i Be eo THE ELEMENTS OF EMBRYOLOGY. PART I. THE ELEMENTS OF EMBRYOLOGY. BY M. FOSTER, M.A, MD., FRS., ae FELLOW OF AND PRHELECTOR IN PHYSIOLOGY IN TRINITY COLLEGE, CAMBRIDGE, AND FRANCIS M. BALFOUR, B.A... FELLOW OF TRINITY COLLEGE, CAMBRIDGE. London: MACMILLAN AND CO. 1874. [All Rights reserved.] Cambridge: PRINTED BY C, J. CLAY, M.A. AT THE UNIVERSITY PRESS. TO THOMAS HENRY HUXLEY AS A LITTLE TOKEN OF OUR APPRECIATION OF HIS WORTI AND OF HIS MUCH KINDNESS TO OURSELVES THIS BOOK IS RESPECIFULLY DEDICATED BY THE AUTHORS. PREFACE. Tn this volume we offer to the public the first part of what we hope may serve as a systematic introduction to the study of Embryology. Some apology is perhaps necessary for the separate publication of a part only of the whole subject; but we trust that the following reasons will justify the course we have adopted. Those who have paid attention to recent embryological researches must be aware of what we may venture to call the ‘tumultuous condition of many parts of the subject, and of the extreme difficulty in many cases of forming a clear and decided judgment without the aid of independent observations. It is this necessity of having repeatedly to work over contested points with a view to reconcile diametrically opposed statements, or to verify startling announcements, which has rendered so laborious the task we have undertaken, and which so much delays its com- pletion. On the other hand, whoever wishes to have a sound foundation of embryological knowledge cannot do better than gain a thorough insight into the development of the bird. The practical advantages offered by the hen’s egg b2 Viil PREFACE. altogether outweigh the theoretical objections to beginning with the avian type. In many respects, it might be thought desirable to commence with a holoblastic ovum; but the large food-yolk of the bird’s egg is in many ways a great assistance to the study of changes going on in the blasto- derm, The chick is of all embryos the best to begin with ; when its history has once been mastered, the subsequent study of other forms becomes an easy matter. We venture to hope therefore that we shall meet with general approval, in having described at considerable length the history of the chick, and in hastening the publication of our account, by bringing it forward in a separate form. In the earlier chapters, especially, we have gone into very considerable detail; and in order to make the account intelligible to the beginner, have not been deterred by the fear of wearying our readers with elementary and recapitu- latory statements. Debated matters and details of minor importance have been put in small print; these may be omitted by the student in reading the book for the first time. Though we have sometimes introduced names in connection with important observations, we have not thought it necessary to do this systematically. For recent or debated statements however, the authorities are always cited. The worth of such a book as this will be very small if the student simply contents himself with reading what is written; and to facilitate the only really useful mode of study, that of actual observation, a few practical instructions have been added in an appendix. The readiness with which the development of the skull can be studied in the chick renders it, in spite of obvious PREFACE. 1x objections, a suitable introduction to the important subject of cranial morphology. It is with this view that we have given a separate chapter on the skull, which we hope may serve as an introduction to the study of Mr Parker’s elaborate memoirs. In the remaining parts, which we shall do our best to complete as soon as possible, the several histories will be treated with much greater brevity, and much more space will be given to theoretical considerations. The figures, whose source is not acknowledged in the text, were drawn by Miss A. B. Balfour, except a few by ourselves. The drawing on wood was executed partly by Mr Allchin, but chiefly by Mr Collings; and all the drawings were cut by Mr Cooper. We have to thank those gentlemen for the trouble they have taken in a matter in which, for many reasons, the result never seems commensurate with the labour. We are much indebted to Professor Huxley for having kindly looked over the proofs of the Chapter on the Skull. The work took its origin in a course of lectures delivered by myself, but many causes prevented my taking the task seriously in hand, until I was joined by my friend and former pupil Mr F. M. Balfour, whose share in the matter has, to say the least, been no less than my own. M. FOSTER. TABLE OF CONTENTS. RPEOUUORION cau) isl oe. 2 le 1. bly ee Pps to. PART I. THE HISTORY OF THE CHICK. CHAPTER I. THE STRUCTURE OF THE HEN’S EGG, AND THE CHANGES WHICH TAKE PLACE UP TO THE BEGINNING OF INCUBATION . 4 pp. 11—26. 1. The shell. 2. The shell-membrane. 3. The albumen. 4. The vitelline membrane. 5. The yolk. 6. The yellow yolk. 7. The white yolk. 8. The white yolk-spheres. 9. The structure of the blastoderm. to. Recapitulation. 11. The ovarian ovum. 12. The descent of the ovum along the oviduct. 13. The impregnation of the ovum. 14. Segmentation. 15. The formation of the upper and lower layers, CHAPTER II. A Brier SUMMARY OF THE WHOLE History OF INCUBATION, pp. 27—42. 1. The embryo is formed in the area pellucida. 2. The epiblast, mesoblast, and hypoblast. 3. The extension of the blastoderm over the yolk. 4. The vascular area. 5. The head-fold and the other folds by means of which the embryonic sac is formed. 6. The outward shape of the embryo. 7. The formation of the neural tube and alimentary canal: somatopleure and splanch- nopleure. 