necessitated by the large size of the yolk cells (vide Professor Haeckel's paper, Taf. II. fig. 30). In these instances there is no commencement of an ingrowth in the manner supposed for meroblastic ova.

When the food-yolk becomes more bulky, and the hypoblast does not completely segment, it is not easy to understand why an ingrowth, which had no existence in the former case, should occur; nor where it is to come from. Such an ingrowth as is supposed to exist by Professor Haeckel would, in fact, break the continuity of development between meroblastic and holoblastic ova, and thus destroy one of the most important results of the Gastræa theory.

It is quite easy to suppose, as I have done, that in the cases of discoidal segmentation, the hypoblast (including the yolk) becomes enclosed by the epiblast in precisely the same manner as in the cases of unequal segmentation.

But even if Professor Haeckel supposes that in the unsegmented food-yolk a fresh element is added to the ovum, it [Pg 283] remains quite unintelligible to me how an ingrowth of cells from a circumferential line, to form a layer which had no previous existence, can be equivalent to, or derived from, the invagination of a layer, which exists before the process of invagination begins, and which remains continuous throughout it.

If Professor Haeckel's views should eventually turn out to be in accordance with the facts of vertebrate development, it will, in my opinion, be very difficult to reduce them into conformity with the Gastræa theory.

Although some space has been devoted to an attempt to refute the views of Professor Haeckel on this question, I wish it to be clearly understood that my disagreement from his opinions concerns matters of detail only, and that I quite accept the Gastræa theory in its general bearings.

* * * * *

Observations upon the formation of the layers in Elasmobranchii have hitherto been very few in number. Those published in my preliminary account of these fishes are, I believe, the earliest[175].

Since then there has been published a short notice on the subject by Dr Alex. Schultz[176]. His observations in the main accord with my own. He apparently speaks of the nuclei of the yolk as cells, and also of the epiblast being more than one cell deep. In Torpedo alone, amongst the genera investigated by me, is the layer of epiblast, at about the age of the last described embryo, composed of more than a single row of cells.

[Pg 284] EXPLANATION OF PLATE 7.

Complete List of Reference Letters.

c. Cells formed in the yolk around the nuclei of the yolk. ep. Epiblast. er. Embryonic ring. es. Embryo swelling. hy. Hypoblast. ll. Lower layer cells. ly. Line separating the yolk from the blastoderm. m. Mesoblast. mg. Medullary groove. n´. Nuclei of yolk. na. Cells to form ventral wall of alimentary canal which have been derived from the yolk. nal. Cells formed around the nuclei of the yolk which have entered the hypoblast. sc. Segmentation cavity. vp. Combined lateral and vertebral plate of mesoblast.

Fig. 1. Longitudinal section of a blastoderm at the first appearance of the segmentation cavity.

Fig. 2. Longitudinal section through a blastoderm after the layer of cells has disappeared from the floor of the segmentation cavity. bd. Large cell resting on the yolk, probably remaining over from the later periods of segmentation. Magnified 60 diameters. (Hardened in chromic acid.)

The section is intended to illustrate the fact that the nuclei form a layer in the yolk under the floor of the segmentation cavity. The roof of the segmentation cavity is broken.

Fig. 2a. Portion of same blastoderm highly magnified, to shew the characters of the nuclei of the yolk n´ and the nuclei in the cells of the blastoderm.

Fig. 2b. Large knobbed nucleus from the same blastoderm, very highly magnified.

Fig. 2c. Nucleus of yolk from the same blastoderm.

Fig. 3. Longitudinal section of blastoderm of same stage as fig. 2. (Hardened in chromic acid.)

Fig. 4. Longitudinal section of blastoderm slightly older than fig. 2. Magnified 45 diameters. (Hardened in osmic acid.)

It illustrates (1) the characters of the epiblast; (2) the embryonic swelling; (3) the segmentation cavity.

Fig. 5. Longitudinal section through a blastoderm at the time of the first appearance of the embryonic rim, and before the formation of the medullary groove. Magnified 45 diameters.

Fig. 5a. Section through the periphery of the embryonic rim of the blastoderm of which fig. 5 represents a section.

Fig. 6. Section through the embryonic rim of a blastoderm somewhat younger than that represented on Pl. 8, fig. B.

Fig. 7. Section through the most projecting portion of the embryonic rim of a blastoderm of the same age as that represented on Pl. 8, fig. B. The section is drawn on a very considerably smaller scale than that on fig. 5. It is intended to illustrate the growth of the embryonic rim and the disappearance of the segmentation cavity.

Fig. 7a. Section through peripheral portion of the embryonic rim of the same blastoderm, highly magnified. It specially illustrates the formation of a cell (c) around a nucleus in the yolk. The nuclei of the blastoderm have been inaccurately rendered by the artist.

[Pg 285] Figs. 8a, 8b, 8c. Three sections of the same embryo. Inserted mainly to illustrate the formation of the mesoblast as two independent lateral masses of cells; only half of each section is represented. 8a is the most posterior of the three sections. In it the mesoblast forms a large mass on each side, imperfectly separated from the hypoblast. In 8b, from the anterior part of the embryo, the main mass of mesoblast is far smaller, and only forms a cap to the hypoblast at the highest point of the medullary fold. In 8c a cap of mesoblast is present, similar to that in 8b, though much smaller. The sections of these embryos were somewhat oblique, and it has unfortunately happened that while in 8a one side is represented, in 8b and 8c the other side is figured, had it not been for this the sections 8b and 8c would have been considerably longer than 8a.

Fig. 9. Longitudinal section of an embryo belonging to a slightly later stage than B.

This section passes through one of the medullary folds. It illustrates the continuity of the hypoblast with the remaining lower layer cells of the blastoderm.

Figs. 10a, 10b, 10c. Three sections of the same embryo belonging to a stage slightly later than B, Pl. 8. The space between the mesoblast and the hypoblast has been made considerably too great in the figures of the three sections.

10a. The most posterior of the three sections. It shews the posterior flatness of the medullary groove and the two isolated vertebral plates.

10b. This section is taken from the anterior part of the same embryo and shews the deep medullary groove and the commencing formation of the ventral wall of the alimentary canal from the nuclei of the yolk.

10c shews the disappearance of the medullary groove and the thinning out of the mesoblast plates in the region of the head.

Fig. 11. Small portion of the blastoderm and the subjacent yolk of an embryo at the time of the first appearance of the medullary groove × 300. It shews two large nuclei of the yolk (n) and the protoplasmic network in the yolk between them; the network is seen to be closer round the nuclei than in the intervening space. There are no areas representing cells around the nuclei.

Fig. 12. Nucleus of the yolk in connection with the protoplasmic network hardened in osmic acid.

Fig. 13. Portion of posterior end of a blastoderm of stage B, shewing the formation of cells around the nuclei of the yolk.

Fig. 14. Section through part of a young Scyllium egg, about 1/15th of an inch in diameter.

nl. Protoplasmic network in yolk. zp. Zona pellucida. ch. Structureless chorion. fep. Follicular epithelium. x. Structureless membrane external to this.

[116] Qy. Journal of Microsc. Science, Oct. 1874. [This Edition, No. V.]

[117] Centr. f. Med. Wiss. No. 38, 1875.

[118] Professor Bambeke (Poissons Osseux, Mém. Acad. Belgique 1875) describes a cavity in the blastoderm of Leuciscus rutilus, which he regards as the true segmentation cavity, but not as identical with the segmentation cavity of Osseous Fishes, usually so called. Its relations are the same as those of my segmentation cavity at this stage. This paper came into my hands at too late a period for me to be able to do more than refer to it in this place.

[119] Loc. cit.

[120] Loc. cit.

[121] Loc. cit.

[122] Qy. Journal of Micros. Science, Oct. 1874. [This Edition, No. V.]

[123] Loc. cit. Probably Dr Schultz, here as in other cases, has mistaken nuclei for cells.

[124] Loc. cit.

[125] Prof. Haeckel (Die Gastrula u. die Eifurchung d. Thiere, Jenaische Zeitschrift, Vol. IX.) has unfortunately copied a figure from my preliminary paper (loc. cit.) (repeated now), which I had carefully avoided using for the purpose of describing the formation of the layers on account of the epiblast cells in the original having been much altered by the chromic acid, as a result of which the whole section gives a somewhat erroneous impression of the condition of the blastoderm at this stage. I take this opportunity of pointing out that the colouration employed by Professor Haeckel to distinguish the layers in this section is not founded on my statements, but is, on the contrary, in entire opposition to them. From the section as represented by Professor Haeckel it might be gathered that I considered the lower layer cells to be divided into two parts, one derived from the epiblast, while the other constituted the hypoblast. Not only is no such division present at this period, but no part of the lower layer cells, or the mesoblast cells into which they become converted, can in any sense whatever be said to be derived from the epiblast.

[126] Entwicklungsgeschichte der Najaden, Sitz. d. k. Akad. Wien, 1875.

[127] Morphologische Jahrbuch, Vol. 1. Heft 3.

[128] Développement des Mammifères, Bul. de l'Acad. de Belgique, XL. No. 12, 1875.

[129] Qy. Journal Microsc. Science, Oct. 1874. [This Edition, No. V.]

[130] Götte, Der Keim d. Forelleneies, Arch. f. Mikr. Anat. Vol. IX.; Haeckel, Die Gastrula u. die Eifurchung d. Thiere, Jenaische Zeitschrift, Bd. IX.

[131] When writing my earlier paper I did not feel so confident about the mode of formation of the hypoblast as I now do, and even doubted the possibility of determining it from sections. The facts now brought forward are I hope sufficient to remove all scepticism on this point.

[132] Owing to the small size of the plates this section has been drawn on a considerably smaller scale than that represented in fig. 5.

[133] Quart. Journ. of Microscop. Science, July, 1875. [This Edition, No. VI.]

[134] Oellacher, Zeit. f. Wiss. Zoologie, Bd. XXIII. Götte, Archiv f. Mikr. Anat. Vol. IX. Haeckel, loc. cit.

[135] This floor appears in most Osseous Fish to be only partially formed. Vide Götte, loc. cit.

[136] Loc. cit.

[137] Loc. cit.

[138] Loc. cit.

[139] Loc. cit.

[140] Loc. cit.

[141] Professor Lieberkühn (Gesellschaft zu Marburg, Jan. 1876) finds in Mammalia a bilateral arrangement of the mesoblast, which he compares with that described by me in Elasmobranchii. In Mammalia, however, he finds the two masses of mesoblast connected by a very thin layer of cells, and is apparently of opinion that a similar thin layer exists in Elasmobranchii though overlooked by me. I can definitely state that, whatever may be the condition of the mesoblast in Mammalia, in Elasmobranchii at any rate no such layer exists.

[142] Loc. cit.

[143] Quart. Journ. of Microsc. Science, Oct., 1874. [This Edition, No. V.]

[144] Embryologische Studien an Würmern u. Arthropoden. Mémoires de l'Acad. S. Pétersbourg. Vol. XIV. 1873.

[145] Archiv für Mikr. Anat. Vol. VII.

[146] Jenaische Zeitschrift, Vol. IX. 1875. A bilateral development of mesoblast, according to Professor Haeckel (loc. cit.), occurs in some Osseous Fish. Hensen, Zeit. für Anat. u. Entw. Vol. 1., has recently described the mesoblast in Mammalia as consisting of independent lateral masses.

[147] Archiv für Mikr. Anat. Vol. XI.

[148] A protoplasmic network resembling in its essential features the one just described has been noticed by many observers in other ova. Fol has figured and described a network or sponge-like arrangement of the protoplasm in the eggs of Geryonia. (Jenaische Zeitschrift, Vol. VII.) Metschnikoff (Zeitschrift f. Wiss. Zoologie, 1874) has demonstrated its presence in the ova of many Siphonophoriæ and Medusæ. Flemming (Entwicklungsgeschichte der Najaden, Sitz. der k. Akad. Wien, 1875) has found it in the ovarian ova of fresh-water mussels (Anodonta and Unio), but regards it as due to the action of reagents, since he fails to find it in the fresh condition. Amongst vertebrates it has been carefully described by Eimer (Archiv für Mikr. Anat., Vol. VIII.) in the ovarian ova of Reptiles. Eimer moreover finds that it is continuous with prolongations from cells of the epithelium of the follicle in which the ovum is contained. According to him remnants of this network are to be met with in the ripe ovum, but are no longer present in the ovum when taken from the oviduct.

[149] Wirbelthiereier mit partieller Dottertheilung. Müller's Arch. 1861.

[150] Erste Anlage des Wirbelthierleibes.

[151] Eierstock u. Ei.

[152] Entwicklungsgeschichte der Unke. The important researches of Götte on the development of the ovum, though meriting the most careful attention, do not admit of discussion in this place.

[153] Annals and Magaz. of Natural History, Vol. XI. 1873, p. 81.

[154] Archiv f. Mikr. Anat. Vol. X.

[155] Quart. Journ. of Micr. Science, Oct. 1874.

[156] Archiv f. Mikr. Anat. Vol. XXI.

[157] Comparison, &c., Quart. Journ. Micr. Science, July, 1875. [This Edition, No. VI.]

[158] Loc. cit.

[159] This figure, together with figs. 2 and 3, are reproduced from my paper upon the comparison of the early stages of development in vertebrates.

[160] Quart. Journ. of Micr. Science, July, 1875. [This Edition, No. VI.]

[161] Die Gastrula u. Eifurchung d. Thiere, Jenaische Zeitschrift, Vol. IX.

[162] For instance, in Crustaceans it does not in some cases appear certain whether an invagination is the typical gastrula invagination, or only an invagination by which, at a period subsequent to the gastrula invagination, the hind gut is frequently formed.

[163] Loc. cit.

[164] Loc. cit.

[165] The references in this quotation are to the figures in the original.

[166] A short statement by Kowalevsky on this subject in a note to his account of the development of Ascidians, would seem to indicate that the type of development of Osseous Fishes is precisely the same as that of Elasmobranchii. Kowalevsky says, Arch. f. Mikr. Anat. Vol. VII. p. 114, note 5, According to my observations on Osseous Fishes the germinal wall consists of two layers, an upper and lower, which are continuous with one another at the border. From the upper one develops skin and nervous system, from the lower hypoblast and mesoblast. This statement, which leaves unanswered a number of important questions, is too short to serve as a basis for supporting my views, but so far as it goes its agreement with the facts of Elasmobranch development is undoubtedly striking.

[167] The eggs of the Osseous Fishes have, I believe, undergone changes of the same character, but not to the same extent, as those of Mammalia, which, according to the views expressed both by Professor Haeckel and myself, are degenerated from an ovum with a large food-yolk. The grounds on which I regard the eggs of Osseous Fishes as having undergone an analogous change, are too foreign to the subject to be stated here.

[168] I find myself unable without figures to understand Dr Rauber's (Centralblatt für Med. Wiss. 1874, No. 50; 1875, Nos. 4 and 17) views with sufficient precision to accord to them either my assent or dissent. It is quite in accordance with the view propounded in my paper (loc. cit.) to regard, with Dr Rauber and Professor Haeckel, the thickened edge of the blastoderm as the homologue of the lip of the blastopore in Amphioxus; though an invagination, in the manner imagined by Professor Haeckel, is no necessary consequence of this view. If Dr Rauber regards the whole egg of the bird as the homologue of that of Amphioxus, and the inclosure of the yolk by the blastoderm as the equivalent to the process of invagination in Amphioxus, then his views are practically in accordance with my own.

[169] I have suggested in a previous paper (Comparison, &c., Quart. Journal of Micr. Science, July, 1875) that the position occupied by the embryo of Birds at the centre, and not at the periphery, of the blastoderm may be due to an abbreviation of the process by which the Elasmobranch embryos cease to be situated at the edge of the blastoderm (vide p. 296 and Pl. 9, fig. 1, 2). Assuming this to be the real explanation of the position of the embryo in Birds, I feel inclined to repeat a speculation which I made some time ago with reference to the primitive streak in Birds (Quart. Journ. of Micr. Science, 1873, p. 280). In Birds there is, as is well known, a structure called the primitive streak, which has been shewn by the observations of Dursy, corroborated by my observations (loc. cit.), to be situated behind the medullary groove, and to take no part in the formation of the embryo. I further shewed that the peculiar fusion of epiblast and mesoblast, called by His the axis cord, was confined to this structure and did not occur in other parts of the blastoderm. Nearly similar results have been recently arrived at by Hensen with reference to the primitive streak in Mammals. The position of the primitive streak immediately behind the embryo suggests the speculation that it may represent the line along which the edges of the blastoderm coalesced, so as to give to the embryo the central position which it has in the blastoderms of Birds and Mammals, and that the peculiar fusion of epiblast and mesoblast at this point may represent the primitive continuity of epiblast and lower layer cells at the dorsal lip of the anus of Rusconi in Elasmobranchii. I put this speculation forward as a mere suggestion, in the hope of elucidating the peculiar structure of the primitive streak, which not improbably may be found to be the keystone to the nature of the blastoderm of the higher vertebrates.

[170] Vide p. 296 and Plate 9, fig. 1 and 2, and Self, Comparison, &c., loc. cit.

[171] The relation of the anus of Rusconi and blastopore in Elasmobranchii was fully explained in the paper above quoted. It was there clearly shewn that neither the one nor the other exactly corresponds with the blastopore of Amphioxus, but that the two together do so. Professor Haeckel states that in the Osseous Fish investigated by him the anus of Rusconi and the blastopore coincide. This is not the case in the Salmon.

[172] Développement Embryonnaire des Mammifères, Bulletin de l'Acad. r. d. Belgique, 1875.

[173] Loc. cit.

[174] For an explanation of these terms, vide Prof. Haeckel's original paper or the abstract in Quart. Journ. of Micr. Science for January, 1876.

[175] I omit all reference to a paper published in Russian by Prof. Kowalevsky. Being unable to translate it, and the illustrations being too meagre to be in themselves of much assistance, it has not been possible for me to make any use of it.

[176] Centralblatt f. Med. Wiss. No. 33, 1875.

[Pg 286]

CHAPTER IV.

The General Features of the Elasmobranch Embryo at Successive Stages.

No complete series of figures, representing the various stages in development of an Elasmobranch Embryo, has hitherto been published. With the view of supplying this deficiency Plate 8 has been inserted. The embryos represented in this Plate form a fairly complete series, but do not all belong to a single species. Figs. A, B, C, D, E, F, H, I represent embryos of Pristiurus; G being an embryo of Torpedo. The remaining figures, excepting K, which is a Pristiurus embryo, are embryos of Scyllium canicula. The embryos A-I were very accurately drawn from nature by my sister, Miss A. B. Balfour. Unfortunately the exceptional beauty and clearness of the originals is all but lost in the lithographs. To facilitate future description, letters will be employed in the remainder of these pages to signify that an embryo being described is of the same age as the embryo on this Plate to which the letter used refers. Thus an embryo of the same age as L will be spoken of hereafter as belonging to stage L.

A.

This figure represents a hardened blastoderm at a stage when the embryo-swelling (e.s.) has become obvious, but before the appearance of the medullary groove. The position of the segmentation cavity is indicated by a slight swelling of the blastoderm (s.c). The shape of the blastoderm, in hardened specimens, is not to be relied upon, owing to the traction which the blastoderm undergoes during the process of removing the yolk from the egg-shell.

B.

B is the view of a fresh blastoderm. The projecting part of this, already mentioned as the 'embryonic rim', is indicated [Pg 287] by the shading. At the middle of the embryonic rim is to be seen the rudiment of the embryo (m.g.). It consists of an area of the blastoderm, circumscribed on its two sides and at one end, by a slight fold, and whose other end forms part of the edge of the blastoderm. The end of the embryo which points towards the centre of the blastoderm is the head end, and that which forms part of the edge of the blastoderm is the tail end. To retain the nomenclature usually adopted in treating of the development of the Bird, the fold at the anterior end of the embryo may be called the head fold, and those at the sides the side folds. There is in Elasmobranchii no tail fold, owing to the position of the embryo at the periphery of the blastoderm, and it is by the meeting of the three above-mentioned folds only, that the embryo becomes pinched off from the remainder of the blastoderm. Along the median line of the embryo is a shallow groove (m.g.), the well-known medullary groove of vertebrate embryology. It flattens out both anteriorly and posteriorly, and is deepest in the middle part of its course.

C.

This embryo resembles in most of its features the embryo last described. It is, however, considerably larger, and the head fold and side folds have become more pronounced structures. The medullary groove is far deeper than in the earlier stage, and widens out anteriorly. This anterior widening is the first indication of a distinction between the brain and the remainder of the central nervous system, a distinction which arises long before the closure of the medullary canal.

D.