8. Theamnion. g. The allantois, x CONTENTS. CHAPTER III. THE CHANGES WHICH TAKE PLACE DURING THE First Day or IncuBATION, PP. 43—57- 1. Variations in the progress of development. 2. The embryonic shield. 3. The formation of the epiblast, mesoblast and hypoblast. 4. The primitive streak, the primitive groove. 5. The head-fold, the medullary groove, me- dullary folds, and notochord. 6. The amnion; the changes taking place in the three layers. 7. The increase of the head-fold. 8. The closure of the me- dullary canal. 9, 10. The cleavage of the mesoblast: formation of splanchno- pleure and somatopleure. 11. The protovertebre. 12. The formation of the vascular area. 13. Recapitulation. CHAPTER IV. THE CHANGES WHICH TAKE PLACE DURING THE SEconD Day, pp. 58—83. 1. Increasing distinctness and prominence of the embryo. 2. The first cere- bral vesicle. 3. The increase in the number of protovertebre. 4. The first rudiments of the alimentary canal. 5. The formation of the heart. 6. The formation of blood-vessels’ the ompHalo-niesaraic veins and arteries, the sinus terminalis. 7. Changes taking place in the cells of the several layers. 8. The rudiment of the Wolffian duct. 9g. Recapitulation of the changes during the first half of the second day. to. Increasing prominence of the embryo; the tail-fold and the lateral folds. 11. Continued closure of the medullary canal. 12. The optic vesicles. 13. The second and third cerebral vesicles. 14. Change of position of the optic vesicles. 15. The vesicles of the cerebral hemispheres. 16. The cranial flexure. 17. The rudiment of the ear, or auditory sac. 18. Changes in the heart. 19. The primitive aorte and first pair of aortic arches, the omphalo-mesaraic vessels, the sinus terminalis. 20. The second and third pair of aortic arches. 21. The Wolffian duct. 22. The amnion. 23. Recapitulation. CHAPTER V. Tse CHANGES WHICH TAKE PLACE DURING THE THIRD Day, pp. 84—140. i. The diminution of the albumen. 2. The spreading of the opaque and vascular areas. 3. The'vascular area. 4. The continued folding in of the embryo. 5. The increase of the amnion. 6. The change in the position of the embryo. 7. The curvature of the body. 8. The cranial flexure. 9. Growth of the vesicles of the cerebral hemispheres; the third ventricle, pineal gland, infundibulum and pituitary body, the cerebellum and medulla oblongata. 1o. Changes in the spinal cord. 11. The formation of the eye. Histological changes in the retina, optic nerve, and lens. 12. The formation of the ear, 13. The nasal pits. 14. The visceral clefts and folds. 15. The aortic arches. 16. Changes in the heart; the Ductus Cuvieri and cardinal veins. 17. The folding in of the alimentary canal; the formation of the tail. 18. The lungs. 19. The liver. 20. The pancreas and spleen. 21. The thyroid body. 22. Changes in the trunk of the embryo. 23. Separation of the muscle-plates from the protovertebre. 24. Growth of the intermediate pitas 25. The cranial nerves. 26. The Wolffian duct. 27. Recapi- ulation. : CONTENTS. xlil CHAPTER VI. THE CHANGES WHICH TAKE PLACE DURING THE FourtH Day, pp. 141—173- s, Appearances on opening the eg 2. Growth of the amnion. 3. Narrowing of the splanchnic stalk. a: " Increase in the cranial flexure. 5. The first appearance of the limbs. 6. Growth of the head. 7. Changes in the nasal pits. 8. Formation of the mouth. 9. The cranial nerves. ro. The allantois. rz. Changes in the protovertebre; the spinal ganglia. 12. The secondary segmentation of the vertebral column. 13. Changes in the notochord. 14. Ossification of the vertebre. 15. The ribs. 16. Changes in the muscle-plates. 17. The Wolffian body and duct. 18. The duct of Miiller. 39. The kidneys. 20. ‘The ovaries and testes. 21. Changes in the arterial system. 22. Changes in the venous system; the veins of the liver. 23. Changes in the heart; the ventricular septum. 24. . Recapitalation. CHAPTER VII. THE CHANGES WHICH TAKE PLACE ON THE FIFTH Day, pp. 174—199. 1. Appearances on opening the ege. 2. The growth of the limbs. 3. The cranium; the investing mass and trabecule. 4. Changes in the face; formation of the nose and nasal passages. 5. Appearance of the anus. 6. Changes in the spinal cord; the formation of the grey and white columns, and of the posterior and anterior fissures. 7. Changes in the heart; the rudiment of the auricular septum, the division of the bulbus arteriosus into aorta and pulmonary artery, the formation of the semilunar valves. 8. Changes in the heart during the sixth day. 9. Subsequent changes in the heart; the completion of the auricular septum, the arrangement of the openings of the vene cave. 10. Histological differentiation; the fate of the three primary layers. 11. Recapitulation. CHAPTER VIII. From THe SrxtH# Day TO THE END OF INCUBATION, pp. 200—224. 1. The commencement of distinct avian differentiation. 2. The foetal appendages during the sixth and seventh days. 3. During the eighth, ninth _and tenth days. 4. From the eleventh to the sixteenth day. 5. From the sixteenth day onwards. 6. The changes in the general form of the embryo during the sixth and seventh days. 7. During the eighth, ninth and tenth days. 8. From the eleventh day onwards ; feathers, ossifications. 9. Changes in the venous system before and after the commencement of pulmonary respi- ration. ro. Changes in the arterial system, the modifications of the aortic arches. 11. Summary of the chief phases of the circulation. 12. EHx- clusion from the egg. XIV CONTENTS. CHAPTER IX. Tue DEVELOPMENT OF THE SKULL, pp. 225—238. 1, 2. The primordial cranium. 3, 4. The investing mass of Rathke. 5. The trabecule cranii. 6. The cartilages of the first visceral arch. 7. ‘The maxillary process. 