This embryo is far larger than the one last described, but the increase in length does not cause it to project beyond the edge of the blastoderm, but has been due to a growth inwards towards the centre of the blastoderm. The head is now indicated by an anterior enlargement, and the embryo also widens out posteriorly. The posterior widening (t.s.) is formed by a pair of rounded prominences, one on each side of the middle line. These are very conspicuous organs during the earlier stages of development, and consist of two large aggregations of mesoblast cells. [Pg 288] In accordance with the nomenclature adopted in my preliminary paper[177], they may be called 'tail-swellings'. Between the cephalic enlargements and the tail-swellings is situated the rudimentary trunk of the embryo. It is more completely pinched off from the blastoderm than in the last described embryo. The medullary groove is of a fairly uniform size throughout the trunk of the embryo, but flattens out and vanishes completely in the region of the head. The blastoderm in Pristiurus and Scyllium grows very rapidly, and has by this stage attained a very considerable size; but in Torpedo its growth is very slow.

E and F.

These two embryos may be considered together, for, although they differ in appearance, yet they are of an almost identical age; and the differences between the two are purely external. E appears to be a little abnormal in not having the cephalic region so distinctly marked off from the trunk as is usual. The head is proportionally larger than in the last stage, and the tail-swellings remain as conspicuous as before. The folding off from the blastoderm has progressed rapidly, and the head and tail are quite separated from it. The medullary groove has become closed posteriorly in both embryos, but the closing has extended further forwards in F than in E. In F the medullary folds have not only united posteriorly, but have very nearly effected a fresh junction in the region of the neck. At this point a second junction of the two medullary folds is in fact actually effected before the posterior closing has extended forwards so far. The later junction in the region of the neck corresponds in position with the point, where in the Bird the medullary folds first unite. No trace of a medullary groove is to be met with in the head, which simply consists of a wide flattened plate. Between the two tail-swellings surface views present the appearance of a groove, but this appearance is deceptive, since in sections no groove, or at most a very slight one, is perceptible.

G.

During the preceding stages growth in the embryo is very slow, and considerable intervals of time elapse before any [Pg 289] perceptible changes are effected. This state of things now becomes altered, and the future changes succeed each other with far greater rapidity. One of the most important of these, and one which first presents itself during this stage, is the disappearance of the yolk-spherules from the embryonic cells, and the consequently increased transparency of the embryo. As a result of this, a number of organs, which in the earlier stages were only to be investigated by means of sections, now become visible in the living embryo.

The tail-swellings (t.s.) are still conspicuous objects at the posterior extremity of the embryo. The folding off of the embryo from the yolk has progressed to such an extent that it is now quite possible to place the embryo on its side and examine it from that point of view.

The embryo may be said to be attached to the yolk by a distinct stalk or cord, which in the succeeding stages gradually narrows and elongates, and is known as the umbilical cord (so.s.). The medullary canal has now become completely closed, even in the region of the brain, where during the last stage no trace of a medullary groove had appeared. Slight constrictions, not perceptible in views of the embryo as a transparent object, mark off three vesicles in the brain. These vesicles are known as the fore, mid, and hind brain. From the fore-brain there is an outgrowth on each side, the first rudiment of the optic vesicle (op.).

The mesoblast on each side of the body is divided into a series of segments, known as protovertebræ or muscle-plates, the first of which lies a little behind the head. The mesoblast of the tail has not as yet undergone this segmentation. There are present in all seventeen segments. These first appeared at a much earlier date, but were not visible owing to the opacity of the embryo.

Another structure which became developed in even a younger embryo than C is now for the first time visible in the living embryo. This is the notochord: it extends from almost the extreme posterior to the anterior end of the embryo. It lies between the ventral wall of the spinal canal and the dorsal wall of the intestine; and round its posterior end these two walls become continuous with each other (vide fig.). Anteriorly the [Pg 290] termination of the notochord cannot be seen, it can only be traced into a mass of mesoblast at the base of the brain, which there separates the epiblast from the hypoblast. The alimentary canal (al.) is completely closed anteriorly and posteriorly, though still widely open to the yolk-sac in the middle part of its course. In the region of the head it exhibits on each side a slight bulging outwards, the rudiment of the first visceral cleft. This is represented in the figure by two lines (I v.c.). The visceral clefts at this stage consist of a pair of simple diverticula from the alimentary canal, and there is no communication between the throat and the exterior.

H.

The present embryo is far larger than the last, but it has not been possible to represent this increase in size in the drawings. Accompanying this increase in size, the folding off of the embryo from the yolk has considerably progressed, and the stalk which unites the embryo with the yolk is proportionately narrower and longer than before.

The brain is now very distinctly divided into the three lobes, whose rudiments appeared during the last stage. From the foremost of these, the optic vesicles now present themselves as well-marked lateral outgrowths, towards which there appears a growing in, or involution, from the external skin (op.) to form the lens. The opening of this involution is represented by the dark spot in the centre.

A fresh organ of sense, the auditory sac, now for the first time becomes visible as a shallow pit in the external skin on each side of the hind-brain (au.v.). The epiblast which is involuted to form this pit becomes much thickened, and thereby the opacity, indicated in the figure, is produced.

The muscle-plates have greatly increased in number by the formation of fresh segments in the tail. Thirty-eight of them were present in the embryo figured. The mesoblast at the base of the brain has increased in quantity, and there is still a certain mass of unsegmented mesoblast which forms the tail-swellings. The first rudiment of the heart becomes visible during this stage as a cavity between the mesoblast of the splanchnopleure and the hypoblast (ht.).

[Pg 291] The fore and hind guts are now longer than they were. A slight pushing in from the exterior to form the mouth has appeared (m.), and an indication of the future position of the anus is afforded by a slight diverticulum of the hind gut towards the exterior some little distance from the posterior end of the embryo (an.). The portion of the alimentary canal behind this point, though at this stage large, and even dilated into a vesicle at its posterior end (al.v.), becomes eventually completely atrophied. In the region of the throat the rudiment of a second visceral cleft has appeared behind the first; neither of them are as yet open to the exterior. The number of visceral clefts present in any given Pristiurus embryo affords a very easy and simple way of determining its age.

I.

A great increase in size is again to be noticed in the embryo, but, as in the case of the last embryo, it has not been possible to represent this in the figure. The stalk connecting the embryo with the yolk has become narrower and more elongated, and the tail region of the embryo proportionately far longer than in the last stage. During this stage the first spontaneous movements of the embryo take place, and consist in somewhat rapid excursions of the embryo from side to side, produced by a serpentine motion of the body.

The cranial flexure, which commenced in stage G, has now become very evident, and the mid-brain[178] begins to project in the same manner as in the embryo fowl on the third day, and will soon form the anterior termination of the long axis of the embryo. The fore-brain has increased in size and distinctness, and the anterior part of it may now be looked on as the unpaired rudiment of the cerebral hemispheres.

Further growths have taken place in the organs of sense, especially in the eye, in which the involution for the lens has made considerable progress. The number of the muscle-plates has again increased, but there is still a region of unsegmented [Pg 292] mesoblast in the tail. The thickened portions of mesoblast which caused the tail-swellings are still to be seen and would seem to act as the reserve from which is drawn the matter for the rapid growth of the tail, which occurs soon after this. The mass of the mesoblast at the base of the brain has again increased. No fresh features of interest are to be seen in the notochord. The heart is now much more conspicuous than before, and its commencing flexure is very apparent. It now beats actively. The hind gut especially is much longer than in the last specimen; and the point where the anus will appear is very easily detected by the bulging out of the gut towards the external skin at that point (an.). The alimentary vesicle, first observable during the last stage, is now a more conspicuous organ (al.v.). Three visceral clefts, none of which are as yet open to the exterior, may now be seen.

K.

The figures G, H, I are representations of living and transparent embryos, but the remainder of the figures are drawings of opaque embryos which were hardened in chromic acid.

The stalk connecting the embryo with the yolk is now, comparatively speaking, quite narrow, and is of sufficient length to permit the embryo to execute considerable movements.

The tail has grown immensely, but is still dilated terminally. This terminal dilatation is mainly due to the alimentary vesicle, but the tract of gut connecting this with the gut in front of the anus is now a solid rod of cells and very soon becomes completely atrophied.

The two pairs of limbs have appeared as elongated ridges of epiblast. The anterior pair is situated just at the front end of the umbilical stalk; and the posterior pair, which is the more conspicuous of the two, is situated some little distance behind the stalk.

The cranial flexure has greatly increased, and the angle between the long axis of the front part of the head and of the body is less than a right angle. The conspicuous mid-brain forms the anterior termination of the long axis of the body. The thin roof of the fourth ventricle may in the figure be noticed behind the mid-brain. The auditory sac is nearly closed and its [Pg 293] opening is not shewn in the figure. In the eye the lens is completely formed.

Owing to the opacity of the embryo, the muscle-plates are only indistinctly indicated, and no other features of the mesoblast are to be seen.

The mouth is now a deep pit, whose borders are almost completely formed by the thickening in front of the first visceral cleft, which may be called the first visceral arch or mandibular arch.

Four visceral clefts are now visible, all of which are open to the exterior, but in a transparent embryo one more, not open to the exterior, would have been visible behind the last of these.

L.

This embryo is considerably older than the one last described, but growth is not quite so rapid as might be gathered from the fact that L is nearly twice as long as K, since the two embryos belong to different genera; and the Scyllium embryos, of which L is an example, are larger than Pristiurus embryos. The umbilical stalk is now quite a narrow elongated structure, whose subsequent external changes are very unimportant, and consist for the most part merely in an increase in its length.

The tail has again grown greatly in length, and its terminal dilatation together with the alimentary vesicle contained in it, have both completely vanished. A dorsal and ventral fin are now clearly visible; they are continuous throughout their whole length. The limbs have grown and are more easily seen than in the previous stage.

Great changes have been effected in the head, resulting in a diminution of the cranial flexure. This diminution is nevertheless apparent rather than real, and is chiefly due to the rapid growth of the rudiment of the cerebral hemispheres. The three main divisions of the brain may still be clearly seen from the surface. Posteriorly is situated the hind-brain, now consisting of the medulla oblongata and cerebellum. At the anterior part of the medulla is to be seen the thin roof of the fourth ventricle, and anteriorly to this again the roof becomes thickened to form the rudiment of the cerebellum. In front of the hind-brain lies the mid-brain, the roof of which is formed by the [Pg 294] optic lobes, which are still situated at the front end of the long axis of the embryo.

Beyond the mid-brain is placed the fore-brain, whose growth is rapidly rendering the cranial flexure imperceptible.

The rudiments of the nasal sacs are now clearly visible as a pair of small pits. The pits are widely open to the exterior, and are situated one on each side, near the front end of the cerebral hemispheres. Five visceral clefts are open to the exterior, and in them the external gills have commenced to appear (L´).

The first cleft is no longer similar to the rest, but has commenced to be metamorphosed into the spiracle.

Accompanying the change in position of the first cleft, the mandibular arch has begun to bend round and enclose the front as well as the side of the mouth. By this change in the mandibular arch the mouth becomes narrowed in an antero-posterior direction.

M.

Of this embryo the head alone has been represented. Two views of it are given, one (M) from the side and the other (M´) from the under surface. The growth of the front part of the head has considerably diminished the prominence of the cranial flexure. The full complement of visceral clefts is now present—six in all. But the first has already atrophied considerably, and may easily be recognized as the spiracle. In Scyllium, there are present at no period more than six visceral clefts. The first visceral arch on each side has become bent still further round, to form the front border of the mouth. The opening of the mouth has in consequence become still more narrowed in an antero-posterior direction. The width of the mouth in this direction, serves for the present and for some of the subsequent stages as a very convenient indication of age.

N.

The limbs, or paired fins, have now acquired the general features and form which they possess in the adult.

The unpaired fins have now also become divided in a manner not only characteristic of the Elasmobranchii but even of the genus Scyllium.

[Pg 295] There is a tail fin, an anal fin and two dorsal fins, both the latter being situated behind the posterior paired fins.

In the head may be noticed a continuation of the rapid growth of the anterior part.

The mouth has become far more narrow and slit-like; and with many other of the organs of the period commences to approach the form of the adult.

The present and the three preceding stages shew the gradual changes by which the first visceral arch becomes converted into the rudiments of the upper and of the lower jaw. The fact of the conversion was first made known through the investigations of Messrs Parker and Gegenbaur.

O.

In this stage the embryo is very rapidly approaching the form of the adult.

This is especially noticeable in the fins, which project in a manner quite characteristic of the adult fish. The mouth is slit-like, and the openings of the nasal sacs no longer retain their primitive circular outline. The external gills project from all the gill-slits including the spiracle.

P.

The head is rapidly elongating by the growth of the snout, and the divisions of the brain can no longer be seen with distinctness from the exterior, and, with the exception of the head and of the external gills, the embryo almost completely resembles the adult.

Q.

The snout has grown to such an extent, that the head has nearly acquired its adult shape. In the form of its mouth the embryo now quite resembles the adult fish.

* * * * *

This part of the subject may be conveniently supplemented by a short description of the manner in which the blastoderm encloses the yolk. It has been already mentioned that the growth of the blastoderm is not uniform. The part of it in the immediate neighbourhood of the embryo remains comparatively stationary, while the growth elsewhere is very rapid. From [Pg 296] this it results that that part of the edge of the blastoderm where the embryo is attached forms a bay in the otherwise regular outline of the edge of the blastoderm. By the time that one-half of the yolk is enclosed the bay is a very conspicuous feature (Pl. 9, fig. 1). In this figure bl. points to the blastoderm, and yk. to the part of the yolk not yet enclosed by the blastoderm.

Shortly subsequent to this the bay becomes obliterated by its two sides coming together and coalescing, and the embryo ceases to lie at the edge of the yolk.

This stage is represented on Pl. 9, fig. 2. In this figure there is only a small patch of yolk not yet enclosed (yk), which is situated at some little distance behind the embryo. Throughout all this period the edge of the blastoderm has remained thickened, a feature which persists till the complete investment of the yolk, which takes place shortly after the stage last figured. In this thickened edge a circular vein arises, which brings back the blood from the yolk-sac to the embryo. The opening in the blastoderm (Pl. 9, fig. 2, yk.), exposing the portion of the yolk not yet enclosed, may be conveniently called the blastopore, according to Professor Lankester's nomenclature.

The interesting feature which characterizes the blastopore in Elasmobranchii is the fact of its not corresponding in position with the opening of the anus of Rusconi. We thus have in Elasmobranchii two structures, each of which corresponds in part with the single structure in Amphioxus which may be called either blastopore or anus of Rusconi, which yet do not in Elasmobranchii coincide in position. It is the blastopore of Elasmobranchii which has undergone a change of position, owing to the unequal growth of the blastoderm; while the anus of Rusconi retains its normal situation. In Osseous Fishes the blastopore undergoes a similar change of position. The possibility of a change in position of this structure is peculiarly interesting, in that it possibly serves to explain how the blastopore of different animals corresponds in different cases with the anus or the mouth, and has not always a fixed situation[179].

[Pg 297] EXPLANATION OF PLATES 8 and 9.

Complete List of Reference Letters.

a. Arteries of yolk sac (red). al. Alimentary cavity. alv. Alimentary vesicle at the posterior end of the alimentary canal. an. Point where anus will appear. auv. Auditory vesicle. bl. Blastoderm. ch. Notochord. es. Embryo-swelling. h. Head. ht. Heart. m. Mouth. mg. Medullary groove. mp. Muscle-plate or protovertebra. op. Eye. sc. Segmentation cavity. sos. Somatic stalk. ts. Tail-swelling. v. Veins of yolk sac (blue). vc. Visceral cleft. I. vc. 1st visceral cleft. x. Portion of blastoderm outside the arterial circle in which no blood-vessels are present. yk. Yolk.

Plate 8.

Fig. A. Surface view of blastoderm of Pristiurus hardened in chromic acid.

Fig. B. Surface view of fresh blastoderm of Pristiurus.

Figs. C, D, E, and F. Pristiurus embryos hardened in chromic acid.

Fig. G. Torpedo embryo viewed as a transparent object.

Figs. H, I. Pristiurus embryos viewed as transparent objects.

Fig. K. Pristiurus embryo hardened in chromic acid.

The remainder of the figures are representations of embryos of Scyllium canicula hardened in chromic acid. In every case, with the exception of the figures marked P and Q, two representations of the same embryo are given; one from the side and one from the under surface.

Plate 9.

Fig. 1. Yolk of a Pristiurus egg with blastoderm and embryo. About two-thirds of the yolk have been enveloped by the blastoderm. The embryo is still situated at the edge of the blastoderm, but at the end of a bay in the outline of this. The thickened edge of the blastoderm is indicated by a darker shading. Two arteries have appeared.

Fig. 2. Yolk of an older Pristiurus egg. The yolk has become all but enveloped by the blastoderm, and the embryo ceases to lie at the edge of the blastoderm, owing to the coalescence of the two sides of the bay which existed in the earlier stage. The circulation is now largely developed. It consists of an external arterial ring, and an internal venous ring, the latter having been developed in the thickened edge of the blastoderm. Outside the arterial ring no vessels are developed.

Fig. 3. The yolk has now become completely enveloped by the blastoderm. The arterial ring has increased in size. The venous ring has vanished, owing to the complete enclosure of the yolk by the blastoderm. The point where it existed is still indicated (y) by the brush-like termination of the main venous trunk in a number of small branches.

Fig. 4. Diagrammatic projection of the vascular system of the yolk sac of a somewhat older embryo.

The arterial ring has grown much larger and the portion of the yolk where no vessels exist is very small (x). The brush-like termination of the venous trunk is still to be noticed.

The two main trunks (arterial and venous) in reality are in close contact as in fig. 5, and enter the somatic stalk close together.

The letter a which points to the venous (blue) trunk should be v and not a.

Fig. 5. Circulation of the yolk sac of a still older embryo, in which the arterial circle has ceased to exist, owing to the space outside it having become smaller and smaller and finally vanished.

[177] Quart. Journ. Micr. Science, Oct. 1874. [This Edition, No. V.]

[178] The part of the brain which I have here called mid-brain, and which unquestionably corresponds to the part called mid-brain in the embryos of higher vertebrates, becomes in the adult what Miklucho-Maclay and Gegenbaur called the vesicle of the third ventricle or thalamencephalon. I shall always speak of it as the mid-brain.

[179] For a fuller discussion of this question vide Self, A comparison of the early stages of development in vertebrates. Quart. Journ. of Micr. Science, July, 1875. [This Edition, No. VI.]

[Pg 298]

CHAPTER V.

Stages B to G.

The present chapter deals with the history of the development of the Elasmobranch embryo from the period when the medullary groove first arises till that in which it becomes completely closed, and converted into the medullary canal. The majority of the observations recorded were made on Pristiurus embryos, a few on embryos of Torpedo. Where nothing is said to the contrary the statements made apply to the embryos of Pristiurus only.

The general external features for this period have already been given in sufficient detail in the last chapter; and I proceed at once to describe consecutively the history of the three layers.

General Features of the Epiblast.

At the commencement of this period, during the stage intermediate between B and C, the epiblast is composed of a single layer of cells. (Pl. 10, fig. 1.)

These are very much elongated in the region of the embryo, but flattened in other parts of the blastoderm. Throughout they contain numerous yolk-spherules.

In a Torpedo embryo of this age (as determined by the condition of the notochord) the epiblast presents a very different structure. It is composed of small spindle-shaped cells several rows deep. The nuclei of these are very large in proportion to the cells containing them, and the yolk-spherules are far less numerous than in the cells of corresponding Pristiurus embryos.

During stage C the condition of the epiblast does not undergo any important change, with the exception of the layer [Pg 299] becoming much thickened, and its cells two or three deep in the anterior parts of the embryo. (Pl. 10, fig. 2.)

In the succeeding stages that part of the epiblast, which will form the spinal cord, gradually becomes two or three cells deep. This change is effected by a decrease in the length of the cells as compared with the thickness of the layer. In the earlier stages the cells are wedge-shaped with an alternate arrangement, so that a decrement in the length of the cells at once causes the epiblast to be composed of two rows of interlocking cells.

The lateral parts of the epiblast which form the epidermis of the embryo are modified in quite a different manner to the nervous parts of the layer, becoming very much diminished in thickness and composed of a single row of flattened cells. (Pl. 10, fig. 3.)

Till the end of stage F, the epiblast cells and indeed all the cells of the blastoderm retain their yolk-spherules, but the epiblast begins to lose them and consequently to become transparent in stage G.

Medullary Groove.

During stage B the medullary groove is shallow posteriorly, deeper in the middle part, and flattened out again at the extreme anterior end of the embryo. (Pl. 7, fig. 10a, b, c.)

A similar condition obtains in the stage between B and C, but the canal has now in part become deeper. Anteriorly no trace of it is to be seen. In stage C it exhibits the same general features. (Pl. 10, fig. 2a, 2b, 2c.)

By stage D we find important modifications of the canal.