8. The mandibular arch. 9. The hyoid arch. Io. The cartilages of the third visceral arch. 11. Changes in the cranium during the fifth and sixth days. 12. During and after the seventh day. 13. The condition of the cranium at about the middle of the second week. 14. LEcto- steal and endosteal ossifications of the cartilaginous cranium. 15. Formation of the membrane bones. 16. Progress of ossification during the second and third weeks. 17: Fenestration of the ethmo-presphenoid cartilage. 18. Ossifi- cations in the prootics and alisphenoid. 19. Changes in the basitemporals. Formation of the vomer. 20. The changes which take place immediately after exclusion from the egg. 21. Further changes in the splint bones. Coalescence of the bones after birth. Table of bones classified according to their mode of ossification. APPENDIX. PRACTICAL INSTRUCTIONS FOR STUDYING THE DEVELOPMENT OF THE CHICK, Pp. 239—267. I. Incubators. II. Examination of a 36 to 48 hours embryo. III. Examination of an embryo of about 48—50 hours. IV. Ofan embryo at the end of the third day. V. Of an embryo of the fourth day. VI. Of a blastoderm of 20 hours. VII. Of an unincubated blastoderm. VIII. Of the process of segmentation. IX. Of the later oes of the embryo. X. Study of the development of the blood-vessels. ERRATUM. p- 124, in the description of Fig. 39 B, for ‘Superior vertebral’ substitute ‘Jugular,’ FIG. To. LIST OF ILLUSTRATIONS. PAGE Draeramuatic Section of an Unincubated Fowl’s Egg . . eee A. Yellow yolk-sphere filled with fine granules. -B. White yolk- spheres and spherules of various sizes and presenting different ap- pearances . C - 2 - . . : : See ON Section of a Blastoderm of a Fowl’s Egg at the commencement of Incubation , - 5 : : “ . . - 9 5S Section through the Germinal Disc of the ripe Ovarian Ovum of a Fowl while yet enclosed in its Capsule . 2 ° : = c LG) Surface Views of the early Stages of the Segmentation in a Fowl’s Egg . - . - - - : f - : ‘ om ee tw Surface View of the Germinal Disc of a Hen’s Egg during the later Stages of Segmentation . . ° . * . : - 23 Section of the Germinal Disc of a Fowl during the later Stages of Segmentation . : - - - C : - Wea Ato N. A series of purely diagrammatic representations introduced to facilitate the comprehension of the manner in which the body of the embryo is formed, and of the various relations of the yolk-sac, amnion and allantois. ° 4 : : . : : 29— 32 Diagrammatic Longitudinal Section through the Axis of an Embryo. 33 Section of a Blastoderm at right angles to the long axis of the Embryo after eight hours’ Incubation . - z . : Sects 19. 20. 21. LIST OF ILLUSTRATIONS. Surface View of the Pellucid Area of a Blastoderm of 18 hours Transverse Section of a Blastoderm incubated for 18 hours . Transverse Section through the Dorsal Region of an Embryo of the Second Day * : 5 . 5 5 > ; An Embryo Chick of the First Day (about thirty-six hours) viewed from below as a transparent object 2 é c Embryo of the Chick at 36 hours viewed from above as an opaque object . ‘ . : Diagrammatic Longitudinal Section through the Axis of an Embryo. A, B. Two consecutive Sections of a 36 hours Embryo illustrating the formation of the heart . 5 Transverse Section of an Embryo at the end of the Second Day passing through the region of bulbus arteriosus. ‘ > Surface View from below of a small portion of the posterior end of the pellucid area of a 36 hours Chick . 3 ° Transverse Section through the Dorsal Region of an Embryo of 45 hours . : 5 “ . 5 : “ : . . 6 Embryo of the Chick at 36 hours viewed from above as an Opaque Object 4 : 0 : - Bi : d Head of a Chick at the End of the Second Day viewed from below as a Transparent Object . . = 3 Diagram of the Circulation of the Yolk-Sac at the end of the Third Day of Incubation . : . : - 5 Chick of the Third Day (54 hours) viewed from underneath as a Transparent Object . : 3 : c 5 . Head of a Chick of the Third Day viewed sideways as a Transparent Object . . . z : . : : . 5 . Section through the Hind-Brain of a Chick at the end of the Third Day of Incubation . aM 24 5 3 ‘ : . ; Diagrammatic Sections illustrating the Formation of the Eye. ; Diagrammatic Section of the Eye and the Optic Nerve at an early “ Blage va ae 8S F : ’ , - s é ‘ 5 PAGE 49 51 tn art on LIST OF ILLUSTRATIONS. XVil Fig. PAGE 29. Diagrammatic representation of the Eye of the Chick of about the Third Day as seen when-the head is viewed from underneath as a transparent object - : 5 : : . 2 . - 99 30. D, E, F. Diagrammatic Sections of the Eye of the Chick of about the Third Day . 3 : ~ c A « 7 4 - 100 31. Section of the Eye of Chick at the Fourth Day . é : = To4 32. Section through the Hind-Brain of a Chick at the end of the Third »Day of Incubation - .- . . c . : : : - 10 33. Two Views of the membranous Labyrinth of Columba domestica. A from the exterior, B from the interior ' ¢ F é a 1x2 34. Transverse Section of the Head of a Foetal Sheep (16 mm, in _length) in the region of the Hind-Brain 5 : 0 , a ts) 35. Section of the Head of a Foetal Sheep (20 mm. in length) a ae Tre: 36. Section through ‘the internal Ear of an Embryonic Sheep (28 mm. in length) . - 2 - : 5 : F - 116 37. Head of an Embryo Chick ‘of thé Third Day viewed sideways as an Opaque Object . - C . ° : - A : 2 irs 38. The same Head as shewn in Fig. 37, seen from the Front 4 PI 39 A. Diagram of the Arterial Circulation on the Third Day : stor 39 B.’ Diagram of the Venous Circulation on the Third Day - - 24 40. Section of the Tail-end of an Embryo (Chick) of the Third Day . 125 41. Section through the Dorsal Region of an Embryo at the commence- ‘mentofthe Third Day =. . : F ‘ A . . 