It is still shallow behind and deep in the dorsal region, Pl. 10, figs. 3d, 3e, 3f; but the anterior flattened area in the last stage has grown into a round flat plate which may be called the cephalic plate, Pl. 8, D and Pl. 10, figs. 3a, 3b, 3c. This plate becomes converted into the brain. Its size and form give it a peculiar appearance, but the most remarkable feature about it is the ventral curvature of its edges. Its edges do not, as might be expected, bend dorsalwards towards each other, but become sharply bent in a ventral direction. This feature is for the first [Pg 300] time apparent at this stage, but becomes more conspicuous during the succeeding ones, and attains its maximum in stage F (Pl. 10, fig. 5), in which it might almost be supposed that the edges of the cephalic plate were about to grow downwards and meet on the ventral side of the embryo.

In the stages subsequent to D the posterior part of the canal deepens much more rapidly than the rest (vide Pl. 10, fig. 4, taken from the posterior end of an embryo but slightly younger than F), and the medullary folds unite and convert the posterior end of the medullary groove into a closed canal (Pl. 8, fig. F), while the groove is still widely open elsewhere[180]. The medullary canal does not end blindly behind, but simply forms a tube not closed at either extremity. The importance of this fact will appear later.

In a stage but slightly subsequent to F nearly the whole of the medullary canal becomes formed. This occurs in the usual way by the junction and coalescence of the medullary folds. In the course of the closing of the medullary groove the edges of the cephalic plate lose their ventral curvature and become bent up in the normal manner (vide Pl. 10, fig. 6, a section taken through the posterior part of the cephalic plate), and the enlarged plate merely serves to enclose a dilated cephalic portion of the medullary canal. The closing of the medullary canal takes place earlier in the head and neck than in the back. The anterior end of the canal becomes closed and does not remain open like the posterior end.

Elasmobranch embryos resemble those of the Sturgeon (Acipenser) and the Amphibians in the possession of a spatula-like cephalic expansion: but so far as I am aware a ventral flexure in the medullary plates of the head has not been observed in other groups.

The medullary canal in Elasmobranchii is formed precisely on the type so well recognised for all groups of vertebrates with the exception of the Osseous Fishes. The only feature in any respect peculiar to these fishes is the closing of their medullary canal first commencing behind, and then at a second point in the [Pg 301] cervical region. In those vertebrates in which the medullary folds do not unite at approximately the same time throughout their length, they appear usually to do so first in the region of the neck.

Mesoblast.

The separation from the hypoblast of two lateral masses of mesoblast has already been described. Till the close of stage C the mesoblast retains its primitive bilateral condition unaltered. Throughout the whole length of the embryo, with the exception of the extreme front part, there are present two plates of rounded mesoblast cells, one on each side of the medullary groove. These plates are in very close contact with the hypoblast, and also follow with fair accuracy the outline of the epiblast. This relation of the mesoblast plates to the epiblast must not however be supposed to indicate that the medullary groove is due to growth in the mesoblast: a view which is absolutely negatived by the manner of formation of the medullary groove in the head. Anteriorly the mesoblast plates thin out and completely vanish.

In stage D, the plates of mesoblast in the trunk undergo important changes. The cells composing them become arranged in two layers (Pl. 10, fig. 3), a splanchnic layer adjoining the hypoblast (sp), and a somatic layer adjoining the epiblast[181] (so). Although these two layers are distinctly formed, they do not become separated at this stage in the region of the trunk, and in the trunk no true body-cavity is formed.

By stage D the plates of mesoblast have ceased to be quite isolated, and are connected with the lower layer cells of the general blastoderm.

Moreover the lower layer cells outside the embryo now exhibit distinct traces of a separation into two layers, one continuous with the hypoblast, the other with the mesoblast. Both layers are composed of very flattened cells, and the mesoblast layer is often more than one cell deep, and sometimes exhibits a mesh-like arrangement of its elements.

[Pg 302] Coincidentally with the appearance of a differentiation into a somatic and splanchnic layer the mesoblast plates become partially split by a series of transverse lines of division into protovertebræ. Only the proximal regions of the plates become split in this way, while their peripheral parts remain quite intact. As a result of this each plate becomes divided into a proximal portion adjoining the medullary canal, which is divided into protovertebræ, and may be called the vertebral plate, and a peripheral portion not so divided, which may be called the lateral plate. These two parts are at this stage quite continuous with each other; and, as will be seen in the sequel, the body-cavity originally extends uninterruptedly to the summit of the vertebral plates.

By stage D at the least ten protovertebræ have appeared.

In Torpedo the mesoblast commences to be divided into two layers much earlier than in Pristiurus; and even before stage C this division is more or less clearly marked.

In the head and tail the condition of the mesoblast is by no means the same as in the body.

In the tail the plates of mesoblast become considerably thickened and give rise to two projections, one on each side, which have already been alluded to as caudal or tail-swellings; vide Pl. 8, figs. D, F, and Pl. 10, fig. 3f and fig. 4, ts.

These masses of mesoblast are neither divided into protovertebræ, nor do they exhibit any trace of a commencing differentiation into somatopleure and splanchnopleure.

In the head, so far as I have yet been able to observe, the mesoblastic plates do not at this stage become divided into protovertebræ. The other changes exhibited in the cephalic region are of interest, mainly from the fact that here appears a cavity in the mesoblast directly continuous with the body-cavity (when that cavity becomes formed), but which appears at a very much earlier date than the body-cavity. This cavity can only be looked on in the light of a direct continuation of the body or peritoneal cavity into the head. Theoretical considerations with reference to it I propose reserving till I have described the changes which it undergoes in the subsequent periods.

Pl. 10, figs. 3a, 3b and 3c exhibit very well the condition of the mesoblast in the head at this period. In fig. 3c, a section [Pg 303] taken through the back part of the head, the mesoblast plates have nearly the same form as in the sections immediately behind. The ventral continuation of the mesoblast formed by the lateral plate has, however, become much thinner, and the dorsal or vertebral portion has acquired a more triangular form than in the sections through the trunk (figs. 3d and 3e).

In the section (fig. 3b) in front of this the ventral portion of the plate is no longer present, and only that part exists which corresponds with the vertebral division of the primitive plate of mesoblast.

In this a distinct cavity, forming part of the body-cavity, has appeared.

In a still anterior section (fig. 3a) no cavity is any longer present in the mesoblast; whilst in sections taken from the foremost part of the head no mesoblast is to be seen (vide Pl. 10, fig. 5, taken from the front part of the head of the embryo represented in Pl. 8, fig. F).

A continuation of the body-cavity into the head has already been described by Oellacher[182] for the Trout: but he believes that the cavity in this part is solely related to the formation of the pericardial space.

The condition of the mesoblast undergoes no important change till the end of the period treated of in this chapter. The masses of mesoblast which form the tail-swellings become more conspicuous (Pl. 10, fig. 4); and indeed their convexity is so great that the space between them has the appearance of a median groove, even after the closure of the neural canal in the caudal region.

In embryos of stage G, which may be considered to belong to the close of this period, eighteen protovertebræ are present both in Pristiurus and Torpedo embryos.

The Alimentary Canal.

The alimentary canal at the commencement of this period (stage B) forms a space between the embryo and the yolk, ending blindly in front, but opening posteriorly by a widish slit-like aperture, which corresponds to the anus of Rusconi (Pl. 7, fig. 7).

[Pg 304] The cavity anteriorly has a more or less definite form, having lateral walls, as well as a roof and floor (Pl. 7, figs. 10b and 10c). Posteriorly it is not nearly so definitely enclosed (Pl. 7, fig. 10a). The ventral wall of the cavity is formed by yolk. But even in stage B there are beginnings of a cellular ventral wall derived from an ingrowth of cells from the two sides.

By stage C considerable progress has been made in the formation of the alimentary canal. Posteriorly it is as flattened and indefinite as during stage B (Pl. 10, figs. 2b and 2c). But in the anterior part of the embryo the cavity becomes much deeper and narrower, and a floor of cells begins to be formed for it (Pl. 10, fig. 2); and, finally, in front, it forms a definite space completely closed in on all sides by cells (Pl. 10, fig. 2a). Two distinct processes are concerned in effecting these changes in the condition of the alimentary cavity. One of these is a process of folding off the embryo from the blastoderm. The other is a simple growth of cells independent of any folding. To the first of these processes the depth and narrowness of the alimentary cavity is due; the second is concerned in forming its ventral wall. The combination of the two processes produces the peculiar triangular section which characterises the anterior closed end of the alimentary cavity at this stage. The process of the folding off of the embryo from the blastoderm resembles exactly the similar process in the embryo bird. The fold by which the constricting off of the embryo is effected is a perfectly continuous one, but may be conveniently spoken of as composed of a head fold and two lateral folds.

Of far greater interest than the nature of these folds is the formation of the ventral wall of the alimentary canal. This, as has been said, is effected by a growth of cells from the two sides to the middle line (Pl. 10, fig. 2). The cells for this are however not derived from pre-existing hypoblast cells, but are formed spontaneously around nuclei of the yolk. This fact can be determined in a large number of sections, and is fairly well shewn in Pl. 10, fig. 2, na. The cells are formed in the yolk, as has been already mentioned, by a simple aggregation of protoplasm around pre-existing nuclei.

The cells being described are in most cases formed close to the pre-existing hypoblast cells, but often require to undergo a [Pg 305] considerable change of position before attaining their final situation in the wall of the alimentary canal.

I have already alluded to this feature in the formation of the ventral wall of the alimentary cavity. Its interest, as bearing on the homology of the yolk, is considerable, owing to the fact that the so-called yolk-cells of Amphibians play a similar part in supplying the ventral epithelium of the alimentary cavity, as do the cells derived from the yolk in Elasmobranchii.

The fact of this feature being common to the yolk-cells of Amphibians and the yolk of Elasmobranchii, supplies a strong argument in favour of the homology of the yolk-cells in the one case with the yolk in the other[183].

[Pg 306] The history of the alimentary canal during the remainder of this period may be told briefly.

The folding off and closing of the alimentary canal in the anterior part of the body proceeds rapidly, and by stage D not only is a considerable tract of alimentary canal formed, but a great part of the head is completely folded off from the yolk (Pl. 10, fig. 3a). By stage F a still greater part is folded off. The posterior part of the alimentary canal retains for a long period its primitive condition. It is not until stage F that it begins to be folded off behind. After the folding has once commenced it proceeds with great rapidity, and before stage G the hinder part of the alimentary canal becomes completely closed in.

The folding in of the gut is produced by two lateral folds, and the gut is not closed posteriorly.

It may be remembered that the neural canal also remained open behind. Thus both the neural and alimentary canals are open behind; and, since both of them extend to the posterior [Pg 307] end of the body, they meet there, their walls coalesce, and a direct communication from the neural to the alimentary canal is instituted. The process may be described in another way by saying that the medullary folds are continuous round the end of the tail with the lateral walls of the alimentary canal; so that, when the medullary folds unite to form a canal, this canal becomes continuous with the alimentary canal, which is closed in at the same time. In whatever way this arrangement is produced, the result of it is that it becomes possible to pass in a continuously closed passage along the neural canal round the end of the tail and into the alimentary canal. A longitudinal section shewing this feature is represented on Pl. 10, fig. 7.

This communication between the neural and alimentary canals, which is coupled, as will be seen in the sequel, with the atrophy of a posterior segment of the alimentary canal, is a feature of great interest which ought to throw considerable light upon the meaning of the neural canal. So far as I know, no suggestion as to the origin of it has yet been made. It is by no means confined to Elasmobranchii, but is present in all the vertebrates whose embryos are situated at the centre and not at the periphery of the blastoderm. It has been described by Goette[184] in Amphibians and by Kowalevsky, Owsjannikow and Wagner[185] in the Sturgeon (Acipenser). The same arrangement is also stated by Kowalevsky[186] to exist in Osseous Fishes and Amphioxus. The same investigator has shewn that the alimentary and neural canals communicate in larval Ascidians, and we may feel almost sure that they do so in the Marsipobranchii.

The Reptilia, Aves, and Mammalia have usually been distinguished from other vertebrates by the possession of a well-developed allantois and amnion. I think that we may further say that the lower vertebrates, Pisces and Amphibia, are to be distinguished from the three above-mentioned groups of higher [Pg 308] vertebrates, by the positive embryonic character that their neural and alimentary canals at first communicate posteriorly. The presence or absence of this arrangement depends on the different positions of the embryo in the blastoderm. In Reptiles, Birds and Mammals, the embryo occupies a central position in the blastoderm, and not, as in Pisces and Amphibia, a peripheral one at its edge. We can, in fact, only compare the blastoderm of the Bird and the Elasmobranch, by supposing that in the blastoderm of the Bird there has occurred an abbreviation of the processes, by which the embryo Elasmobranch is eventually placed in the centre of the blastoderm: as a result of this abbreviation the embryo Bird occupies from the first a central position in the blastoderm[187].

The peculiar relations of the blastoderm and embryo, and the resulting relations of the neural and alimentary canal, appear to me to be features of quite as great an importance for classification as the presence or absence of an amnion and allantois.

General Features of the Hypoblast.

There are but few points to be noticed with reference to the histology of the hypoblast cells. The cells of the dorsal wall of the alimentary cavity are columnar and form a single row. Those derived from the yolk to form the ventral wall are at first roundish, but subsequently assume a more columnar form.

[Pg 309] The Notochord.

One of the most interesting features in the Elasmobranch development is the formation of the notochord from the hypoblast. All the steps in the process by which this takes place can be followed with great ease and certainty.

Up to stage B the hypoblast is in contact with the epiblast immediately below the medullary groove, but exhibits no trace of a thickening or any other formation at that point.

Between stage B and C the notochord first arises.

In the hindermost sections of this stage the hypoblast retains a perfectly normal structure and uniform thickness throughout. In the next few sections (Pl. 10, fig. 1c, ch´) a slight thickening is to be observed in the hypoblast, immediately below the medullary canal. The layer, which elsewhere is composed of a single row of cells, here becomes two cells deep, but no sign of a division into two layers exhibited.

In the next few sections the thickening of the hypoblast becomes much more pronounced; we have, in fact, a ridge projecting from the hypoblast towards the epiblast (Pl. 10, fig. 1b, ch´).

This ridge is pressed firmly against the epiblast, and causes in it a slight indentation. The hypoblast in the region of the ridge is formed of two layers of cells, the ridge being entirely due to the uppermost of the two.

In sections in front of this a cylindrical rod, which can at once be recognised as the notochord and is continuous with the ridge just described, begins to be split off from the hypoblast. It is difficult to say at what point the separation of this rod from the hypoblast is completed, since all intermediate gradations between complete separation and complete attachment are to be seen.

Where the separation first appears, a fairly thick bridge of hypoblast is left connecting the two lateral halves of the layer, but anteriorly this bridge becomes excessively delicate and thin (Pl. 10, fig. 1a), and in some cases is barely visible except with high powers.

From the series of sections represented, it is clear that the [Pg 310] notochord commences to be separated from the hypoblast anteriorly, and that the separation gradually extends backwards.

The posterior extremity of the notochord remains for a long time attached to the hypoblast; and it is not till the end of the period treated of in this chapter that it becomes completely free.

A sheath is formed around the notochord, very soon after its formation, at a stage intermediate between stages C and D. This sheath is very delicate, though it stains with both osmic acid and hæmatoxylin. I conclude from its subsequent history, that it is to be regarded as a product of the cells of the notochord, but at the same time it should be stated that it precisely resembles membrane-like structures, which I have already described as being probably artificial.

Towards the end of this period the cells of the notochord become very much flattened vertically, and cause the well-known stratified appearance which characterises the notochord in longitudinal sections. In transverse sections the outlines of the cells of the notochord appear rounded.

Throughout this period the notochord cells are filled with yolk-spherules, and near its close small vacuoles make their appearance in them.

An account of the development of the notochord, substantially similar to that I have just given, appeared in my preliminary paper[188] on the development of the Elasmobranch fishes.

To the remarks which were there made, I have little to add. There are two possible views, which can be held with reference to the development of the notochord from the hypoblast.

We may suppose that this is the primitive mode of development of the notochord, or we may suppose that the separation of the notochord from the hypoblast is due to a secondary process.

If the latter view is accepted, it will be necessary to maintain that the mesoblast becomes separated from the hypoblast as three separate masses, two lateral, and one median, and that the latter becomes separated much later than the two former.

We have, I think, no right to assume the truth of this view without further proof. The general admission of assumptions of this kind is apt to lead to an injurious form of speculation, in [Pg 311] which every fact presenting a difficulty in the way of some general theory is explained away by an arbitrary assumption, while all the facts in favour of it are taken for granted. It is however clear that no theory can ever be fairly tested so long as logic of this kind is permitted. If, in the present instance, the view is adopted that the notochord has in reality a mesoblastic origin, it will be possible to apply the same view to every other organ derived from the hypoblast, and to say that it is really mesoblastic, but has become separated at rather a late period from the hypoblast.

If, however, we provisionally reject this explanation, and accept the other alternative, that the notochord is derived from the hypoblast, we must be prepared to adopt one of two views with reference to the development of the notochord in other vertebrates. We must either suppose that the current statements as to the development of the notochord in other vertebrates are inaccurate, or that the notochord has only become secondarily mesoblastic.

The second of these alternatives is open to the same objections as the view that the notochord has only apparently a hypoblastic source in Elasmobranchii, and, provisionally at least, the first of them ought to be accepted. The reasons for accepting this alternative fall under two heads. In the first place, the existing accounts and figures of the development of the notochord exhibit in almost all cases a deficiency of clearness and precision. The exact stage necessary to complete the series never appears. It cannot, therefore, at present be said that the existing observations on the development of the notochord afford a strong presumption against its hypoblastic origin.

In the second place, the remarkable investigations of Hensen[189], on the development of the notochord in Mammalia, render it very probable that, in this group, the notochord is developed from the hypoblast.

Hensen finds that in Mammalia, as in Elasmobranchii, the mesoblast forms two independent lateral masses, one on each side of the medullary canal.

After the commencing formation of the protovertebræ the hypoblast becomes considerably thickened beneath the medullary [Pg 312] groove; and, though he has not followed out all the steps of the process by which this thickening is converted into the notochord, yet his observations go very far towards proving that it does become the notochord.

Against the observations of Hensen, there ought, however, to be mentioned those of Lieberkühn[190]. He believes that the two lateral masses of mesoblast, described by Hensen (in an earlier paper than the one quoted), are in reality united by a delicate layer of cells, and that the notochord is formed from a thickening of these.

Lieberkühn gives no further statements or figures, and it is clear that, even if there is present the delicate layer of mesoblast, which he fancies he has detected, yet this cannot in any way invalidate such a section as that represented on Pl. X. fig. 40, of Hensen's paper.

In this figure of Hensen's, the hypoblast cells become distinctly more columnar, and the whole layer much thicker immediately below the medullary canal than elsewhere, and this independently of any possible layer of mesoblast.

It appears to me reasonable to conclude that Lieberkühn's statements do not seriously weaken the certainty of Hensen's results.

In addition to the observations of Hensen's on Mammalia, those of Kowalevsky and Kuppfer on Ascidians may fairly be pointed to as favouring the hypoblastic origin of the notochord.

It is not too much to say that at the present moment the balance of evidence is in favour of regarding the notochord as a hypoblastic organ.

This conclusion is, no doubt, rather startling, and difficult to understand. The only feature of the notochord in its favour is the fact of its being unsegmented[191].

Should it eventually turn out that the notochord is developed in most vertebrates from the mesoblast, and only exceptionally from the hypoblast, the further question will have to be settled [Pg 313] as to whether it is primitively a hypoblastic or a mesoblastic organ; but, from whatever layer it has its source, an excellent example will be afforded of an organ changing from the layer in which it was originally developed into another distinct layer.

EXPLANATION OF PLATE 10.

Complete List of Reference Letters.

al. Alimentary canal. ch. Chorda dorsalis or notochord. ch´. Ridge of hypoblast, which will become separated off as the notochord. ep. Epiblast. hy. Hypoblast. lp. Coalesced lateral and vertebral plate of mesoblast. mg. Medullary groove. n. Nucleus of yolk. na. Cells formed around the nuclei of the yolk to enter into the ventral wall of the alimentary canal. nc. Neural or medullary canal. pv. Protovertebra. so. Somatopleure. sp. Splanchnopleure. ts. Mesoblast of tail-swelling. yk. Yolk-spherules.

Figs. 1a, 1b, 1c. Three sections from the same embryo belonging to a stage intermediate between B and C, of which fig. 1a is the most anterior. × 96 diameters.

The sections illustrate (1) The different characters of the medullary groove in the different regions of the embryo. (2) The structure of the coalesced lateral and vertebral plates. (3) The mode of formation of the notochord as a thickening of the hypoblast (ch´), which eventually becomes separated from the hypoblast as an elliptical rod (1a, ch).