126 42. Diagram of a portion of the Digestive Tract of a Chick upon the ESungn Lea ee ee A ae Ca en 2) 43- Four diagrams illustrating the Formation of the Lungs . ; 6) HAG) 44. Geet Aosta the Dorsal Region of an Embryo at the end of the Third Day . c : : - 4 . : : - 135 45. Head of an Embryo Chick of the Third Day (seventy-five hours) viewed sideways as a transparent object : 4 . : =) lai 46. Embryo at the end of the Fourth Day seen as a transparent object. _. ‘ 5 4 : 5 ; 3 : F = Tag XVili LIST OF ILLUSTRATIONS. FIG. PAGE 47- Section through the Lumbar Region of an Embryo at the End of the Fourth Day . : 3 ° * ° . . - ~ 144 48. A. Head of an Embryo Chick of the Fourth Day viewed from below as an opaque object. B. The same seen sideways . - 146 49. Longitudinal Section of the Tail-end of an Embryo Chick at the commencement of the Third Day 5 5 5 . So arts! 50. Longitudinal Section of the Tail-end of an Embryo Chick at the middle of the Third Day . . . . : ° . » 149 51. Section of the intermediate Cell-mass on the Fourth Day - . 165 52. State of Arterial Circulation on the Fifth or Sixth Day . . - 169 53. Diagram of the Venous Circulation at the Commencement of the Fifth Day . 5 . . 5 . : 5 . . eeLO 54. Heart of a Chick on the Fourth Day of Incubation viewed from the Ventral Surface . a . : . . : : 5 GP 55. View from above of the Investing Mass and of the Trabecule on the Fourth Day of Incubation . ° . . . ¢ ae lay 56. A. Head of an Embryo Chick of the Fourth Day viewed from below as an opaque object. B. The same seen sideways. ; 180 57. Head of a Chick at the Sixth Day from below . 5 : LoL 58. Head of a Chick of the Seventh Day from below . f . - 82 59. Section through the Spinal Cord of a Seven Days Chick 5 - 188 60. Two views of the Heart of a Chick upon the Fifth Day of Incuba- tion . : . ° 5 . . ° . : . = 192 61. Heart of a Chick upon the Sixth Day of Incubation, from the Ventral Surface . Sorte ones ia cu Se - = 103 62. Diagram of the Venous Circulation at the Commencement of the Fifth Day . 5 . : a 2 . . ‘. : . 206 63. Diagram of the Venous Circulation during the later days of Incu- bation . . 5 : : . ‘ 5 : . - 208 64. Diagram of the Venous Circulation of the Chick after the com- mencement of Respiration by means of the Lungs . . Ana 63. State of Arterial Circulation on the Fifth or Sixth Day . * . 212 FIG. LIST OF ILLUSTRATIONS. Diagram of the Condition of the Arches of the Aorta towards the Close of Incubation , . ‘ ‘ . Diagram of the Arterial System of the Adult Fowl View from above of the Investing Mass and of the Trabeculz on the Fourth Day of Incubation . . 6 F 2 > ‘ View from below of the Paired Appendages of the Skull of a Fowl on the Fourth Day of Incubation 5 . Side view of the Cartilaginous Cranium of a Fowl on the Seventh Day of Incubation . ‘ < . Embryonic Skull of a Fowl during the Second Week of Incubation (third stage) from below. : : . . 1X PAGE 216 219 226 INTRODUCTION. Every living being passes in the course of its life through a series of changes of shape and structure. These changes may, in their completest form, be considered as constituting a morphological cycle, beginning with the ovum and ending with the ovum again. Among many living beings and especially among verte- brate animals by far by the greater part of the life of the individual is spent in one particular phase, which is not only of longer duration than the rest, but also of much more importance, inasmuch as during it the greater part of the ‘work’ of the living being is done. This is generally spoken of as the adult stage, and in most cases immediately precedes, or is peculiarly associated with, the completion of the morpho- - logical cycle in the appearance of a new ovum. The word embryology may be generally taken to mean the study of the successive morphological phases through which a living being passes from the ovum to the adult stage, or the study of the gradual ‘development’ of the ovum to the adult form ; though, especially among some of the so- called lower forms of life, its meaning must be so extended as to embrace all the morphological phases of an individual life. Embryology is thus a part of and a necessary intro- duction to the wider study of ‘Generation.’ As a matter of E. : 2 ON EMBRYOLOGY. history we find that the study of it sprang out of the various attempts to solve the problems of why and how living beings come into existence. It would be beyond the scope of this work to enter at all fully into any account of the earlier of these inquiries from those of Aristotle downwards; but it may be of some use to point out the chief steps by which in modern times embryology has been established as a distinct branch of knowledge. From the very first, incubated bird’s eggs, and especially hen’s eggs, owing to their abundance at all seasons, and the ease with which they could be examined, became special objects of study. Aristotle examined the growing chick within the egg, and gave the name of punctwm saliens to the ‘bloody palpitating point, which marks the growing heart in the early days of incubation. Since his time all observers have had recourse to the hen’s egg; and though it may be urged that the highly specialised characters of the avian type unfit it for so general a purpose as that of serving as the foundation of embryology, the practical advantages of the bird’s egg over either the mammalian or any other ovum, are so many, that it must always continue to be, as it has been, a chief object of study. From the time of Aristotle down to that of Fabricius of Aquapendente so little progress in real observation of facts had been made, that we find the latter anatomist (De Forma- tione Ovi et Pulli, 1621) describing the chick as being formed out of the chalazze of the white of the egg; a view which lived long afterwards, and whose influence may still be recognized in the names ‘tread’ or ‘treadle’ which the housewife sometimes gives to those portions of thickened albumen. Harvey was the first to clearly establish that the essential part of the hen’s egg, that out of which the embryo pro- INTRODUCTION. 3 ceeded, was the cicatricula. This Fabricius had looked upon as a blemish, a scar left by a broken peduncle. In his Anatomical Exercises on the Generation of Animals (1651), Harvey describes the little cicatricula as expanding under the influence, of incubation into a wider structure, which he calls the eye of the egg; and at the same time sepa- rating into a colliquamentum. In this colliquamentum, according to him, there appears, as the first rudiment of the embryo, the heart or punctwm saliens, together with the blocd-vessels. These gradually gather round them the solid parts of the body of the chick. Harvey clearly was of opinion that the embryo arose, by the successive formation of parts, out of the homogeneous nearly liquid colliquamentum. He was an early advocate of the doctrine of epigenesis. Notwithstanding the weight of Harvey’s authority, the doctrine of epigenesis subsequently gave way to that of evolution, according to which the embryo pre-existed, even though invisible, in the ovum, and the changes which took place during incubation consisted not in a formation of parts, but in a growth, 7.e. in an expansion with concomitant changes, of the already existing germ. Of this theory Malpighi is frequently said to have been the founder. In a limited sense this is true. In his letter to the Royal Society of London, De Formatione Pulli in Ovo (1672), he confesses himself compelled to admit that even in unincu- bated eggs an embryo was present (Quare pulli stamina in ovo pre-existere, altioremque originem nacta esse fateri convenit). Yet he evidently struggled against such a con- clusion, and instead of developing a consistent theory of evolution, left the earliest stages of the embryo as too mysterious to be profitable objects of study, and contented himself with tracing out the events of later days. From his descriptions it is clear that his so-called unincubated eggs 1—2 4 ON EMBRYOLOGY. had under the warmth of summer already made considerable progress in development. The man who first logically worked out a theory of evolution and became its most distinguished and zealous advocate was Haller (Sur la Formation du Cour dans le Poulet, 1758, and Hlementa Physiologie, Liber xxix. 1766). This great anatomist insisted that the embryo existed even in the unincubated egg though in a rudimentary form, and indeed invisible. He supposed that it was a vermiform structure composed of all the essential parts of a full-grown animal in an undeveloped state, and that the effect of incu- bation was to educe or evolve these undeveloped organs into an adult condition. The same views were urged with cha- racteristic extravagance by Bonnet (Considérations sur les corps organisés, 1762). This doctrine of evolution or predilineation, as it was called at the time, was doomed to be overthrown even in Haller’s own day. In an inaugural dissertation entitled Theoria Generationis, published 1759, Casper Frederick Wolff laid the foundations of not only modern Embryology, but modern Histology. He shewed that the cicatricula of the unincubated hen’s egg con- sisted of a congeries of particles (such as we now call cells) all alike, or divisible into groups only, and that anything like distinct rudiments of an embryo were wholly absent. Out of these particles the embryo was built up by means of a series of successive changes (several of which he described in detail, especially in his work on the Formation of the Alimentary Canal, 1768), part being added to part, and parts once formed being modified into fresh parts. Thus the old imperfect theory of evolution was supplanted by a view, which, under the term of epigenesis, was in reality a more complete and truer theory of evolution. Wolff also shewed that all the parts as well of plants as of animals could be conceived of INTRODUCTION. 5 as being arrangements of these particles or cells variously modified, and that all the phenomena of the form and structure of living beings were to be regarded as the results of a variable nutritive energy, to which he gave the name vis essentialis. Haller complained of Wolff, that he had attempted to make a great leap instead of being contented with small on- ward steps. Wolff’s leap proved too great for his time. While his insight into the fundamental doctrines of histology re- mained for the most part without fruit till the next century, so also the way he opened up in embryology was successfully followed by no one for many years after. In 1816 that admirable teacher Dollinger, of Wiirzburg, induced Pander to take up the study of the incubated hen’s ege. We owe to Pander (Dissertatio Inauguralis sistens Mistoriam Metamorphoseos quam Ovum Incubatum prioribus guinque diebus subit, and Beitrdge zur Entwickelungsge- schichte des Hiihnchens im Hie) a clear and excellent descrip- tion of many of the changes which take place during the early days of incubation. It was he who introduced the term blastoderm. He too first drew attention to the distinction of the three layers, serous, mucous, and vascular. But his greatest merit perhaps consisted in the fact of his studies having been the exciting cause of those of Von Baer. Coming to Wiirzburg to study under Dollinger, and finding Pander busily engaged in his embryological work, Von Baer enthusiastically took up the same subject, and thenceforward devoted the greater part of his life to it. Of the results of his labours, which are embodied in his