Fig. 2. Section through the anterior part of an embryo belonging to stage C. The section is mainly intended to illustrate the formation of the ventral wall of the alimentary canal from cells formed around the nuclei of the yolk. It also shews the shallowness of the medullary groove in the anterior part of the body.

Figs. 2a, 2b, 2c. Three sections from the same embryo as fig. 2. Fig. 2a is the most anterior of the three sections and is taken through a point shortly in front of fig. 2. The figures illustrate the general features of an embryo of stage C, more especially the complete closing of the alimentary canal in front and the triangular section which it there presents.

Fig. 3. Section through the posterior part of an embryo belonging to stage D. × 86 diameters.

It shews the general features of the layers during the stage, more especially the differentiation of somatic and splanchnic layers of the mesoblast.

Figs. 3a, 3b, 3c, 3d, 3e, 3f. Sections of the same embryo as fig. 3 (× 60 diameters). Fig. 3 belongs to part of the embryo intermediate between figs. 3e and 3f.

The sections shew the features of various parts of the embryo. Figs. 3a, 3b and 3c belong to the head, and special attention should be paid to the presence of a cavity in the mesoblast in 3b and to the ventral curvature of the medullary folds.

Fig. 3d belongs to the neck, fig. 3e to the back, and fig. 3f to the tail.

Fig. 4. Section through the region of the tail at the commencement of stage F. × 60 diameters.

The section shews the character of the tail-swellings and the commencing closure of the medullary groove.

[Pg 314] Fig. 5. Transverse section through the anterior part of the head of an embryo belonging to stage F (× 60 diameters). It shews (1) the ventral curvature of the medullary folds next the head. (2) The absence of mesoblast in the anterior part of the head. hy points to the extreme front end of the alimentary canal.

Fig. 6. Section through the head of an embryo at a stage intermediate between F and G. × 86 diameters.

It shews the manner in which the medullary folds of the head unite to form the medullary canal.

Fig. 7. Longitudinal and vertical section through the tail of an embryo belonging to stage G.

It shews the direct communication which exists between the neural and alimentary canals.

The section is not quite parallel to the long axis of the embryo, so that the protovertebræ are cut through in its anterior part, and the neural canal passes out of the section anteriorly.

Fig. 8. Network of nuclei from the yolk of an embryo belonging to stage H.

[180] Vide Preliminary Account, etc. Q. Jl. Micros. Science, Oct. 1874, Pl. 14, 8a. [This Edition, No. V. Pl. 3, 8a.] This and the other section from the same embryo (stage F) may be referred to. I have not thought it worth while repeating them here.

[181] I underestimated the distinctness of this formation in my earlier paper, loc. cit., although I recognised the fact that the mesoblast cells became arranged in two distinct layers.

[182] Zeitschrift f. wiss. Zoologie, 1873.

[183] Nearly simultaneously with Chapter III. of the present monograph on the Development of Elasmobranchii, which dealt in a fairly complete manner with the genesis of cells outside the blastoderm, there appeared two important papers dealing with the same subject for Teleostei. One of these, by Professor Bambeke, Embryologie des Poissons Osseux, Mém. Cour. Acad. Belgique, 1875, which appeared some little time before my paper, and a second by Dr Klein, Quart. Jour. of Micr. Sci. April, 1876. In both of these papers a development of nuclei and of cells is described as occurring outside the blastoderm in a manner which accords fairly well with my own observations.

The conclusions of both these investigators differ however from my own. They regard the finely granular matter, in which the nuclei appear, as pertaining to the blastoderm, and morphologically quite distinct from the yolk. From their observations we can clearly recognise that the material in which the nuclei appear is far more sharply separated off from the yolk in Osseous Fish than in Elasmobranchii, and this sharp separation forms the main argument for the view of these authors. Dr Klein admits, however, that this granular matter (which he calls parablast) graduates into the typical food-yolk, though he explains this by supposing that the parablast takes up part of the yolk for the purpose of growth.

It is clear that the argument from a sharp separation of yolk and parablast cannot have much importance, when it is admitted (1) that in Osseous Fish there is a gradation between the two substances, while (2) in Elasmobranchii the one merges slowly and insensibly into the other.

The only other argument used by these authors is stated by Dr Klein in the following way. The fact that the parablast has, at the outset, been forming one unit with what represents the archiblast, and, while increasing has spread i.e. grown over the yolk which underlies the segmentation-cavity, is, I think, the most absolute proof that the yolk is as much different from the parablast as it is from the archiblast. This argument to me merely demonstrates that certain of the nutritive elements of the yolk become in the course of development converted into protoplasm, a phenomenon which must necessarily be supposed to take place on my own as well as on Dr Klein's view of the nature of the yolk. My own views on the subject have already been fully stated. I regard the so-called yolk as composed of a larger or smaller amount of food-material imbedded in protoplasm, and the meroblastic ovum as a body constituted of the same essential parts as a holoblastic ovum, though divided into regions which differ in the proportion of protoplasm they contain. I do not propose to repeat the positive arguments used by me in favour of this view, but content myself with alluding to the protoplasmic network found by Schultz and myself extending through the whole yolk, and to the similar network described by Bambeke as being present in the eggs of Osseous Fish after deposition but before impregnation. The existence of these networks is to me a conclusive proof of the correctness of my views. I admit that in Teleostei the 'parablast' contains more protoplasm than the homologous material in the Elasmobranch ovum, while it is probable that after impregnation the true yolk of Teleostei contains little or no protoplasm; but these facts do not appear to me to militate against my views.

I agree with Prof. Bambeke in regarding the cells derived from the sub-germinal matter as homologous with the so-called yolk-cells of the Amphibian embryo.

I have recently, in some of the later stages of development, met with very peculiar nuclei of the yolk immediately beneath the blastoderm at some little distance from the embryo, Pl. 10, fig. 8. They were situated not in finely sub-germinal matter, but amongst large yolk-spherules. They were very large, and presented still more peculiar forms than those already described by me, being produced into numerous long filiform processes. The processes from the various nuclei were sometimes united together, forming a regular network of nuclei quite unlike anything that I have previously seen described.

The sub-germinal matter, in which the nuclei are usually formed, becomes during the later stages of development far richer in protoplasm than during the earlier. It continually arises at fresh points, and often attains to considerable dimensions, no doubt by feeding on yolk-spherules. Its development appears to be determined by the necessities of growth in the blastoderm or embryo.

[184] Entwicklungsgeschichte der Unke.

[185] Mélanges Biologiques de l'Académie Pétersbourg, Tome VII.

[186] Archiv. f. mikros. Anat. Vol. VII. p. 114. In the passage on this point Kowalevsky states that in Elasmobranchii the neural and alimentary canals communicate. This I believe to be the first notice published of this peculiar arrangement.

[187] Vide Note on p. 281, also p. 295, and Pl. 9, figs. 1 and 2, and Comparison, &c., Qy. Jl. of Micros. Sci. July, 1875, p. 219. [This Edition, No. VI. p. 125.] These passages give an account of the change of position of the Elasmobranch embryo, and the Note on p. 281 contains a speculation about the nature of the primitive streak with its contained primitive groove. I have suggested that the primitive streak is probably to be regarded as a rudiment at the position where the edges of the blastoderm coalesced to give to the embryos of Birds and Mammals the central position which they occupy.

If my hypothesis should turn out to be correct, various, now unintelligible, features about the primitive streak would be explained: such as its position behind the embryo, the fusion of the epiblast and mesoblast in it, the groove it contains, &c.

The possibility of the primitive streak representing the blastopore, as it in fact does according to my hypothesis, ought also to throw light on E. Van Beneden's recent researches on the development of the Mammalian ovum.

In order clearly to understand the view here expressed, the reader ought to refer to the passages above quoted.

[188] Loc. cit.

[189] Zeitschrift f. Anat. u. Entwicklungsgeschichte, Vol. I. p. 366.

[190] Sitz. der Gesell. zu Marburg, Jan. 1876.

[191] In my earlier paper I suggested that the endostyle of Ascidians afforded an instance of a supporting organ being derived from the hypoblast. This parallel does not hold since the endostyle has been shewn to possess a secretory function. I never intended (as has been imagined by Professor Todaro) to regard the endostyle as the homologue of the notochord.

[Pg 315]

CHAPTER VI.

Development of the Trunk during Stages G to K.

By the stage when the external gills have become conspicuous objects, the rudiments of the greater number of the important organs of the body are definitely established.

Owing to this fact the first appearance of the external gills forms a very convenient break in the Elasmobranch development; and in the present chapter the history is carried on to the period of this occurrence.

While the last chapter dealt for the most part with the formation of the main organic systems from the three embryonic layers, the present one has for its subject the gradual differentiation of these systems into individual organs. In treating of the development of the separate organs a divergence from the plan of the last chapter becomes necessary, and the following arrangement has been substituted for it. First of all an account is given of the development of the external epiblast, which is followed by a description of the organs derived from the mesoblast and of the notochord.

External Epiblast.

During stages G to I the epiblast[192] is formed of a single layer of flattened cells; and in this, as in the earlier stages, it deserves to be especially noticed that the epiblast is never more than one cell deep, and is therefore incapable of presenting any differentiation into nervous and epidermic layers. (Pl. 11, figs. 1-5.)

[Pg 316] The cells which compose it are flattened and polygonal in outline, but more or less spindle-shaped in section. They present a strong contrast to the remaining embryonic cells of the body in possessing a considerable quantity of clear protoplasm, which in most other cells is almost entirely absent. Their granular nucleus is rounded or oval, and typically contains a single nucleolus. Frequently, however, two nucleoli are present, and when this is the case an area free from granules is to be seen around each nucleolus, and a dark line, which could probably be resolved into granules by the use of a sufficiently high magnifying power, divides the nucleus into two halves. These appearances probably indicate that nuclei, in which two nucleoli are present, are about to divide.

The epiblast cells vary in diameter from .022 to .026 Mm. and their nuclei from .014 to .018 Mm. They present a fairly uniform character over the greater part of the body. In Torpedo they present nearly the same characters as in Pristiurus and Scyllium, but are somewhat more columnar. (Pl. 11, fig. 7.)

Along the summit of the back from the end of the tail to the level of the anus, or slightly beyond this, epiblast cells form a fold—the rudiment of the embryonically undivided dorsal fin—and the cells forming this, unlike the general epiblast cells, are markedly columnar; they nevertheless, here as elsewhere, form but a single layer. (Pl. 11, fig. 3 and 5, df.) Although at this stage the dorsal fin is not continued as a fold anteriorly to the level of the anus, yet a columnar thickening or ridge of epiblast, extending along the median dorsal line nearly to the level of the heart, forms a true morphological prolongation of the fin.

On the ventral side of the tail is present a rudiment of the ventral unpaired fin, which stops short of the level of the anus, but, though less prominent, is otherwise quite similar to the dorsal fin and continuous with it round the end of the tail. At this stage the mesoblast has no share in forming either fin.

In many sections of the tail there may be seen on each side two folds of skin, which are very regular, and strongly simulate the rudimentary fins just described. The cells composing them are, however, not columnar, and the folds themselves are merely artificial products due to shrinking.

[Pg 317] At a stage slightly younger than K an important change takes place in the epiblast.

From being composed of a single layer of cells it becomes two cells deep. The two layers appear first of all anteriorly, and subsequently in the remaining parts of the body. At first, both layers are formed of flattened cells (Pl. 11, figs. 8, 9); but at a stage slightly subsequent to that dealt with in the present chapter, the cells of the inner of the two layers become columnar, and thus are established the two strata always present in the epidermis of adult vertebrates, viz. an outer layer of flattened cells and an inner one of columnar cells[193].

The history of the epiblast in Elasmobranchii is interesting, from the light which it throws upon the meaning of the nervous and epidermic layers into which the epiblast of Amphibians and some other Vertebrates is divided. The Amphibians and Elasmobranchii present the strongest contrast in the development of their epiblast, and it is worth while shortly to review and compare the history of the layer in the two groups.

In Amphibians the epiblast is from the first divided into an outer stratum formed of a single row of flattened cells, and an inner stratum composed of several rows of more rounded cells. These two strata were called by Stricker the nervous and epidermic layers, and these names have been very generally adopted.

Both strata have a share in forming the general epiblast, and though eventually they partially fuse together, there can be but little doubt that the horny layer of the adult epiblast, where such can be distinguished[194], is derived from the epidermic layer of the embryo, and the mucous layer of the epiblast from the embryonic nervous layer. Both layers of the epiblast assist in the formation of the cerebro-spinal nervous system, and there also at first fuse together[195], though the epidermic layer probably separates itself again, as the central epithelium of the spinal canal. The lens and auditory sac are derived exclusively from the nervous layer [Pg 318] of the epidermis, while this layer also has the greater share in forming the olfactory sac.

In Elasmobranchii the epiblast is at first uniformly composed of a single row of cells. The part of the layer which will form the central nervous system next becomes two or three cells deep, but presents no distinction into two layers; the remaining portions of the layer remain, as before, one cell deep. Although the epiblast at first presents this simple structure, it eventually, as we have seen, becomes divided throughout into two layers, homologous with the two layers which arise so early in Amphibians. The outer one of the two forms the horny layer of the epidermis and the central epithelium of the neural canal. The inner one, the mucous layer of the epidermis and the nervous part of the brain and spinal cord. Both layers apparently enter into the formation of the organs of sense.

While there is no great difficulty in determining the equivalent parts of the epidermis in Elasmobranchii and Amphibians, it still remains an open question in which of these groups the epiblast retains its primitive condition.

Though it is not easy to bring conclusive proofs on the one side or the other, the balance of argument appears to me to be decidedly in favour of regarding the condition of the epiblast in Elasmobranchii, and most other Vertebrates, as the primitive one, and its condition in Amphibians as a secondary one, due to the throwing back of the differentiation of their epiblast into two layers to a very early period in their development.

In favour of this view are the following points: (1) That a primitive division of the epiblast into two layers is unknown in the animal kingdom, except amongst Amphibians and (?) Osseous Fish. (2) That it appears more likely for a particular feature of development to be thrown back to an earlier period, than for such an important feature as a distinction between two primary layers to be absolutely lost during an early period of development, and then to reappear again in later stages.

The fact of the epiblast of the neural canal being divided, like the remainder of the layer, into nervous and epidermic parts, cannot, I think, be used as an argument in favour of the opposite view to that here maintained.

It seems probable that the central canal of the nervous [Pg 319] system arose as an involution from the exterior, and therefore that the epidermis lining it is in reality merely a part of the external epidermis, and as such is naturally separated from the true nervous structures adjacent to it[196].

Leaving the general features of the external skin, I pass to the special organs derived from it during the stage just anterior to K.

The unpaired Fins. The unpaired fins have grown considerably, and the epiblast composing them becomes, like the remainder of the layer, divided into two strata, both however composed of more or less columnar cells. The ventral fin has now become more prominent than the dorsal fin; but the latter extends forward as a fold quite to the anterior part of the body.

The paired Fins. Along each side of the body there appears during this stage a thickened line of epiblast, which from the first exhibits two special developments: one of these just in front of the anus, and a second and better marked one opposite the front end of the segmental duct. These two special thickenings are the rudiments of the paired fins, which thus arise as special developments of a continuous ridge on each side, precisely like the ridges of epiblast which form the rudiments of the unpaired fins.

Similar thickenings to those in Elasmobranchii are found at the ends of the limbs in the embryos of both Birds and Mammals, in the form of caps of columnar epiblast[197].

The ridge, of which the limbs are special developments, is situated on a level slightly ventral to that of the dorsal aorta, and extends from just behind the head to the level of the anus. It is not noticeable in surface views, but appears in sections as a portion of the epiblast where the cells are more columnar than elsewhere; precisely resembling in this respect the forward continuation of the dorsal fin. At the present stage the posterior thickenings of this ridge which form the abdominal fins are so slight as to be barely visible, and their real nature can only be detected by a careful comparison between sections of this and the succeeding stages. The rudiments of the anterior pair of [Pg 320] limbs are more visible than those of the posterior, though the passage between them and the remainder of the ridges is most gradual. Thus at first the rudiments of both the limbs are nothing more than slight thickenings of the epiblast, where its cells are more columnar than elsewhere. During stage K the rudiments of both pairs of limbs, but especially of the anterior pair, grow considerably, while at the same time the thickened ridge of epiblast which connects them together rapidly disappears. The thoracic limbs develop into an elongated projecting fold of epiblast, in every way like the folds forming the unpaired fins; while at the same time the cells of the subjacent mesoblast become closely packed, and form a slight projection, at the summit of which the fold of the epiblast is situated (Pl. 11, fig. 9). The maximum projection of the thoracic fin is slightly in advance of the front end of the segmental duct. The abdominal fins do not, during stage K, develop quite so fast as the thoracic, and at its close are merely elongated areas where the epiblast is much thickened, and below which the mesoblast is slightly condensed. In the succeeding stages they develop into projecting folds of skin, precisely as do the thoracic fins.

The features of the development of the limbs just described, are especially well shewn in Torpedo; in the embryos of which the passage from the general linear thickening of epiblast into the but slightly better marked thickening of the thoracic fin is very gradual, and the fact of the limb being nothing else than a special development of the linear lateral thickening is proved in a most conclusive manner.

If the account just given of the development of the limbs is an accurate record of what really takes place, it is not possible to deny that some light is thrown by it upon the first origin of the vertebrate limbs. The facts can only bear one interpretation, viz.: that the limbs are the remnants of continuous lateral fins.

The unpaired dorsal fin develops as a continuous thickening, which then grows up into a projecting fold of columnar cells. The greater part of this eventually atrophies, but three separate lobes are left which form the two dorsal fins and the upper lobe of the caudal fin.

The development of the limbs is almost identically similar [Pg 321] to that of the dorsal fins. There appears a lateral linear thickening of epiblast, which however does not, like the similar thickening of the fins, grow into a distinct fold. Its development becomes confined to two special points, at each of which is formed a continuous elongated fold of columnar cells precisely like the fold of skin forming the dorsal fins. These two folds form the paired fins. If it be taken into consideration that the continuous lateral fin, of which the rudiment appears in Elasmobranchii, does not exist in any adult Vertebrate, and also that a continuous dorsal fin exists in many Fishes, the small differences in development between the paired fins and the dorsal fins will be seen to be exactly those which might have been anticipated beforehand. Whereas the continuous dorsal fin, which often persists in adult fishes, attains a considerable development before vanishing, the originally continuous lateral one has only a very ephemeral existence.

While the facts of development strongly favour a view which would regard the limbs as remnants of a primitively continuous lateral fin, there is nothing in the structure of the limbs of adult Fishes which is opposed to this view. Externally they closely resemble the unpaired fins, and both their position and nervous supply appear clearly to indicate that they do not belong to one special segment of the body. They appear rather to be connected with a varying number of segments; a fact which would receive a simple explanation on the hypothesis here adopted[198].

My researches throw no light on the nature of the skeletal parts of the limb, but the suggestion which has been made by Günther[199] with reference to the limb of Ceratodus (the most primitive known), that it is a modification of a series of parallel rays, would very well suit the view here proposed.

Dr Dohrn[200] in speaking of the limbs, points out the difficulties [Pg 322] in the way of supposing that they can have originated de novo, and not by the modification of some pre-existing organ, and suggests that the limbs are modified gill-arches; a view similar to which has been hinted at by Professor Gegenbaur[201].

Dr Dohrn has not as yet given the grounds for his determination, so that any judgment on his views is premature.

None of my observations on Elasmobranchii lends any support to these views; but perhaps, while regarding the limbs as the remains of a continuous fin, it might be permissible to suppose that the pelvic and thoracic girdles are altered remnants of the skeletal parts of some of the gill-arches which have vanished in existing Vertebrates.

The absence of limbs in the Marsipobranchii and Amphioxus, for reasons already insisted upon by Dr Dohrn[202], cannot be used as an argument against limbs having existed in still more primitive Vertebrates.

Though it does not seem probable that a dorsal and ventral fin can have existed contemporaneously with lateral fins (at least not as continuous fins), yet, judging from such forms as the Rays, there is no reason why small balancing dorsal and caudal fins should not have co-existed with fully developed lateral fins.

Mesoblast. G-K.

The mesoblast in stage F forms two independent lateral plates, each with a splanchnic and somatic layer, and divided, as before explained, into a vertebral portion and a parietal portion. At their peripheral edge these plates are continuous with the general mesoblastic tissue of the non-embryonic part of the blastoderm; except in the free parts of the embryo, where they are necessarily separated from the mesoblast of the yolk-sac, and form completely independent lateral masses of cells.