Entwickelungsgeschichte der Thiere, 1828, 1837, this simply may be said. Von Baer found the true line of inquiry already marked out by Wolff. He followed up that line so sedulously and with such success, that nearly all the work which has been done since his day up to the present time, in Vertebrate 6 ON EMBRYOLOGY. Embryology, may be regarded as little more than an ex- tension, with corrections, of his observations. Were it de- sirable to re-publish Von Baer’s work, the corrections and expansions of matters of fact necessary to bring it up to the present time, as the phrase goes, would, with some few exceptions, be of minor importance, though they might be many. ‘The theoretical considerations embodied in his Scholia through which he interprets the morphological sig- nificance of embryological facts are of great and lasting importance, though they need some modifications in order to bring them into harmony with the theory of natural selection. Since Von Baer’s time, the advances made in Vertebrate Em- bryology, through the elaborate work of Remak, the labours of Rathke, Allen Thomson and others, the admirable lectures of Kolliker, and the researches of more recent inquirers, though many and varied, cannot be said to constitute any epochs in the history of the subject, such as that which was marked by Von Baer, and before him by Wolff. We may perhaps make an exception in favour of the discovery by Purkinje, of the germinal vesicle in the fowl’s ovarian ovum (1825). This led to Von Baer’s discovery of the mammalian ovum (1827), which first rendered possible a consistent view of mammalian gene- ration. The study of invertebrate embryology has, on the other hand, during the last few years produced the most striking results. In the following pages we propose to follow in the path thus marked out by the history of the subject. We begin with the chick as being the animal which has been most studied, and the study of which is easiest, and most fruitful for the beginner. The first part accordingly will be devoted to a description of the changes undergone by an incubated hen’s egg, especially during the early days of incubation. We oo? shall endeavour to explain, with such details as are necessary, INTRODUCTION. Ff the manner in which the embryo is formed, and the way in which the rudiments of the most important organs of the chick arise. We shall follow a chronological order, tracing out the changes day by day (or with even shorter periods), during the first few days. We are convinced that this method (adopted by Von Baer) is on the whole the one which most commends itself to the learner. It has of course its disad- vantages ; and in several instances we have found it desirable when describing, at its appropriate date, the most striking phase in the development of an organ, at once to follow up the subsequent history, instead of giving it piecemeal after- wards. But the general advantages of the chronological method, especially when the reading of such a book as this is rendered really useful by an accompanying actual exami- nation of incubated eggs, are so great that they far outweigh the evil of any such slight irregularities. After tracing out the history of the several organs, no farther than is necessary to give a clear idea of the general course of events in each case, we propose to treat the changes and incidents of the latter days of incubation with great brevity, not attempting any special account of avian development, except in the case of the skull. And even this will be treated summarily. The First Part will therefore really be an introduction to the general facts of vertebrate embryology, the chick being taken as an example. In the Second Part we purpose to consider the embryonic histories of other vertebrates, in so far as these differ from that of the bird; and then to treat of the development of special organs in a more complete manner. The Third Part will be devoted to an exposition of the main facts of invertebrate embryology, and to the discussion of general morphological considerations, 8 : ON EMBRYOLOGY. The reader will scarcely fail to notice that the First Part especially is entirely confined to a simple description of observed facts, no attempt whatever being made to interpret their meanings. We have purposely pursued this course, because any interpretation of the facts of the bird’s develop- ment is impossible, or at least illusory, till the history of other animals, vertebrate and invertebrate, has been studied. When all the facts are before him the reader will be in a position to judge of the interpretations offered. PAIL | THE HISTORY OF THE CHICK. CHAPTER I. THE STRUCTURE OF THE HEN’S EGG, AND THE CHANGES WHICH TAKE PLACE UP TO THE BEGINNING OF INCUBATION. 1. In a hen’s egg quite newly laid we meet with the following structures. Most external is the shell (Fig. 1, s.), composed of an organic basis, impregnated with calcic salts. It is sufficiently porous to allow of the interchange of gases between its interior and the external air, and thus the chemical processes of respiration, feeble at first, but gradually increasing in intensity, are carried on during the whole period of incubation. According to Nathusius, Zeztsch. f. Wiss. Zool. Vol. XVIII. p. 225—270, XIX. 322—348, XX. 106—120, XXI. 330—355, the egg-shell of birds consists of an outer thinner and an inner thicker layer. The outer layer varies con- siderably in its consistency in different species. It is soft and pliant in the hen, but in many other birds, as for instance the ostrich, is hard and friable. It is frequently striated both vertically and transversely. Pigment when present is confined to this layer. The inner layer is thicker; and