During the stages G and H, the two layers of which the mesoblast is composed cease to be in contact, and leave between them a space which constitutes the commencement of the body-cavity (Pl. 10, fig. 1). From the very first this cavity is more or less clearly divided into two distinct parts; one of them [Pg 323] in the vertebral portion of the plates of mesoblast, the other in the parietal. The cavity in the parietal part of the plates alone becomes the true body-cavity. It extends uninterruptedly through the anterior parts of the embryo, but does not appear in the caudal region, being there indicated only by the presence of two layers in the mesoblast plates. Though fairly wide below, it narrows dorsally before becoming continuous with the cavity in the vertebral plates. The line of junction of the vertebral and parietal plates is a little ventral to the dorsal summit of the alimentary canal (Pl. 10, fig. 5). Owing to the fact that the vertebral plates are split up into a series of segments (protovertebræ), the section of the body-cavity they enclose is necessarily also divided into a series of segments, one for each protovertebra.

Thus the whole body-cavity consists of a continuous parietal space which communicates by a series of apertures with a number of separate cavities enclosed in the protovertebræ. The cavity in each of the protovertebræ is formed of a narrowed dorsal and a dilated ventral segment, the latter on the level of the dorsal aorta (Pl. 11, fig. 5). Cavities are present in all the vertebral plates with the exception of a few far back in the tail; and exist in part of the caudal region posterior to that in which a cavity in the parietal plate is present.

Protovertebræ. Each protovertebra[203] or vertebral segment of the mesoblast plate forms a flattened rectangular body, ventrally continuous with the parietal plate of mesoblast. During stage G the dorsal edge of the protovertebræ is throughout on about a level with the ventral third of the spinal cord. Each vertebral plate is composed of two layers, a somatic and a splanchnic, and encloses the already-mentioned section of the body-cavity. The cells of both layers of the plate are columnar, and each consists of a very large nucleus, invested by a delicate layer of protoplasm.

Before the end of stage H the inner or splanchnic wall of the protovertebra loses its simple constitution, owing to the middle part of it, opposite the dorsal two-thirds of the notochord, undergoing [Pg 324] peculiar changes. These changes are indicated in transverse sections (Pl. 11, figs. 5 and 6, mp´), by the cells in the part we are speaking of acquiring a peculiar angular appearance, and becoming one or two deep; and the meaning of the changes is at once shewn by longitudinal horizontal sections. These prove (Pl. 12, fig. 10) that the cells in this situation have become elongated in a longitudinal direction, and, in fact, form typical spindle-shaped embryonic muscle-cells, each with a large nucleus. Every muscle-cell extends for the whole length of a protovertebra, and in the present stage, or at any rate in stage I, acquires a peculiar granulation, which clearly foreshadows transverse striation (Pl. 12, figs. 11-13).

Thus by stage H a small portion of the splanchnopleure which forms the inner layer of each protovertebra, becomes differentiated into a distinct band of longitudinal striated muscles; these almost at once become functional, and produce the peculiar serpentine movements of the embryo, spoken of in a previous chapter, p. 291.

It may be well to say at once that these muscles form but a very small part of the muscles which eventually appear; which latter are developed at a very much later period from the remaining cells of the protovertebræ. The band developed at this stage appears to be a special formation, which has arisen through the action of natural selection, to enable the embryo to meet its respiratory requirements, by continually moving about, and so subjecting its body to fresh oxydizing influences; and as such affords an interesting example of an important structure acquired during and for embryonic life.

Though the cavities in the protovertebræ are at first perfectly continuous with the general body-cavity, of which indeed they merely form a specialized part, yet by the close of stage H they begin to be constricted off from the general body-cavity, and this process is continued rapidly, and completed shortly after stage I, and considerably before the commencement of stage K (Pl. 11, figs. 6 and 8). While this is taking place, part of the splanchnic layer of each protovertebra, immediately below the muscle-band just described, begins to proliferate, and produce a number of cells, which at once grow in between the muscles and the notochord. These cells are very easily [Pg 325] seen both in transverse and longitudinal sections, and form the commencing vertebral bodies (Pl. 11, fig. 6, and Pl. 12, figs. 10 and 11, Vr).

At first the vertebral bodies have the same segmentation as the protovertebræ from which they sprang; that is to say, they form masses of embryonic cells separated from each other by narrow slits, continuous with the slits separating the protovertebræ. They have therefore at their first appearance a segmentation completely different from that which they eventually acquire (Pl. 12, fig. 11).

After the separation of the vertebral bodies from the protovertebræ, the remaining parts of the protovertebræ may be called muscle-plates; since they become directly converted into the whole voluntary muscular system of the trunk. At the time when the cavity of the muscle-plates has become completely separate from the body-cavity, the muscle-plates themselves are oblong structures, with two walls enclosing the cavity just mentioned, in which the original ventral dilatation is still visible. The outer or somatic wall of the plates retains its previous simple constitution. The splanchnic wall has however a somewhat complicated structure. It is composed dorsally and ventrally of a columnar epithelium, but in its middle portion of the muscle-cells previously spoken of. Between these and the central cavity of the plates the epithelium forming the remainder of the layer commences to insert itself; so that between the first-formed muscle and the cavity of the muscle-plate there appears a thin layer of cells, not however continuous throughout.

At the end of the period K the muscle-plates have extended dorsally two-thirds of the way up the sides of the spinal cord, and ventrally to the level of the segmental duct. Their edges are not straight, but are bent into an angular form, with the apex pointing forwards. Vide Pl. 12, fig. 17, mp.

Before the end of the period a number of connective-tissue cells make their appearance, and extend upwards from the dorsal summit of the muscle-plates around the top of the spinal cord. These cells are at first rounded, but become typical branched connective-tissue cells before the close of the period (Pl. 11, figs. 7 and 8).

Between stages I and K the bodies of the vertebræ rapidly [Pg 326] increase in size and send prolongations downwards and inwards to meet below the notochord.

These soon become indistinguishably fused with other cells which appear in the area between the alimentary cavity and the notochord, but probably serve alone to form the vertebral bodies, while the cells adjoining them form the basis for the connective tissue of the kidneys, &c.

The vertebral bodies also send prolongations dorsalwards between the sides of the spinal cord and the muscle-plates. These grow round till they meet above the spinal and enclose the dorsal nerve-roots. They soon however become fused with the dorsal prolongations from the muscle-plates, at least so far as my methods of investigation enable me to determine; but it appears to me probable that they in reality remain distinct, and become converted into the neural arches, while the connective-tissue cells from the muscle-plates form the adjoining subcutaneous and inter-muscular connective tissue.

All the cells of the vertebral rudiments become stellate and form typical embryonic connective-tissue. The rudiments however still retain their primitive segmentation, corresponding with that of the muscle-plates, and do not during this period acquire their secondary segmentation. Their segmentation is however less clear than it was at an earlier period, and in the dorsal part of the vertebral rudiments is mainly indicated by the dorsal nerve-roots, which always pass out in the interval between two vertebral rudiments. Vide Pl. 12, fig. 12, pr.

Intermediate Cell-mass. At about the period when the muscle-plates become completely free, a fusion takes place between the somatopleure and splanchnopleure immediately above the dorsal extremity of the true body-cavity (Pl. 11, fig. 6). The cells in the immediate neighbourhood of this fusion form a special mass, which we may call the intermediate cell-mass—a name originally used by Waldeyer for the homologous cells in the Chick. Out of it are developed the urinogenital organs and the adjoining tissues. At first it forms little more than a columnar epithelium, but by the close of the period is divided into (1) An epithelium on the free surface; from this are derived the glandular parts of the kidneys and functional parts of the genital glands; and (2) a subjacent stroma which forms the [Pg 327] basis for the connective-tissue and vascular parts of these organs.

To the history of these parts a special section is devoted; and I now pass to the description of the mesoblast which lines the body-cavity and forms the connective tissue of the body-wall, and the muscular and connective tissue of the wall of the alimentary canal.

Body-cavity and Parietal Plates. By the close of stage H, as has been already mentioned, a cavity is formed between the somatopleure and splanchnopleure in the anterior part of the trunk, which rapidly widens during the succeeding stages. Anteriorly, it invests the heart, which arises during stage G, as a simple space between the ventral wall of the throat and the splanchnopleure (Pl. 11, fig. 4). Posteriorly it ends blindly.

This cavity forms in the region of the heart the rudiment of the pericardial cavity. The remainder of the cavity forms the true body-cavity.

Immediately behind the heart the alimentary canal is still open to the yolk-sac, and here naturally the two lateral halves of the body-cavity are separated from each other. In the tail of the embryo no body-cavity has appeared by stage I, although the parietal plates of mesoblast are distinctly divided into somatic and splanchnic layers. In the caudal region the lateral plates of mesoblast of the two sides do not unite ventrally, but are, on the contrary, quite disconnected. Their ventral edge is moreover much swollen (Pl. 11, fig. 1). At the caudal swelling the mesoblast plates cease to be distinctly divided into somatopleure and splanchnopleure, and more or less fuse with the hypoblast of the caudal vesicle (Pl. 11, fig. 2).

Between stages I and K the body-cavity extends backwards behind the point where the anus is about to appear, though it never reaches quite to the extreme end of the tail. The backward extension of the body-cavity, as is primitively the case everywhere, is formed of two independent lateral halves (Pl. 11, fig. 9a). Anteriorly, opposite the hind end of the small intestine, these two lateral halves unite ventrally to form a single cavity in which hangs the small intestine (Pl. 11, fig. 8) suspended by a very short mesentery.

[Pg 328] The most important change which takes place in the body-cavity during this period is the formation of a septum which separates off a pericardial cavity from the true body-cavity.

Immediately in front of the liver the splanchnic and somatic walls of the body come into very close contact, and I believe unite over the greater part of their extent. The septum so formed divides the original body-cavity into an anterior section or pericardial cavity, and a posterior section or true body-cavity. There is left, however, on each side dorsally a rather narrow passage which serves to unite the pericardial cavity in front with the true body-cavity behind.

In Pl. 11, fig. 8a, there is seen on one side a section through this passage, while on the other side the passage is seen to be connected with the pericardial cavity.

It is not possible from transverse sections to determine for certain whether the septum spoken of is complete. An examination of longitudinal horizontal sections from an embryo belonging to the close of the stage K has however satisfied me that this septum, by that stage at any rate, is fully formed.

The two lateral passages spoken of above probably unite in the adult to form the passage connecting the pericardial with the peritoneal cavity, which, though provided with but a single orifice into the pericardial cavity, divides into two limbs before opening into the peritoneal cavity.

The body-cavity undergoes no further changes of importance till the close of the period.

Somatopleure and Splanchnopleure. Both the somatic and splanchnic walls of the body-cavity during stage I exhibit a simple uniform character throughout their whole extent. They are formed of columnar cells where they line the dorsal part of the body-cavity, but ventrally of more rounded and irregular cells (Pl. 11, fig. 5).

In them may occasionally be seen aggregations of very peculiar and large cells with numerous highly refracting spherules; the cells forming these are not unlike the primitive ova to be described subsequently, but are probably large cells derived from the yolk.

It is during the stage intermediate between I and K that the first changes become visible which indicate a distinction between [Pg 329] an epithelium (endothelium) lining the body-cavity and the connective tissue adjoining this.

There are at first but very few connective-tissue cells between the epithelium of the somatic layer of the mesoblast and the epiblast, but a connection between them is established by peculiar protoplasmic processes which pass from the one to the other (Pl. 11, fig. 8). Towards the end of stage K, however, there appears between the two a network of mesoblastic cells connecting them together. In the rudimentary outgrowth to form the limbs the mesoblast cells of the somatic layer are crowded in an especially dense manner.

From the first the connective-tissue cells around the hypoblastic epithelium of the alimentary tract are fairly numerous (Pl. 11, fig. 8), and by the close of this period become concentrically arranged round the intestinal epithelium, though not divided into distinct layers. A special aggregation of them is present in the hollow of the rudimentary spiral valve.

Behind the anal region the two layers of the mesoblast (somatic and splanchnic) completely fuse during stage K, and form a mass of stellate cells in which no distinction into two layers can be detected (Pl. 11, figs. 9c, 9d).

The alimentary canal, which at first lies close below the aorta, becomes during this period gradually carried further and further from this, remaining however attached to the roof of the body-cavity by a thin layer of the mesoblast of the splanchnopleure formed of an epithelium on each side, and a few interposed connective-tissue cells. This is the mesentery, which by the close of stage K is of considerable length in the region of the stomach, though shorter elsewhere.

* * * * *

The above account of the protovertebræ and body-cavity applies solely to the genera Pristiurus and Scyllium. The changes of these parts in Torpedo only differ from those of Pristiurus in unimportant though fairly noticeable details. Without entering into any full description of these it may be pointed out that both the true body-cavity and its continuations into the protovertebræ appear later in Torpedo than in Pristiurus and Scyllium. In some cases even the muscle-plates become definitely separated and independent before the true body-cavity has appeared. As [Pg 330] a result of this the primitive continuity of the body-cavity and cavity of the muscle-plates becomes to a certain extent masked, though its presence may easily be detected by the obvious continuity which at first exists between the somatic and splanchnic layers of mesoblast and the two layers of the muscle-plate. In the muscle-plate itself the chief point to be noticed is the fact that the earlier formed bands of muscles (mp´) arise very much later, and are less conspicuous, in Torpedo than in the genera first described. They are however present and functional.

The anatomical relations of the body-cavity itself are precisely the same in Torpedo as in Pristiurus and Scyllium, and the pericardial cavity becomes separated from the peritoneal in the same way in all the genera; the two lateral canals connecting the two cavities being also present in all the three genera. The two independent parietal plates of mesoblast of the posterior parts of the body have ventrally a swollen edge, as in Pristiurus, and in this a cavity appears which forms a posterior continuation of the true body-cavity.

Resumé. The primitive independent mesoblast plates of the two sides of the body become divided into two layers, a somatic and a splanchnic (Hautfaserblatt and Darmfaserblatt). At the same time in the dorsal part of the mesoblast plate a series of transverse splits appear which mark out the limits of the protovertebræ and serve to distinguish a dorsal or vertebral part of the plate from a ventral or parietal part.

Between the somatic and splanchnic layers of the mesoblast plate a cavity arises which is continued quite to the summit of the vertebral part of the plate. This is the primitive body-cavity; and at first the cavity is divided into two lateral and independent halves.

The next change which takes place is the complete separation of the vertebral portion of the plate from the parietal; thereby the upper segmented part of the body-cavity becomes isolated and separated from the lower and unsegmented part. In connection with this change in the constitution of the body-cavity there are formed a series of rectangular plates, each composed of two layers, a somatic and a splanchnic, between which is the cavity originally continuous with the body-cavity. The splanchnic layer of the plates buds off cells to form the rudiments [Pg 331] of the vertebral bodies which are originally segmented in the same planes as the protovertebræ. The plates themselves remain as the muscle-plates and develop a special layer of muscle (mp´) in their splanchnic layer.

In the meantime the parietal plates of the two sides unite ventrally throughout the intestinal and cardiac regions of the body, and the two primitively isolated cavities contained in them coalesce. Posteriorly however the plates do not unite ventrally, and their contained cavities remain distinct.

At first the pericardial cavity is quite continuous with the body-cavity; but by the close of the period included in the present chapter it becomes separated from the body-cavity by a septum in front of the liver, which is however pierced dorsally by two narrow channels.

The parts derived from the two layers of the mesoblast (not including special organs or the vascular system) are as follow:—

From the somatic layer are formed

(1) A considerable part of the voluntary muscular system of the body.

(2) The dermis.

(3) A large part of the intermuscular connective tissue.

(4) Part of the peritoneal epithelium.

From the splanchnic layer are formed

(1) A great part of the voluntary muscular system.

(2) Part of the intermuscular connective tissue (?).

(3) The axial skeleton.

(4) The muscular and connective-tissue wall of the alimentary tract.

(5) A great part of the peritoneal epithelium.

General Considerations. In the history which has just been given of the development of the mesoblast, there are several points which appear to me to throw light upon the primitive origin of that layer. Before entering into these it is however necessary to institute a comparison between the history of the mesoblast in Elasmobranchii and in other Vertebrates, in order to distinguish as far as possible the primitive and the secondary characters present in the various groups.

[Pg 332] Though the Mammals are to be looked on as the most differentiated group amongst the Vertebrates, yet in their embryonic history they retain many very primitive features, and, as has been recently shewn by Hensen[204], present numerous remarkable approximations to the Elasmobranchii. We find accordingly[205] that the primitive lateral plates of mesoblast undergo nearly the same changes in these two groups. In Mammals there is at first a continuous cavity extending through both the parietal and vertebral portions of each plate, and dividing the plates into a somatic and a splanchnic layer: this cavity is the primitive body-cavity. The vertebral portion of each plate with its contained cavity then becomes divided off from the parietal. The later development of these parts is not accurately known, but it seems that the outer portion of each vertebral plate, composed of two layers (somatic and splanchnic) enclosing between them a remnant of the primitive body-cavity, becomes separated off as a muscle-plate. The remainder forms a vertebral rudiment, &c. Thus the extension of the body-cavity into the vertebral portion of the mesoblast, and the constriction of the vertebral portion of the cavity from the remainder, are as distinctive features of Mammals as they are of the Elasmobranchii.

In Birds[206] the horizontal splitting of the mesoblast into somatic and splanchnic layers appears, as in Mammals, to extend at first to the summit of the protovertebræ, but these bodies become so early separated from the parietal plates that this fact has usually been overlooked or denied; but even on the second day of incubation the outer layer of the protovertebræ is continuous with the somatic layer of the lateral plates, and the inner layer and kernel of the protovertebræ with the splanchnic layer of the lateral plates[207]. After the isolation of the protovertebræ the primitive position of the split which separated their somatic and splanchnic layers becomes obscured, but when [Pg 333] on the third day the muscle-plates are formed they are found to be constituted of two layers, an inner and an outer, which enclose between them a central cavity. This remarkable fact, which has not received much attention, though noticeable in most figures, receives a simple explanation as a surviving rudiment on Darwinian principles. The central cavity of the muscle-plate is, in fact, a remnant of the vertebral extension of the body-cavity, and is the same cavity as that found in the muscle-plates of Elasmobranchii. The two layers of the muscle-plate also correspond with the two layers present in Elasmobranchii, the one belonging to the somatic, the other to the splanchnic layer of mesoblast. The remainder of the protovertebræ internal to the muscle-plates is very large in Birds, and is the equivalent of that portion of the protovertebræ which in Elasmobranchii is split off to form the vertebral bodies[208] (Pl. 11, figs. 6, 7, 8, Vr). Thus, though the history of the development of the mesoblast is not precisely the same for Birds as for Elasmobranchii, yet the differences between the two groups are of such a character as to prove in a striking manner that the Avian development is a derivation from a more primary form, like that of the Elasmobranchii.

According to the statements of Bambeke and Götte, the Amphibians present rather remarkable peculiarities in the development of their muscular system. Each side-plate of mesoblast is divided into a somatic and a splanchnic layer, continuous throughout the vertebral and parietal portions of the plate. The vertebral portions (protovertebræ) of the plates soon become separated from the parietal, and form an independent mass of cells constituted of two layers, which were originally continuous with the somatic and splanchnic layers of the parietal plates. The outer or somatic layer of the vertebral plates is formed of a single row of cells, but the inner or splanchnic layer is made up of a central kernel of cells and an inner single layer. This central kernel is the first portion of the vertebral body to undergo [Pg 334] any change, and it becomes converted into the main dorso-lateral muscles of the body, which apparently correspond with the muscles derived from the whole muscle-plate of the Elasmobranchii. From the inner layer of the splanchnic division there are next formed the main internal ventral muscles, rectus abdominis, &c., as well as the chief connective-tissue elements of the parts surrounding the spinal cord. The outer layer of the vertebral plates forms the dermis and subcutaneous connective tissue, as well as some of the superficial muscles of the trunk and the muscles of the limbs.

Dr Götte appears to think that the vertebral plates in Amphibians present a perfectly normal development very similar to that of other Vertebrates. The divergences between Amphibians and other Vertebrates appear, however, to myself, to be very great, and although the very careful account given by Dr Götte is probably to be relied on, yet some further explanation than he has offered of the development of these parts amongst the Amphibians would seem to be required.

A primary stage in which the two layers of the vertebral plates are continuous with the somatic and splanchnic layers of the body-wall is equally characteristic of Amphibians, Elasmobranchii and Mammals. In the subsequent development, however, a great difference between the types becomes apparent, for whereas in Elasmobranchii both layers of the vertebral plates combine to form the muscle-plates, out of which the great dorso-lateral muscles are formed, in Amphibians what appear to be the equivalent muscles are derived from a few of the cells (the kernel) of the inner layer of the vertebral plates only. The cells which form the lateral muscles in Amphibians might be thought to correspond in position with the cells which become, in Elasmobranchii, converted into the special early formed band of muscles (m.p´.), rather than, as their development seems to indicate, with the whole Elasmobranch muscle-plates[209].

[Pg 335] Osseous Fishes are stated to agree with Amphibians in the development of their protovertebræ and muscular system[210], but further observations on this point are required.

Though the development of the general muscular system and muscle-plates does not, according to existing statements, take place on quite the same type throughout the Vertebrate subkingdom, yet the comparison which has been instituted between Elasmobranchii and other Vertebrates appears to prove that there are one or two common features in their development, which may be regarded as primitive, and as having been inherited from the ancestors of Vertebrates. These features are (1) The extension of the body-cavity into the vertebral plates, and subsequent enclosure of this cavity between the two layers of the muscle-plates; (2) The primitive division of the vertebral plate into a somatic and a splanchnic layer, and the formation of a large part of the voluntary muscular system out of the splanchnic layer.

* * * * *

The ultimate derivation of the mesoblast forms one of the numerous burning questions of modern embryology, and there are advocates to be found for almost every one of the possible views the question admits of.

All who accept the doctrine of descent are agreed that primitively only two embryonic layers were present—the epiblast and the hypoblast—and that the mesoblast subsequently appeared as a distinct layer, after a certain complexity of organization had been attained.

The general agreement stops, however, at this point, and the greatest divergence of opinion exists with reference to all further questions which bear on the development of the mesoblast. There appear to be four possibilities as to the origin of this layer.

It may be derived:

(1) entirely from the epiblast,

(2) partly from the epiblast, and partly from the hypoblast,

[Pg 336] (3) entirely from the hypoblast,

(4) or may have no fixed origin.

The fourth of these possibilities may for the present be dismissed, since it can be only maintained should it turn out that all the other views are erroneous. The first possibility is supported by the case of the Cœlenterata, and we might almost say by that of this group only[211].

Amongst the Cœlenterata the mesoblast, when present, is unquestionably a derivative of the epiblast, and when, as is frequently the case, a distinct mesoblast is not present, the muscle-cells form a specialized part of the epidermic cells.

The condition of the mesoblast in these lowly organized animals is exactly what might à priori have been anticipated, but the absence throughout the group of a true body-cavity, or specially developed muscular system of the alimentary tract, prevents the possibility of generalizing for other groups, from the condition of the mesoblast in this one.

In those animals in which a body-cavity and muscular alimentary tract are present, it would certainly appear reasonable to expect the mesoblast to be derived from both the primitive layers: the voluntary muscular system from epiblast, and the splanchnic system from the hypoblast. This view has been taken and strongly advocated by so distinguished an embryologist as Professor Haeckel, and it must be admitted, that on à priori grounds there is much to recommend it; there are, however, so far as I am aware of, comparatively few observed facts in its favour.

Professor Haeckel's own objective arguments in support of his view are as follows:

[Pg 337] (1) From the fact that some investigators derive the mesoblast with absolute confidence from the hypoblast, while others do so with equal confidence from the epiblast, he concludes that it is really derived from both these layers.

(2) A second argument is founded on the supposed derivation of the mesoblast in Amphioxus from both epiblast and hypoblast. Kowalevsky's account (on which apparently Prof. Haeckel's[212] statements are based) appears to me, however, too vague, and his observations too imperfect, for much confidence to be placed in his statements on this head. It does not indeed appear to me that the formation of the layers in Amphioxus, till better known, can be used as an argument for any special view about this question.

(3) Professor Haeckel's own observations on the development of Osseous fish form a third argument in support of his views. These observations do not, however, accord with those of the majority of investigators, and not having been made by means of sections, require further confirmation before they can be definitely accepted.

(4) A fourth argument rests on the fact that the various embryonic layers fuse together to form the primitive streak or axis-cord in higher vertebrates. This he thinks proves that the mesoblast is derived from both the primitive layers. The primitive streak has, however, according to my views, quite another significance to that attributed to it by Professor Haeckel[213]; but in any case Professor Kölliker's researches, and on this point my own observations accord with his, appear to me to prove that the fusion which there takes place is only capable of being used as an argument in favour of an epiblastic origin of the mesoblast, and not of its derivation from both epiblast and hypoblast.

The objective arguments in favour of Professor Haeckel's views are not very conclusive, and he himself does not deny that the mesoblast as a rule apparently arises as a single and undivided mass from one of the two primary layers, and only [Pg 338] subsequently becomes split into somatic and splanchnic strata. This original fusion and subsequent splitting of the mesoblast is explained by him as a secondary condition, a possibility which cannot by any means be thrown on one side. It seems therefore worth while examining how far the history of the somatic and splanchnic layers of the mesoblast in Elasmobranchii and other Vertebrates accords with the supposition that they were primitively split off from the epiblast and the hypoblast respectively.

It is well to consider first of all what parts of the mesoblast of the body might be expected to be derived from the somatic and splanchnic layers on this view of their origin[214].

From the somatic layer of the mesoblast there would no doubt be formed the whole of the voluntary muscular system of the body, the dermis, the subcutaneous connective tissue, and the connective tissue between the muscles. It is probable also, though this point is less certain, that the skeleton would be derived from the somatic layer. From the splanchnic layer would be formed the connective tissue and muscular layers of the alimentary tract, and possibly also the vascular system.

Turning to the actual development of these parts, the discrepancy between theory and fact becomes very remarkable. From the somatic layer of the mesoblast, part of the voluntary muscular system and the dermis is no doubt derived, but the splanchnic layer supplies the material, not only for the muscular wall of the digestive canal and the vascular system, but also for the whole of the axial skeleton and a great part of the voluntary muscular system of the body, including the first-formed muscles. Though remarkable, it is nevertheless not inconceivable, that the skeleton might be derived from the splanchnic mesoblast, but [Pg 339] it is very difficult to understand how there could be formed from it a part of the voluntary muscular system of the body indistinguishably fused with part of the muscular system derived from the somatopleure. No fact in my investigations comes out more clearly than that a great part of the voluntary muscular system is formed from the splanchnic layer of the mesoblast, yet this fact presents a most serious difficulty to the view that the somatic and splanchnic layers of the mesoblast in Vertebrates are respectively derived from the epiblast and hypoblast.

In spite, therefore, of general à priori considerations of a very convincing kind which tell in favour of the double origin of the mesoblast, this view is supported by so few objective facts, and there exists so powerful an array of facts against it, that at present, at least, it seems impossible to maintain it. The full strength of the facts against it will appear more fully in a review of the present state of our knowledge as to the development of the mesoblast in the different groups.

To this I now pass.

In a paper on the Early stages of Development in Vertebrates[215] a short resumé was given of the development of the mesoblast throughout the animal kingdom, which it may be worth while repeating here with a few additions. So far as we know at present, the mesoblast is derived from the hypoblast in the following groups:

Echinoderms (Hensen, Agassiz, Metschnikoff, Selenka, Götte), Nematodes (Bütschli), Sagitta (Kowalevsky, Bütschli), Lumbricus and probably other Annelids (Kowalevsky), Brachiopoda (Kowalevsky), Crustaceans (Bobretzky), Insects (Kowalevsky, Ulianin, Dohrn), Myriapods (Metschnikoff), Tunicates (Kowalevsky, Kuppfer), Petromyzon (Owsjanikoff), Osseous fishes (Oellacher, Götte, Kowalevsky), Elasmobranchii (Self), Amphibians (Remak, Stricker, Götte).

The list includes members from the greater number of the groups of the animal kingdom; the most striking omissions being the Cœlenterates, Mollusks, and the Amniotic Vertebrates. The absence of the Cœlenterates has been already explained and my grounds for regarding the Amniotic Vertebrates as [Pg 340] apparent rather than real exceptions have also been pointed out. The Mollusks, however, remain as a large group, in which we as yet know very little as to the formation of the mesoblast.

Dr Rabl[216], who seems recently to have studied the development of Lymnæus by means of sections, gives some figures shewing the origin of the mesoblast; they are, however, too diagrammatic to be of much service in settling the present question, and the memoirs of Professor Lankester[217] and Dr Fol[218] are equally inconclusive for this purpose, for, though they contain figures of elongated and branched mesoblast cells passing from the epiblast to the hypoblast, no satisfactory representations are given of the origin of these cells. I have myself observed in embryos of Turbo or Trochus similar elongated cells to those figured by Lankester and Fol, but was unable clearly to determine whence they arose. The most accurate observations which we have on this question are those of Professor Bobretzky[219]. In Nassa he finds that the three embryonic layers are all established during segmentation. The outermost and smallest cells form the epiblast, somewhat larger cells adjoining these the mesoblast, and the large yolk-cells the hypoblast. These observations do not, however, demonstrate from which of the primary layers the mesoblast is derived.

The evidence at present existing is clearly in favour of the mesoblast being, in almost all groups of animals, developed from the hypoblast, but strong as this evidence is, it has not its full weight unless the actual manner in which the mesoblast is in many groups derived from the hypoblast, is taken into consideration. The most important of these are the Echinoderms, Brachiopods and Sagitta.

In the Echinoderms the mesoblast is in part formed by cells budded off from the hypoblast, the remainder, however, arises as one or more diverticula of the alimentary tract. From the separate cells first budded off there are formed the cutis, part of the connective tissue and the calcareous skeleton[220]. The diverticula [Pg 341] from the alimentary cavity form the water-vascular system and the somatic and splanchnic layers of mesoblast. The cavity of the diverticula after the separation of the water-vascular system, forms the body-cavity. The outer lining layer of the cavity forms the somatic layer of mesoblast and the voluntary muscles; the inner lining layer the splanchnic mesoblast which unites with the epithelium of the alimentary tract. Though this fundamental arrangement would seem to be universal amongst Echinoderms, considerable variations of it are exhibited in different groups.

There is one outgrowth from the alimentary tract in Synapta; two in Echinoids, Asteroids and Ophiura; three in Comatula, and four (?) in Amphiura. The cavity of the outgrowth usually forms the body-cavity, but sometimes in Ophiura and Amphiura (Metschnikoff) the outgrowths are from the first or soon become solid, and only secondarily acquire a cavity, which is however homologous with the body-cavity of the other groups.

In Sagitta[221] the formation of the mesoblast and the alimentary tract takes place in nearly the same fashion as in the Echinoderms. The simple invaginate alimentary cavity becomes divided into three lobes, a central and two lateral. The two lateral lobes are gradually more and more constricted off from the central one, and become eventually quite separated from it; their cavities remain independent, and form in the adult the body-cavity, divided by a mesentery into two distinct lateral sections. The inner layer of each of the two lateral lobes forms the mesoblast of the splanchnopleure, the outer layer the mesoblast of the somatopleure. The central division of the primitive gastræa cavity remains as the alimentary tract of the adult.

The remarkable observations of Kowalevsky[222] on the development of the Brachiopoda have brought to light the unexpected fact that in two genera at least (Argiope and Terebratula) the mesoblast and body-cavity develop as paired constrictions from [Pg 342] the alimentary tract in a manner almost identically the same as in Sagitta.

It thus appears that, so far as can be determined from the facts at our disposal, the mesoblast in almost all cases is derived from the hypoblast, and in three widely separated groups it arises as a pair of diverticula from the alimentary tract, each diverticulum containing a cavity which eventually becomes the body-cavity. I have elsewhere suggested[223] that the origin of the mesoblast from alimentary diverticula is to be regarded as primitive for all higher animals, and that the more general cases in which the mesoblast becomes split off, as an undivided layer, from the hypoblast, are in reality derivates from this. The chief obstacle in the way of this view arises from the difficulty of understanding how the whole voluntary muscular system can have been derived at first from the alimentary tract. That part of a voluntary system of muscles might be derived from the contractile diverticula of the alimentary canal attached to the body-wall is not difficult to understand, but it is not easy to believe that the secondary system so formed could completely replace the primitive muscular system, derived, as it must have been, from the epiblast. In my paper above quoted will be found various speculative suggestions for removing this difficulty, which I do not repeat here. If it be granted, however, that in Sagitta, Brachiopods, and Echinoderms we have genuine examples of the formation of the whole mesoblast from alimentary diverticula, it is easy to see how the formation of the mesoblast in Vertebrates may be a secondary derivate from an origin of this nature.

An attempt has been already made to shew that the mesoblast in Elasmobranchii is formed in a very primitive fashion, and for this reason the Elasmobranchii appear to be especially adapted for determining whether any signs are exhibited of a derivation of the mesoblast as paired diverticula of the alimentary tract. There are, it appears to me, several such features. In the first place, the mesoblast is split off from the hypoblast not as a single mass but as a pair of distinct masses, comparable with the paired diverticula already alluded to. [Pg 343] Secondly, the body-cavity when it appears in the mesoblast plates, does not arise as a single cavity, but as a pair of cavities, one for each plate of mesoblast, and these cavities remain permanently distinct in some parts of the body, and nowhere unite till a comparatively late period. Thirdly, the primitive body-cavity of the embryo is not confined to the region in which a body-cavity exists in the adult, but extends to the summit of the muscle-plates, at first separating parts which become completely fused in the adult to form the great lateral muscles of the body. It is difficult to understand how the body-cavity could have such an extension as this, on the supposition that it represents a primitive split in the mesoblast between the wall of the gut and the body-wall; but its extension to this part is quite intelligible, on the supposition that it represents the cavities of two diverticula of the alimentary tract, from whose muscular walls the voluntary muscular system has been derived. Lastly, I would point out that the derivation of part of the muscular system from what appears as the splanchnopleure is quite intelligible on the assumed hypothesis, but, as far as I see, on no other.

Such are the main features presented by the mesoblast in Elasmobranchii, which favour the view of its having originally formed the walls of the alimentary diverticula. Against this view of its nature are the facts (1) of the mesoblast plates being at first solid, and (2), as a consequence of this, of the body-cavity never communicating with the alimentary canal. These points, in view of our knowledge of embryological modifications, cannot be regarded as great difficulties to my view. We have many examples of organs, which, though in most cases arising as involutions, yet appear in other cases as solid ingrowths. Such examples are afforded by the optic vesicle, auditory vesicle, and probably also by the central nervous system, of Osseous Fish. In most Vertebrates these organs are formed as hollow involutions from the exterior; in Osseous Fish, however, as solid involutions, in which a cavity secondarily appears.

The segmental duct of Elasmobranchii or the Wolffian duct (segmental duct) of Birds are cases of a similar kind, being organs which must originally have been formed as hollow involutions, but which now arise as solid bodies.

[Pg 344] Only one more instance of this kind need be cited, taken from the Echinoderms.

The body-cavity and the mesoblast investing it arise in the case of most Echinoderms as hollow involutions of the alimentary tract, but in some exceptional groups, Ophiura and Amphiura, are stated to be solid at first and only subsequently to become hollow. Should the accuracy of Metschnikoff's account of this point be confirmed, an almost exact parallel to what has been supposed by me to have occurred with the mesoblast in Elasmobranchii, and other groups, will be supplied.

The tendency of our present knowledge appears to be in favour of regarding the body-cavity in Vertebrates as having been primitively the cavity of alimentary diverticula, and the mesoblast as having formed the walls of the diverticula.

This view, to say the least of it, suits the facts which we know far better than any other theory which has been proposed, and though no doubt the à priori difficulties in its way are very great, yet it appears to me to be sufficiently strongly supported to deserve the attention of investigators. In the meantime, however, our knowledge of invertebrate embryology is so new and imperfect that no certainty on a question like that which has just been discussed can be obtained; and any generalizations made at present are not unlikely to be upset by the discovery of fresh facts.

The only other point in connection with the mesoblast which I would call attention to is the formation of the vertebral bodies.

My observations confirm those of Remak and Gegenbaur, shewing that there is a primary segmentation of the vertebral bodies corresponding to that of the muscle-plates, followed by a secondary segmentation in which the central lines of the vertebral bodies are opposite the partitions between the muscle-plates.

The explanation of these changes is not difficult to find. The primary segmentation of the body is that of the muscle-plates, which must have been present at a time when the vertebral bodies had no existence. As soon however as the notochordal sheath was required to be strong as well as flexible, it necessarily became divided into a series of segments.

The conditions under which the lateral muscles can cause the [Pg 345] flexure of the vertebral column are clearly that each muscle-segment shall be capable of acting on two vertebræ; and this condition can only be fulfilled when the muscle-segments are opposite the intervals between the vertebræ. Owing to this necessity, when the vertebral segments became formed, their centres corresponded, not with the centres of the muscle-plates, but with the inter-muscular septa.

These considerations fully explain the secondary segmentation of the vertebræ by which they become opposite the inter-muscular septa. On the other hand, the primary segmentation is clearly a remnant of the time when no vertebral bodies were present, and has no greater morphological significance than the fact that the cells to form the unsegmented investment of the notochord were derived from the segmented muscle-plates, and only secondarily became fused into a continuous tube.

The Urinogenital System.

The first traces of the urinary system become visible at about the time of the appearance of the third visceral cleft. At about this period the somatopleure and splanchnopleure become more or less fused together at the level of the dorsal aorta, and thus, as has been already mentioned, each of the original plates of mesoblast becomes divided into a vertebral plate and lateral plate (Pl. 11, fig. 6). The mass of cells resulting from this fusion corresponds with Waldeyer's intermediate cell-mass in the Fowl.

At about the level of the fifth protovertebra the first trace of the urinary system appears.

From the intermediate cell-mass a solid knob grows outwards towards the epiblast (woodcut, fig. 4, pd). This knob consists at first of 20-30 cells, which agree in character with the neighbouring cells of the intermediate cell-mass, and are at this period rounded. It is mainly, if not entirely, derived from the somatic layer of the mesoblast.

From this knob there grows backwards a solid rod of cells which keeps in very close contact with the epiblast, and rapidly diminishes in size towards its posterior extremity. Its hindermost part consists in section of at most one or two cells. It keeps so close to the epiblast that it might be supposed to be [Pg 346] derived from that layer were it not for the sections shewing its origin from the knob above mentioned. We have in this rod the commencement of what I have elsewhere[224] called the segmental duct.

Fig. 4. Two sections of a Pristiurus Embryo with three visceral clefts.

The sections are to shew the development of the segmental duct (pd) or primitive duct of the kidneys. In A (the anterior of the two sections) this appears as a solid knob projecting towards the epiblast. In B is seen a section of the column which has grown backwards from the knob in A.

spn. rudiment of a spinal nerve; mc. medullary canal; ch. notochord; X. string of cells below the notochord; mp. muscle-plate; mp´. specially developed portion of muscle-plate; ao. dorsal aorta; pd. segmental duct; so. somatopleura; sp. splanchnopleura; pp. pleuro-peritoneal or body-cavity; ep. epiblast; al. alimentary canal.

My observations shew that the segmental duct is developed in the way just described in both Pristiurus and Torpedo. Its origin in Pristiurus is shewn in the adjoining woodcut, and in Torpedo in Pl. 11, fig. 7, sd.

At a stage somewhat older than I, the condition of the segmental duct has not very materially altered. It has increased considerably in length, and the knob at its front end is both absolutely smaller, and also consists of fewer cells than before (Pl. 11, fig. 7, sd). These cells have become more columnar, and have begun to arrange themselves radially; thus indicating the early appearance of the lumen of the duct. The cells forming the front part of the rod, as well as those of the knob, commence to exhibit a columnar character, but in the hinder part of the [Pg 347] rod the cells are still rounded. In no part of it has a lumen appeared.

At this period also the knob, partly owing to the commencing separation of the muscle-plate from the remainder of the mesoblast, begins to pass inwards and approach the pleuro-peritoneal cavity.

At the same stage the first not very distinct traces of the remainder of the urinary system become developed. These appear in the form of solid outgrowths from the intermediate cell-mass just at the most dorsal part of the body-cavity.

The outgrowths correspond in numbers with the vertebral segments, and are at first quite disconnected with the segmental duct. At this stage they are only distinctly visible in the first few segments behind the front end of the segmental duct. A full description of them will come more conveniently in the next stage.

By a stage somewhat earlier than K important changes have taken place in the urinary system.

The segmental duct has acquired a lumen in its anterior portion, which opens at its front end into the body-cavity. (Pl. 11, fig. 9, sd.) The lumen is formed by the columnar cells spoken of in the last stage, acquiring a radiating arrangement round a central point, at which a small hole appears. After the lumen has once become formed, it rapidly increases in size.

The duct has also grown considerably in length, but its hind extremity is still as thin, and lies as close to the epiblast, as at first. The segmental involutions which commenced to be formed in the last stage, have now appeared for every vertebral segment along the whole length of the segmental duct, and even for two or three segments behind this.

They are simple independent outgrowths arising from the outer and uppermost angle of the body-cavity, and are at first almost without a trace of a lumen; though their cells are arranged as two layers. They grow in such a way as to encircle the oviduct on its inner and upper side (Pl. 11, fig. 8 and Pl. 12, fig. 14b, st). When the hindermost ones are formed, a slight trace of a lumen is perhaps visible in the front ones. At a stage slightly subsequent to this, in Scyllium canicula, I noticed 29 [Pg 348] of them; the first of them arising in the segment immediately behind the front end of the oviduct (Pl. 12, fig. 17, st), and two of them being formed in segments just posterior to the hinder extremity of the oviduct.