its internal surface is marked with rounded processes more or less separated from one another, whose blunt extremities are sunk into the shell-membrane. The presence of these pro- cesses must be considered as universal amongst birds. Vertical sections shew that this layer is composed of alternating horizontal laminz of transparent and opaque material, the opaque laminz being composed of exceedingly minute par- ticles of an organic nature imbedded in a matrix impregnated with calcic salts. Both layers of the shell are pierced by vertical canals, which are simple in Carinate but ramified in Ratite birds. These canals open freely on the exterior surface and also on the interior surface in the pits between the blunt processes of the inner layer. It is probable that the outer openings of these canals become closed by the presence of moisture, so that when the shell is wet neither air nor water can pass through it. If the shell is dry, air will penetrate easily ; and if the upper layer with the free ends of the tubes be rubbed off, both water and air will pass through it without difficulty. In eggs with coloured shells the colouring matter frequently passes into the canals. 12 THE HEN’S EGG. [CHAP. 2. Lining the shell, is the shell-membrane, which is double, being made up of two layers; an outer thicker (Fig. 1, s. m.), and an inner thinner one (¢. s. m.). Both of these layers consist of several laminz of felted fibres of various sizes, intermediate in nature between connective and elastic fibres. Fic. 1. DiaGRAMMATIC SECTION OF AN UNINCUBATED Fow.’s Eca (modified from Allen Thomson). bl. blastoderm. w. y. white yolk. This consists of a central flask-shaped mass and a number of layers concentrically arranged around this. y. y. yellow yolk. v. ¢. vitelline membrane. «x. layer of more fluid albumen immediately surrounding the yolk. w. albumen consisting of alternate denser and more fluid layers. ch.l. chalaza. a. ch. air-chamber at the broad end of the egg. This chamber is merely a space left between the two layers of the shell-membrane. ¢@. s. m. internal layer of shell- membrane. s. m. external layer of shell-membrane. s. shell. Over the greater part of the egg the two layers of the shell-membrane remain permanently in close apposition to each other; but at the broad end they tend to separate, and thus to develope between them a space into which air finds its way. This air-chamber, as it is called, is not to be found’ in perfectly fresh eggs, but makes its appearance in 1] THE WHITE OF THE EGG. 13 eggs which have been kept for some time, whether incubated or not, and gradually increases in size, as the white of the ege shrinks in bulk by evaporation. 3. Immediately beneath the shell-membrane is the white of the egg or albumen (Fig. 1, w.), which is, chemically speaking, a mixture of various forms of proteid material, with fatty, extractive, and saline bodies. Its average composition may be taken as 12°0 p. c. proteid matter, I°5 p. c. fat and extractives, *g p.c. saline matter, chiefly sodic and potassic chlorides, with phos- phates and sulphates, 86:0 p. c. water. The white of the egg when boiled shews in section alternate concentric layers of a transparent and of a finely granular opaque material. In the natural condition, the layers corresponding to these opaque layers are composed of more fluid albumen, while those corresponding to the transparent layers are less fluid, and consist of networks of fibres, con- taining fluid in their meshes. The outer part of the white, especially in eggs which are not perfectly fresh, is more fluid than that nearer the yolk. The innermost layer, however, immediately surrounding the yolk (Fig. 1, x.), is of the more fluid finely granular kind. In eggs which have been hardened a spiral arrangement of the white may be observed, and it is possible to tear off laminz in a spiral direction from left to right, from the broad to the narrow end of the egg. Two twisted cords called the Chalaze (Fig. 1, ch. l.), com- posed of coiled membranous layers of the less fluid albumen, run from the two extremities of the egg to the opposite portions of the yolk. Their inner extremities expand and merge into the layer of denser albumen surrounding the fluid layer next the yolk. Their outer extremities are free, and do not quite reach the outer layer of the white. Thus they cannot serve to suspend the yolk, although they may help to keep it in position, by acting as elastic pads. The interior of each chalaza presents the appearance of a suc- cession of opaque white knots; hence the name chalaze, grandines (hailstones). 4. The yolk is enclosed in the vitelline membrane (Fig. 1, v. t.), a transparent somewhat elastic membrane easily 14 THE HEN’S EGG. [CHAP. thrown into creases and wrinkles. It might almost be called structureless, but under a high power a fine fibrillation is visible, and a transverse section has a dotted or punctated appearance; it is probably therefore composed of fibres. Its affinities are with elastic rather than connective tissue. The vitelline membrane of most vertebrates is perforated by fine pores. These are largest in osseous fishes and much finer in mammals; they have not been found in the vitelline membrane of birds. 