Pl. 12, figs. 16 and 18 represent two longitudinal sections shewing the segmental nature of the involutions and their relation to the segmental duct.

Many of the points which have been mentioned can be seen by referring to Pl. 11 and 12. Anteriorly the segmental duct opens into the pleuro-peritoneal cavity. In the sections behind this there may be seen the segmental duct with a distinct lumen, and also a pair of segmental involutions (Pl. 12, fig. 14a). In the still posterior sections the segmental duct would be quite without a lumen, and would closely adjoin the epiblast.

It seems not out of place to point out that the modes of the development of the segmental duct and of the segmental involutions are strikingly similar. Both arise as solid involutions, from homologous parts of the mesoblast. The segmental duct arises in the vertebral segment immediately in front of that in which the first segmental involution appears; so that the segmental duct appears to be equivalent to a single segmental involution.

The next stage corresponds with the first appearance of the external gills. The segmental duct now communicates by a wide opening with the body-cavity (Pl. 11, fig. 9, sd). It possesses a lumen along its whole length up to the extreme hind end (Pl. 11, fig. 9a). It is, however, at this hinder extremity that the most important change has taken place. This end has grown downwards towards that part of the alimentary canal which still lies behind the anus. This downgrowth is beginning to shew distinct traces of a lumen, and will appear in the next stage as one of the horns by which the segmental ducts communicate with the cloaca (Pl. 11, fig. 9b). All the anterior segmental involutions have now acquired a lumen. But this is still absent in the posterior ones (Pl. 11, fig. 9a).

Owing to the disappearance of the body-cavity in the region behind the anus, the primitive involutions there remain as simple masses of cells still disconnected with the segmental duct (Pl. 11, figs. 9b, 9c and 9d).

[Pg 349] Primitive Ova. The true generative products make their first appearance as the primitive ova between stages I and K.

In the sections of one of my embryos of this stage they are especially well shewn, and the following description is taken from those displayed in that embryo.

They are confined to the region which extends posteriorly nearly to the end of the small intestine and anteriorly to the abdominal opening of the segmental duct.

Their situation in this region is peculiar. There is no trace of a distinct genital ridge, but the ova mainly lie in the dorsal portion of the mesentery, and therefore in a part of the mesoblast which distinctly belongs to the splanchnopleure (Pl. 12, fig. 14a). Some are situated external to the segmental involutions; and others again, though this is not common, in a part of the mesoblast which distinctly belongs to the body-wall (Pl. 12, fig. 14b).

The portion of mesentery, in which the primitive ova are most densely aggregated, corresponds to the future position of the genital ridge, but the other positions occupied by ova are quite outside this. Some ova are in fact situated on the outside of the segmental duct and segmented tubes, and must therefore effect a considerable migration before reaching their final positions in the genital ridge on the inner side of the segmental duct (Pl. 12, fig. 14b).

The condition of the tissue in which the ova appear may at once be gathered from an examination of the figures given. It consists of an irregular epithelium of cells partly belonging to the somatopleure and partly to the splanchnopleure, but passing uninterruptedly from one layer to the other. The cells which compose it are irregular in shape, but frequently columnar (Pl. 12, figs. 14a and 14b).

They are formed of a nucleus which stains deeply, invested by a very delicate layer of protoplasm. At the junction of somatopleure and splanchnopleure they are more rounded than elsewhere. Very few loose connective-tissue cells are present. The cells just described vary from .008 Mm. to .01 Mm. in diameter.

The primitive ova are situated amongst them and stand out with extraordinary clearness, to which justice is hardly done in my figures.

[Pg 350] The normal full-sized ova exhibit the following structure. They consist of a mass of somewhat granular protoplasm of irregular, but more or less rounded, form. Their size varies from .016 - .036 Mm. In their interior a nucleus is present, which varies from .012 - .016 Mm., but its size as a rule bears no relation to the size of the containing cell.

This is illustrated by the subjoined list of measurements.

The numbers given refer to degrees on my micrometer scale.

Since it is the ratio alone which it is necessary to call attention to, the numbers are not reduced to decimals of a millimeter. Each degree of my scale is equal, however, with the object glass employed, to .002 Mm.

This series brings out the result I have just mentioned with great clearness.

In one case we find a cell has three times the diameter of the nucleus 16 : 5½; in another case 10 : 8, the nucleus has only a slightly smaller diameter than the cell. The irrationality of the ratio is fairly shewn in some of my figures, though none of the largest cells with very small nuclei have been represented.

The nuclei are granular, and stain fairly well with hæmatoxylin. They usually contain a single deeply stained nucleolus, but in many cases, especially where large (and this independently [Pg 351] of the size of the cell), they contain two nucleoli (Pl. 12, figs. 14c and 14d), and are at times so lobed as to give an apparent indication of commencing division.

A multi-nucleolar condition of the nuclei, like that figured by Götte[225], does not appear till near the close of embryonic life, and is then found equally in the large ova and in those not larger than the ova which exist at this early date.

As regards the relation of the primitive ova to each other and the neighbouring cells, there are a few points which deserve attention. In the first place, the ova are, as a rule, collected in masses at particular points, and not distributed uniformly (fig. 14a). The masses in some cases appear as if they had resulted from the division of one primitive ovum, but can hardly be adduced as instances of a commencing coalescence; since if the ova thus aggregated were to coalesce, an ovum would be produced of a very much greater size than any which is found during the early stages. Though at this stage no indication is present of such a coalescence of cells to form ova as is believed to take place by Götte, still the origin of the primitive ova is not quite clear. One would naturally expect to find a great number of cells intermediate between primitive ova and ordinary columnar cells. Cells which may be intermediate are no doubt found, but not nearly so frequently as might have been anticipated. One or two cells are shewn in Pl. 12, fig. 14a, x, which are perhaps of an intermediate character; but in most sections it is not possible to satisfy oneself that any such intermediate cells are present.

In one case what appeared to be an intermediate cell was measured, and presented a diameter of .012 Mm. while its nucleus was .008 Mm. Apart from certain features of the nucleus, which at this stage are hardly very marked, the easiest method of distinguishing a primitive ovum from an adjacent cell is the presence of a large quantity of protoplasm around the nucleus. The nucleus of one of the smallest primitive ova is not larger than the nucleus of an ordinary cell (being about .008 Mm. in both). It is perhaps the similarity in the size of the nuclei which renders it difficult at first to distinguish developing primitive ova from ordinary cells. Except with the [Pg 352] very thinnest sections a small extra quantity of protoplasm around a nucleus might easily escape detection, and the developing cell might only become visible when it had attained to the size of a small typical primitive ovum.

It deserves to be noticed that the nuclei even of some of the largest primitive ova scarcely exceed the surrounding nuclei in size. This appears to me to be an argument of some weight in shewing that the great size of primitive ova is not due to the fact of their having been formed by a coalescence of different cells (in which case the nucleus would have increased in the same proportion as the cell); but to an increase by a normal method of growth in the protoplasm around the nucleus.

It appears to me to be a point of great importance certainly to determine whether the primitive ova arise by a metamorphosis of adjoining cells, or may not be introduced from elsewhere. In some of the lower animals, e.g. Hydrozoa, there is no question that the ova are derived from the epiblast; we might therefore expect to find that they had the same origin in Vertebrates. Further than this, ova are frequently capable in a young state of executing amœboid movements, and accordingly of migrating from one layer to another. In the Elasmobranchii the primitive ova exhibit in a hardened state an irregular form which might appear to indicate that they possess a power of altering their shape, a view which is further supported by some of them being at the present stage situated in a position very different from that which they eventually occupy, and which they can only reach by migration. If it could be shewn that there were no intermediate stages between the primitive ova and the adjoining cells (their migratory powers being admitted) a strong presumption would be offered in favour of their having migrated from elsewhere to their present position. In view of this possibility I have made some special investigations, which have however led to no very satisfactory results. There are to be seen in the stages immediately preceding the present one, numerous cells in a corresponding position to that of the primitive ova, which might very well be intermediate between the primitive ova and ordinary cells, but which offer no sufficiently well marked features for a certain determination of their true nature.

[Pg 353] In the particular embryo whose primitive ova have been described these bodies were more conspicuous than in the majority of cases, but at the same time they presented no special or peculiar characters.

In a somewhat older embryo of Scyllium the cells amongst which the primitive ova lay had become very distinctly differentiated as an epithelium (the germinal epithelium of Waldeyer) well separated by what might almost be called a basement membrane from the adjoining connective-tissue cells. Hardly any indication of a germinal ridge had appeared, but the ova were more definitely confined than in previous embryos to the restricted area which eventually forms this. The ova on the average were somewhat smaller than in the previous cases.

In several embryos intermediate in age between the embryo whose primitive ova were described at the commencement of this section and the embryo last described, the primitive ova presented some peculiarities, about the meaning of which I am not quite clear, but which may perhaps throw some light on the origin of these bodies.

Instead of the protoplasm around the nucleus being clear or slightly granular, as in the cases just described, it was filled in the most typical instances with numerous highly refracting bodies resembling yolk-spherules. In osmic acid specimens (Pl. 12, fig. 15) these stain very darkly, and it is then as a rule very difficult to see the nucleus; in specimens hardened in picric acid and stained with hæmatoxylin these bodies are stained of a deep purple colour, but the nucleus can in most cases be distinctly seen. In addition to the instances in which the protoplasm of the ova is quite filled with these bodies, there are others in which they only occupy a small area adjoining the nucleus (Pl. 12, fig. 15a), and finally some in which only one or two of these bodies are present. The protoplasm of the primitive ova appears in fact to present a series of gradations between a state in which it is completely filled with highly refracting spherules and one in which these are completely absent.

This state of things naturally leads to the view that the primitive ova, when they are first formed, are filled with these spherules, which are probably yolk-spherules, but that they [Pg 354] gradually lose them in the course of development. Against this interpretation is the fact that the primitive ova in the younger embryo first described are completely without these bodies; this embryo however unquestionably presented an abnormally early development of the ova; and I am satisfied that embryos present considerable variations in this respect.

If the primitive ova are in reality in the first instance filled with yolk-spherules, the question arises as to whether, considering that they are the only mesoblast cells filled at this period with yolk-spherules, we must not suppose that they have migrated from some peripheral part of the blastoderm into their present position. To this question I can give no satisfactory answer. Against a view which would regard the spherules in the protoplasm as bodies which appear subsequently to the first formation of the ova, is the fact that hitherto no instances in which these spherules were present have been met with in the late stages of development; and they seem therefore to be confined to the first stages.

Notochord.

The changes undergone by the notochord during this period present considerable differences according to the genus examined. One type of development is characteristic of Scyllium and Pristiurus; a second type, of Torpedo.

My observations being far more complete for Scyllium and Pristiurus than for Torpedo, it is to the two former genera only that the following account applies, unless the contrary is expressly stated. Only the development of the parts of the notochord in the trunk are here dealt with; the cephalic section of the notochord is treated of in a subsequent section.

During stage G the notochord is composed of flattened cells arranged vertically, rendering the histological characters of the notochord difficult to determine in transverse sections. In longitudinal sections, however, the form and arrangement of the cells can be recognised with great ease. At the beginning of stage G each cell is composed of a nucleus invested by granular protoplasm frequently vacuolated and containing in suspension numerous yolk-spherules. It is difficult to determine whether [Pg 355] there is only one vacuole for each cell, or whether in some cases there may not be more than one.

Round the exterior of the notochord there is present a distinct though delicate cuticular sheath.

The vacuoles are at first small, but during stage G rapidly increase in size, while at the same time the yolk-spherules completely vanish from the notochord.

As a result of the rapid growth of the vacuoles, the nuclei, surrounded in each case by a small amount of protoplasm, become pushed to the centre of the notochord, the remainder of the protoplasm being carried to the edge. The notochord thus becomes composed during stages H and I (Pl. 11, fig. 4-6) of a central area mainly formed of nuclei with a small quantity of protoplasm around them, and of a thin peripheral layer of protoplasm without nuclei, the widish space between the two being filled with clear fluid. The exterior of the cells is indurated, so that they may be said to be invested by a membrane[226]; the cells themselves have a flattened form, and each extends from the edge to the centre of the notochord, the long axis of each being rather greater than half the diameter of the cord.

The nuclei of the notochord are elliptical vesicles, consisting of a membrane filled with granular contents, amongst which is situated a distinct nucleolus. They stain deeply with hæmatoxylin. Their long diameter in Scyllium is about 0.02 Mm.

The diameter of the whole notochord in Pristiurus during stage I is about 0.1 Mm. in the region of the back, and about 0.08 Mm. near the posterior end of the body.

Owing to the form of its constituent cells, the notochord presents in transverse sections a dark central area surrounded by a lighter peripheral one, but its true structure cannot be unravelled without the assistance of longitudinal sections. In these (Pl. 12, fig. 10) the nuclei form an irregular double row in the centre of the cord. Their outlines are very clear, but those of the individual cells cannot for certain be made out. It is, however, easy to see that the cells have a flattened and wedge-shaped form, with the narrow ends overlapping and interlocking at the centre of the notochord.

[Pg 356] By the close of stage I the cuticular sheath of the notochord has greatly increased in thickness.

During the period intermediate between stages I and K the notochord undergoes considerable transformations. Its cells cease to be flattened, and become irregularly polygonal, and appear but slightly more compressed in longitudinal sections than in transverse ones. The vacuolation of the cells proceeds rapidly, and there is left in each cell only a very thin layer of protoplasm around the nucleus. Each cell, as in the earlier stages, is bounded by a membrane-like wall.

Accompanying these general changes special alterations take place in the distribution of the nuclei and the protoplasm. The nuclei, accompanied by protoplasm, gradually leave the centre and migrate towards the periphery of the notochord. At the same time the protoplasm of the cells forms a special layer in contact with the investing sheath.

The changes by which this takes place can easily be followed in longitudinal sections. In Pl. 12, fig. 11 the migration of the nuclei has commenced. They are still, however, more or less aggregated at the centre, and very little protoplasm is present at the edges of the notochord. The cells, though more or less irregularly polygonal, are still somewhat flattened. In Pl. 12, fig. 12 the notochord has made a further progress. The nuclei now mainly lie at the side of the notochord, where they exist in a somewhat shrivelled state, though still invested by a layer of protoplasm.

A large portion of the protoplasm of the cord forms an almost continuous layer in close contact with the sheath, which is more distinctly visible in some cases than in others.

While the changes above described are taking place the notochord increases in size. At the age of fig. 11 it is in the anterior part of the body of Pristiurus about 0.11 Mm. At the age of fig. 12 it is in the same species 0.12 Mm., while in Scyllium stellare it reaches about 0.17 Mm.

During stage K (Pl. 11, fig. 8) the vacuolation of the cells of the notochord becomes even more complete than during the earlier stages, and in the central cells hardly any protoplasm is present, though a starved nucleus surrounded by a little protoplasm may be found in an occasional corner.

[Pg 357] The whole notochord becomes very delicate, and can with great difficulty be conserved whole in transverse sections.

The layer of protoplasm which appeared during the last stage on the inner side of the cuticular membrane of the notochord becomes during the present stage a far thicker and more definite structure. It forms a continuous layer with irregular prominences on its inner surface; and contains numerous nuclei. The layer sometimes presents in transverse sections hardly any indication of a division into a number of separate cells, but in longitudinal sections this is generally very obvious. The cells are directed very obliquely forwards, and consist of an oblong nucleus invested by protoplasm. The layer formed by them is very delicate and very easily destroyed. In one example its thickness varied from .004 to .006 Mm., in another it reached .012 Mm. The thickness of the cuticular membrane is about .002 Mm. or rather less.

The diameter of a notochord in the anterior part of the body of a Pristiurus embryo of this stage is about 0.21 Mm. Round the exterior of the notochord the mesoblast cells are commencing to arrange themselves as a special sheath.

In Torpedo the notochord at first presents the same structure as in Pristiurus, i.e. it forms a cylindrical rod of flattened cells.

The vacuolation of these cells does not however commence till a relatively very much later period than in Pristiurus, and also presents a very different character (Pl. 11, fig. 7).

The vacuoles are smaller, more numerous, and more rounded than in the other genera, and there can be no question that in many cases there is more than one vacuole in a cell. The most striking point in which the notochord of Torpedo differs from that of Pristiurus consists in the fact that in Torpedo there is never any aggregation of the nuclei at the centre of the cord, but the nuclei are always distributed uniformly through it. As the vacuolation proceeds the differences between Torpedo and the other genera become less and less marked. The vacuoles become angular in form, and the cells of the cord cease to be flattened, and become polygonal.

At my final stage for Torpedo (slightly younger than K) the only important feature distinguishing the notochord from that [Pg 358] of Pristiurus, is the absence of any signs of nuclei or protoplasm passing to the periphery. Around the exterior of the cord there is early found in Torpedo a special investment of mesoblastic cells.

EXPLANATION OF PLATES 11 AND 12.

Complete List of Reference Letters.

al. Alimentary tract. an. Point where anus will be formed. ao. Dorsal aorta. ar. Rudiment of anterior root of spinal nerve. b. Anterior fin. c. Connective-tissue cells. cav. Cardinal vein. ch. Notochord. df. Dorsal fin. ep. Epiblast. ge. Germinal epithelium. ht. Heart. l. Liver. mp. Muscle-plate. mp´. Early formed band of muscles from the splanchnic layer of the muscle-plates. nc. Neural canal. p. Protoplasm from yolk in the alimentary tract. pc. Pericardial cavity. po. Primitive ovum. pp. body-cavity. pr. Rudiment of posterior root of spinal nerve. sd. Segmental duct. sh. Cuticular sheath of notochord. so. Somatic layer of mesoblast. sp. Splanchnic layer of mesoblast. spc. Spinal cord. sp.v. Spiral valve. sr. Interrenal body. st. Segmental tube. sv. Sinus venosus. ua. Umbilical artery. um. Umbilical cord. uv. Umbilical vein. V. Splanchnic vein. v. Blood-vessel. vc. Visceral cleft. vr. Vertebral rudiment. W. White matter of spinal cord. x. Subnotochordal rod (except in fig. 14a). y. Passage connecting the neural and alimentary canals.

Plate 11.

Fig. 1. Section from the caudal region of a Pristiurus embryo belonging to stage H. Zeiss C, ocul. 1. Osmic acid specimen.

It shews (1) the constriction of the subnotochordal rod (x) from the summit of the alimentary canal. (2) The formation of the body-cavity in the muscle-plate and the ventral thickening of the parietal plate.

Fig. 1a. Portion of alimentary wall of the same embryo, shewing the formation of the subnotochord rod (x).

Fig. 2. Section through the caudal vesicle of a Pristiurus embryo belonging to stage H. Zeiss C, ocul. 1.

It shews the bilobed condition of the alimentary vesicle and the fusion of the mesoblast and hypoblast at the caudal vesicle.

Fig. 3a. Sections from the caudal region of a Pristiurus embryo belonging to stage H. Zeiss C, ocul. 1. Picric acid specimen.

It shews the communication which exists posteriorly between the neural and alimentary canals, and also by comparison with 3b it exhibits the dilatation undergone by the alimentary canal in the caudal vesicle.

Fig. 3b. Section from the caudal region of an embryo slightly younger than 3a. Zeiss C, ocul. 1. Osmic acid specimen.

[Pg 359] Fig. 4. Section from the cardiac region of a Pristiurus embryo belonging to stage H. Zeiss C, ocul. 1. Osmic acid specimen.

It shews the formation of the heart (ht) as a cavity between the splanchnopleure and the wall of the throat.

Fig. 5. Section from the posterior dorsal region of a Scyllium embryo, belonging to stage H. Zeiss C, ocul. 1. Osmic acid specimen.

It shews the general features of an embryo of stage H, more especially the relations of the body-cavity in the parietal and vertebral portions of the lateral plate, and the early-formed band of muscle (mp´) in the splanchnic layer of the vertebral plate.

Fig. 6. Section from the œsophageal region of Scyllium embryo belonging to stage I. Zeiss C, ocul. 1. Chromic acid specimen.

It shews the formation of the rudiments of the posterior nerve-roots (pr) and of the vertebral rudiments (Vr).

Fig. 7. Section of a Torpedo embryo belonging to stage slightly later than I. Zeiss C, ocul. 1, reduced 1/3. Osmic acid specimen.

It shews (1) the formation of the anterior and posterior nerve-roots. (2) The solid knob from which the segmental duct (sd) originates.

Fig. 8. Section from the dorsal region of a Scyllium embryo belonging to a stage intermediate between I and K. Zeiss C, ocul. 1. Chromic acid specimen.

It illustrates the structure of the primitive ova, segmental tubes, notochord, etc.

Fig. 8a. Section from the caudal region of an embryo of the same age as 8. Zeiss A, ocul. 1.

It shews (1) the solid œsophagus. (2) The narrow passage connecting the pericardial (pc) and body cavities (pp).

Fig. 9. Section of a Pristiurus embryo belonging to stage K. Zeiss A, ocul. 1. Osmic acid specimen.

It shews the formation of the liver (l), the structure of the anterior fins (b), and the anterior opening of the segmental duct into the body-cavity (sd).

Figs. 9a, 9b, 9c, 9d. Four sections through the anterior region of the same embryo as 9. Osmic acid specimens.

The sections shew (1) the atrophy of the post-anal section of the alimentary tract (9b, 9c, 9d). (2) The existence of the segmental tubes behind the anus (9b, 9c, 9d). With reference to these it deserves to be noted that the segmental tubes behind the anus are quite disconnected, as is proved by the fact that a tube is absent on one side in 9c but reappears in 9d. (3) The downward prolongation of the segmental duct to join the posterior or cloacal extremity of the alimentary tract (9b).

Plate 12.

Fig. 10. Longitudinal and horizontal section of a Scyllium embryo of stage H. Zeiss C, ocul. 1. Reduced by 1/3. Picric acid specimen.

It shews (1) the structure of the notochord; (2) the appearance of the early formed band of muscles (mp´) in the splanchnic layer of the protovertebra.

Fig. 11. Longitudinal and horizontal sections of an embryo belonging to stage I. Zeiss C, ocul. 1. Chromic acid specimen. It illustrates the same points as the previous section, but in addition shews the formation of the rudiments of the vertebral bodies (Vr) which are seen to have the same segmentation as the muscle-plates.

[Pg 360] Fig. 12.[227] Longitudinal and horizontal section of an embryo belonging to the stage intermediate between I and K. Zeiss C, ocul. 1. Osmic acid specimen illustrating the same points as the previous section.

Fig. 13. Longitudinal and horizontal section of an embryo belonging to stage K. Zeiss C, ocul. 1, and illustrating same points as previous section.

Figs. 14a, 14b, 14c, 14d. Figures taken from preparations of an embryo of an age intermediate between I and K, and illustrating the structure of the primitive ova. Figs. 14a and 14b are portions of transverse sections. Zeiss C, ocul. 3 reduced 1/3. Figs. 14c and 14d are individual ova, shewing the lobate form of nucleus. Zeiss F, ocul. 2.

Fig. 15. Osmic acid preparation of primitive ova belonging to stage K. Zeiss immersion No. 2, ocul. 1. The protoplasm of the ova is seen to be nearly filled with bodies resembling yolk-spherules: and one ovum is apparently undergoing division.

Fig. 15a. Picric acid preparation shewing a primitive ovum partially filled with bodies resembling yolk-spherules.

Fig. 16. Horizontal and longitudinal section of Scyllium embryo belonging to stage K. Zeiss A, ocul. 1. Picric acid preparation. The connective-tissue cells are omitted.

The section shews that there is one segmental tube to each vertebral segment.

Fig. 17. Portion of a Scyllium embryo belonging to stage K, viewed as a transparent object.

It shews the segmental duct and the segmental involutions—two of which are seen to belong to segments behind the end of the alimentary tract.

Fig. 18. Vertical longitudinal section of a Scyllium embryo belonging to stage K. Zeiss A, ocul. 1. Hardened in a mixture of osmic and chromic acid. It shews

(1) the commissures connecting together the posterior roots of the spinal nerves;

(2) the junction of the anterior and posterior roots;

(3) the relations of the segmental ducts to the segmental involutions and the alternation of calibre in the segmental tube;

(4) the germinal epithelium lining the body-cavity.

[192] Unless the contrary is stated, the facts recorded in this chapter apply only to the genera Scyllium and Pristiurus.

[193] The layers are known as epidermic (horny) and mucous layers by English writers, and as Hornschicht and Schleimschicht by the Germans. For their existence in all Vertebrates, vide Leydig Ueber allgemeine Bedeckungen der Amphibien, p. 20. Bonn, 1876.

[194] Vide Leydig, loc. cit.

[195] Vide Götte, Entwicklungsgeschichte der Unke.

[196] Vide Self, Development of Spinal Nerves in Elasmobranchii. Phil. Trans. 1876. [This Edition, No. VIII.]

[197] For Birds, vide Elements of Embryology, Foster and Balfour, pp. 144, 145, and for Mammals, Kölliker, Entwicklungsgeschichte, p. 283.

[198] For the nervous supply in fishes, vide Stannius, Peripher. Nerv. System d. Fische. In Osseous Fishes he states that the thoracic fin is supplied by branches from the first three though sometimes from the first four spinal nerves. In Acipenser there are branches from the first six nerves. In Spinax the limb is supplied by the rami anteriores of the fourth and succeeding ten spinal nerves. In the Rays not only do the sixteen anterior spinal nerves unite to supply the fin, but in all there are rami anteriores from thirty spinal nerves which pass to the thoracic limb.

[199] Philosophical Transactions, 1871.

[200] Ursprung d. Wirbelthiere and Functionswechsels.

[201] Grundriss d. Vergleichenden Anat. p. 494.

[202] Loc. cit.

[203] No attempt has been made to describe in detail the different appearances presented by the protovertebræ in the various parts of the body, but in each stage a protovertebra from the dorsal region is taken as typical.

[204] Zeitschrift f. Anat. Entwicklungsgeschichte, Vol. 1.

[205] Hensen loc. cit.

[206] For the history of protovertebræ and muscle-plates in Birds, vide Elements of Embryology, Foster and Balfour. The statement there made that the horizontal splitting of the mesoblast does not extend to the summit of the vertebral plate, must however be regarded as doubtful.

[207] Vide Elements of Embryology, p. 56.

[208] Dr Götte, Entwicklungsgeschichte der Unke, p. 534, gives a different account of the development of the protovertebræ from that in the text. He states that the muscle-plates do not give rise to the main dorso-lateral muscles, but only to some superficial ventral muscles, while the dorso-lateral muscles are according to him formed from part of the kernel of the protovertebræ internal to the muscle-plates. The account given in the text is the result of my own investigations, and accords precisely with the recent statements of Professor Kölliker, Entwicklungsgeschichte, 1876.

[209] The type of development of the muscle-plates of Amphibians would become identical with that of Elasmobranchii if their first-formed mass of muscle corresponded with the early-formed muscles of Elasmobranchii, and the remaining cells of both layers of the protovertebræ became in the course of development converted into muscle-cells indistinguishable from those formed at first. Is it possible that, owing to the distinctness of the first-formed mass of muscle, Dr Götte can have overlooked the fact that its subsequent growth is carried on at the expense of the adjacent cells of the somatic layer?

[210] Ehrlich, Ueber den peripher. Theil d. Urwirbel. Archiv f. Mic. Anat. Vol. XI.

[211] The most important other instances in addition to that of the Cœlenterata which can be adduced in favour of the epiblastic origin of the mesoblast are the Bird and Mammal, in which according to the recent observations of Hensen for the Mammal, and Kölliker for the Mammal and Bird, the mesoblast is split off from the epiblast. If the views I have elsewhere put forward about the meaning of the primitive groove be accepted, the derivation of the mesoblast from the epiblast in these instances would be apparent rather than real, and have no deep morphological significance for the present question.

Other instances may be brought forward from various groups, but none of these are sufficiently well confirmed to be of any value in the determination of the present question.

[212] Vide Anthropogenie, p. 197.

[213] Vide Self, Development of Elasmobranch Fishes, Journal of Anat. and Phys. Vol. X. note on p. 682, and also Review of Professor Kölliker's Entwicklungsgeschichte des Menschen u. d. höheren Thiere, Journal of Anat. And Phys. Vol. X.

[214] Professor Haeckel speaks of the splitting of the mesoblast in Vertebrates into a somatic and splanchnic layer as a secondary process (Gastrula u. Eifurchung d. Thiere), but does not make it clear whether he regards this secondary splitting as taking place along the old lines. It appears to me to be fairly certain that even if the original unsplit condition of the mesoblast is to be regarded as a secondary condition, yet that the splitting of this must take place along the old lines, otherwise a change in the position of the body-cavity in the adult would have to be supposed—an unlikely change producing unnecessary complication. The succeeding argument is based on the assumption that the unsplit condition is a secondary condition, but that the split which eventually appears in this occurs along the old lines, separating the primitive splanchnopleure from the primitive somatopleure.

[215] Quart. Jl. of Micros. Science, July, 1875. [This Edition, No. VI.]

[216] Jenaische Zeitschrift, Vol. IX.

[217] Quart. Jl. of Micros. Science, Vol. XXV. 1874, and Phil. Trans. 1875.

[218] Archives de Zoologie, Vol. IV.

[219] Archiv f. Micr. Anat. Vol. XIII.

[220] The recent researches of Selenka, Zeitschrift f. Wiss. Zoologie, Vol. XXVII. 1876, demonstrate that in Echinoderms the muscles are derived from the cells first split off from the hypoblast, and that the diverticula only form the water-vascular system and the epithelial lining of the body-cavity.

[221] Kowalevsky, Würmer u. Arthropoden, Mém. Acad. Pétersbourg, 1871.

[222] Zur Entwicklungsgeschichte d. Brachiopoden, Protokoll d. ersten Session der Versammlung Russischer Naturforscher in Kasan, 1873. Published in Kaiserliche Gesellschaft Moskau, 1874 (Russian). Abstracted in Hoffmann and Schwalbe, Jahresbericht f. 1873.

[223] Comparison of Early Stages, Quart. Jl. Micros. Science, July, 1875. [This Edition, No. VI.]

[224] Urinogenital Organs of Vertebrates, Journ. of Anat. and Phys. Vol. X. [This Edition, No. VII.]

[225] Entwicklungsgeschichte der Unke, Pl. 1, fig. 8.

[226] This membrane is better looked upon, as is done by Gegenbaur and Götte, as intercellular matter.

[227] The apparent structure in the sheath of the notochord in this and the succeeding figure is merely the result of an attempt on the part of the engraver to represent the dark colour of the sheath in the original figure.

[Pg 361]

CHAPTER VII.

General Development of the Trunk from Stage H to the Close of Embryonic Life.

External Epiblast.

The change already alluded to in the previous chapter (p. 317) by which the external epiblast or epidermis becomes divided into two layers, is completed before the close of stage L.

In the tail region at this stage three distinct strata may be recognized in the epidermis. (1) An outer stratum of flattened horny cells, which fuse together to form an almost continuous membrane. (2) A middle stratum of irregular partly rounded and partly flattened cells. (3) An internal stratum of columnar cells, bounded towards the mesoblast by a distinct basement membrane (Pl. 13, fig. 8), unquestionably pertaining to the epiblast. This layer is especially thickened in the terminal parts of the paired fins (Pl. 13, fig. 1). The two former of these strata together constitute the epidermic layer of the skin, and the latter the mucous layer.

In the anterior parts of the body during stage L the skin only presents two distinct strata, viz. an inner somewhat irregular layer of rounded cells, the mucous layer, and an outer layer of flattened cells (Pl. 13, fig. 8).

The remaining history of the external epiblast, consisting as it does of a record of the gradual increase in thickness of the epidermic strata, and a topographical description of its variations in structure and thickness in different parts, is of no special interest and need not detain us here.

In the late embryonic periods subsequent to stage Q the layers of the skin cease to be so distinct as at an earlier period, [Pg 362] partly owing to the innermost layer becoming less columnar, and partly to the presence of a large number of mucous cells, which have by that stage made their appearance.

I have followed with some care the development of the placoid scales, but my observations so completely accord with those of Dr O. Hertwig[228], that it is not necessary to record them. The so-called enamel layer is a simple product of the thickening and calcification of the basement membrane, and since this membrane is derived from the mucous layer of the epidermis, the enamel is clearly to be viewed as an epidermic product. There is no indication of a gradual conversion of the bases of the columnar cells forming the mucous layer of the epidermis into enamel prisms, as is frequently stated to occur in the formation of the enamel of the teeth in higher Vertebrates.

Lateral line.

The lateral line and the nervous structures appended to it have been recently studied from an embryological point of view by Götte[229] in Amphibians and by Semper[230] in Elasmobranchii.

The most important morphological result which these two distinguished investigators believe themselves to have arrived at is the direct derivation of the lateral nerve from the ectoderm. On this point there is a complete accord between them, and Semper especially explains that it is extremely easy to establish the fact.

As will appear from the sequel, I have not been so fortunate as Semper in elucidating the origin of the lateral nerve, and my observations bear an interpretation not in the least in accordance with the views of my predecessors, though not perhaps quite conclusive against them.

It must be premised that two distinct structures have to be dealt with, viz. the lateral line formed of modified epidermis, and the lateral nerve whose origin is in question.

The lateral line is the first of the two to make its appearance, at a stage slightly subsequent to K, in the form of a [Pg 363] linear thickening of the inner row of cells of the external epiblast, on each side, at the level of the notochord.

This thickening, in my youngest embryo in which it is found, has but a very small longitudinal extension, being present through about 10 thin sections in the last part of the head and first part of the trunk. The thickening, though short, is very broad, measuring about 0.28 Mm. in transverse section, and presents no signs of a commencing differentiation of nervous structures. The large intestinal branch of the vagus can be seen in all the anterior sections in close proximity to this line, and appears to me to give off to it posteriorly a small special branch which can be traced through a few sections, vide Pl. 13, fig. 2, n.l. But this branch is not sufficiently well marked to enable me to be certain of its real character. In any case the posterior part of the lateral line is absolutely without any adjoining nervous structures or traces of such.

The rudiment of the epidermic part of the lateral line is formed of specially elongated cells of the mucous layer of the epiblast, but around the bases of these certain rounder cells of a somewhat curious appearance are intercalated.

There is between this and my next youngest embryo an unfortunately large gap with reference to the lateral line, although in almost every other respect the two embryos might be regarded as belonging to the same stage. The lateral line in the older embryo extends from the hind part of the head to a point well behind the anus, and is accompanied by a nerve for at least two-thirds of its length.

In the foremost section in which it appears the intestinal branch of the vagus is situated not far from it, and may be seen at intervals giving off branches to it. There is no sign that these are otherwise than perfectly normal branches of the vagus. Near the level of the last visceral cleft the intestinal branch of the vagus gives off a fair-sized branch, which from the first occupies a position close to the lateral line though well within the mesoblast (Pl. 13, fig. 3a, n.l). This branch is the lateral nerve, and though somewhat larger, is otherwise much like the nerve I fancied I could see originating from the intestinal branch of the vagus during the previous stage.

It rapidly thins out posteriorly and also approaches closer [Pg 364] and closer to the lateral line. At the front end of the trunk it is quite in contact with it, and a short way behind this region the cells of the lateral line arrange themselves in a gable-like form, in the angle of which the nerve is situated (Pl. 13, figs. 3b, and 3c). In this position the nerve though small is still very distinct in all good sections, and is formed of a rod of protoplasm, with scattered nuclei, in which I could not detect a distinct indication of cell-areas. The hinder part of the nerve becomes continually smaller and smaller, without however presenting any indication of becoming fused with the epiblast, and eventually ceases to be visible some considerable distance in front of the posterior end of the lateral line.

The lateral line itself presents some points of not inconsiderable interest. In the first place, it is very narrow anteriorly and throughout the greater part of its length, but widens out at its hinder end, and is widest of all at its termination, which is perfectly abrupt. The following measurements of it were taken from an embryo belonging to stage L, which though not quite my second youngest embryo is only slightly older. At its hinder end it was 0.17 Mm. broad. At a point not far from this it was 0.09 Mm. broad, and anteriorly it was 0.05 Mm. broad. These measurements clearly shew that the lateral line is broadest at what may be called its growing-point, a fact which explains its extraordinary breadth in the anterior part of the body at my first stage, viz. 0.28 Mm., a breadth which strangely contrasts with the breadth, viz. 0.05 Mm., which it has in the same part of the body at the present stage.

It still continues to form a linear area of modified epidermis, and has no segmental characters. Anteriorly it is formed by the cells of the mucous layer becoming more columnar (Pl. 13 fig. 3a). In its middle region the cells of the mucous layer in it are still simply elongated, but, as has been said above, have a gable-like arrangement, so as partially to enclose the nerve (Pl. 13, fig. 3b). Nearer the hind end of the trunk a space appears in it between its columnar cells and the flattened cells of the outermost layer of the skin (Pl. 13, fig. 3c), and this space becomes posteriorly invested by a very definite layer of cells. The space (Pl. 13, fig. 3d) or lumen has a slit-like section, and is not formed by the closing in of an originally open groove, but by [Pg 365] the formation of a cavity in the midst of the cells of the lateral line. Its walls are formed by a layer of columnar cells on the inner side, and flattened cells on the outer side, both layers however appearing to be derived from the mucous layer of the epidermis. The outer layer of cells attains its greatest thickness dorsally.

During stages M, N, O, the lateral nerve gradually passes inwards into the connective tissue between the dorso-lateral and the ventro-lateral muscles, and becomes even before the close of stage N completely isolated from the lateral line.

The growth of the lateral line itself remains for some time almost stationary; anteriorly the cells retain the gable-like arrangement which characterised them at an earlier period, but cease to enclose the nerve; posteriorly the line retains its original more complicated constitution as a closed canal. In stage O the cells of the anterior part of the line, as well as those of the posterior, commence to assume a tubular arrangement, and the lateral line takes the form of a canal. The tubular form is due to a hollowing out of the lateral line itself and a rearrangement of its cells. As the lateral line becomes converted into a canal it recedes from the surface.

In stage P the first indication of segmental apertures to the exterior make their appearance, vide Pl. 13, fig. 4. The lateral line forms a canal situated completely below the skin, but at intervals (corresponding with segments) sends upwards and outwards prolongations towards the exterior. These prolongations do not during stage P acquire external openings. As is shewn in my figure, a special area of the inner border of the canal of the lateral line becomes distinguished by its structure from the remainder.

No account of the lateral line would be complete without some allusion to the similar sensory structures which have such a wide distribution on the heads of Elasmobranchii; and this is especially important in the present instance, owing to the light thrown by a study of their development on the origin of the nerves which supply the sense-organs of this class. The so-called mucous canals of the head originate in the same way as does the lateral line; they are products of the mucous layer of the epidermis. They eventually form either canals with numerous [Pg 366] openings to the exterior, or isolated tubes with terminal ampulliform dilatations.

I have not definitely determined whether the canal-system of the head arises in connection with the lateral line, or only eventually becomes so connected. The important point to be noticed is, that at first no nervous structures are to be seen in connection with it. In stage O nerves for the mucous canals make their appearance as delicate branches of the main stems. These nerve-stems are very much ramified, and their branches have, in a large number of instances, an obvious tendency towards a particular sense-organ (Pl. 13, figs. 5 and 6).

I have not during stage O been able to detect a case of direct continuity between the two. This is, however, established in the succeeding stage P, in the case of the canals, and the facility with which it may be observed would probably render the embryo Elasmobranch a very favourable object for studying the connection between nerves and terminal sense-organs. The nerve (Pl. 13, fig. 7) dilates somewhat before uniting with the sense-organ, and the protoplasm of the nerve and the sense-organ become completely fused. The basement membrane of the skin is not continuous across their point of junction, and appears to unite with a delicate membrane-like structure, which invests the termination of the nerve. The ampullæ would seem to receive their nervous supply somewhat later than the canals, and the terminal swellings of the nerves supplying them are larger than in the case of the canals, and the connection between the ampullæ and the nerves not so clear. In the case of the head, there can for Elasmobranchii be hardly a question that the nerves which supply the mucous canals grow centrifugally from the original cranial nerve-stems, and do not originate in a peripheral manner from the integument.

This is an important point to make certain of in settling any doubtful features in the nervous supply of the lateral line. Professor Semper[231], with whom as dealing with Elasmobranchii we are more directly concerned, makes the following statement: At the time when at the front end the lateral nerve has already completely separated itself from the ectoderm, and is situated amongst the muscles, it still lies in the middle of the body close [Pg 367] to the ectoderm, and at the hind end of the body is not yet completely segmented off (abgegliedert) from the ectoderm. Although the last sentence of this quotation may seem to be opposed to my statements, yet it appears to me probable that Professor Semper has merely seen the lateral nerve partially enclosed in the ectoderm. This position of the nerve no doubt affords a presumption, but only a presumption, in favour of a direct origin of the lateral nerve from the ectoderm; but against this interpretation of it are the following facts:

(1) That the front part of the lateral line is undoubtedly supplied by branches which arise in the ordinary way from the intestinal branch of the vagus; and we should not expect to find part of the lateral line supplied by nerves which originate in one way, and the remainder supplied by a nerve having a completely different and abnormal mode of origin.