5. The whole space within the vitelline membrane is occupied by the yolk. To the naked eye this appears toler- ably uniform throughout, except at one particular point of its surface, at which may be seen, lying immediately under the vitelline membrane, a small white disc, about 4 mm. in diameter. This is the blastoderm, or cicatricula. A tolerably typical cicatricula in a fecundated egg will shew an outer white rim of some little breadth, and within that a circular transparent area, in the centre of which, again, there is an opacity, varying in appearance, sometimes uniform, and sometimes dotted. The disc is always found to be uppermost whatever be the position of the egg, provided there is no restraint to the rotation of the yolk. The explanation of this is to be sought for in the lighter specific gravity of that portion of the yolk which is in the neighbourhood of the disc, and the phenomenon is not in any way due to the action of the chalaze. A section of the yolk of a hard-boiled egg will shew that it is not perfectly uniform throughout, but that there is a portion of it having the form of a flask, with a funnel- shaped neck, which, when the egg is boiled, does not become so solid as the rest of the yolk, but remains more or less fluid. The expanded neck of this flask-shaped space is situated immediately underneath the disc, while its bulbous enlarge- ment is about the middle of the yolk. We shall return to it directly. 6. The great mass of the yolk is composed’ of what is known as the yellow yolk (Fig. 1, y. y.). This consists of spheres (Fig. 2, A.) of from 25 to 100u’ in diameter, never containing a nucleus, but filled with numerous minute highly refractive granules; these spheres are very delicate and easily 4 4= "OO! mm, I.] THE WHITE YOLK. 15 destroyed by crushing, When boiled or otherwise hardened an situ, they assume a polyhedral form, from mutual pressure. The granules they contain seem to be of an albuminous nature, as they are insoluble in ether or alcohol. Chemically speaking the yolk is characterized by the presence in large quantities of a proteid matter, having many affinities with globulin, and called vitellin, This exists in peculiar association with the remarkable body Lecithin. (Compare Hoppe-Seyler, Hdb. Phys. Chem. Anal.) Other fatty bodies, colouring matters, extractives (and, according to Dareste, starch in small quan- tities), &c. are also present. Miescher (Hoppe-Seyler, Chem. Untersuch. p. 502) states that a considerable quantity of nucleén may be obtained from the yolk, probably from the spherules of the white yolk. Fig. 2. A, Yellow yolk-sphere filled with fine granules. The outline of the sphere has been rendered too bold. White yolk-spheres and spherules of various sizes and presenting different appearances. (It is very difficult in a woodcut to give a satisfactory repre- sentation of these peculiar structures.) 7. The yellow yolk thus forming the great mass of the entire yolk is clothed externally by a thin layer of a different material, known as the white yolk, which at the edge of the blastoderm passes underneath the disc, and becoming thicker at this spot forms, as it were, a bed on which the blastoderm rests. Immediately under the middle of the blastoderm this bed of white yolk is connected, by a narrow neck, with a central mass of similar material, lying in the middle of the yolk (Fig. 1, w. y.). When boiled, or otherwise hardened, the white yolk does not become so sclid as the yellow yolk; hence the appearances to be seen in sections of the hardened yolk. The upper expanded extremity of this neck of white yolk is generally known as the “nucleus of Pander.” Concentric to the outer enveloping layer of white yolk there are within the yolk other inner layers of the same substance, which cause sections of the hardened yolk to 16 THE HEN’S EGG. [CHAP. appear to be composed of alternate concentric thicker lamine of darker (yellow) yolk, and thinner lamine of lighter (white) yolk (Fig. 1, w, y.). 8. The microscopical characters of the white yolk are very different from those of the yellow yolk. It is composed of spheres (Fig. 2, B.) for the most part smaller than those of the yellow yolk (4u—75y), with a highly refractive nucleus- like body often as small as lw in the interior of each; and also of larger spheres, each of which contains a number of spherules, similar to the smaller spheres ; these latter appear- ing to have passed into the larger spheres, by a process of inclusion. There has been a considerable amount of controversy as to whether these elements possess a membrane ; there is little doubt however that there is no membrane present. It has also been disputed as to whether they should be considered as true cells or not. If by definition a cell must contain a nucleus, they can hardly be considered as such, since the characters of the highly refractive bodies con- tained in them have nothing in common with nuclei. We shall give later on reasons for thinking that they may however, as a result of incubation, become veritable cells. Another feature of the white yolk, according to His, is that in the region of the blastoderm it contains numerous large vacuoles filled with fluid; they are sufficiently large to be seen with the naked eye, but do not seem to be present in the ripe ovarian ovum. 9. It is now necessary to return to the blastoderm. In this, as we have already said, the naked eye can distinguish an opaque white rim surrounding a more transparent central area, in the middle of which again is a white spot of variable appearance. In an unfecundated cicatricula the white disc is simply marked with a number of irregular clear spaces, there being no proper division into a transparent centre and an opaque rim. The opaque rim is the commencement of what we shall henceforward speak of as the area opaca; the central trans- parent portion is in the same way the beginning of the area pellucida. At this stage the distinction between these two areas depends entirely on the disposition of the white yolk beneath them, for the blastoderm when lifted up from the white yolk on which it rests appears uniform throughout. In the part corresponding to the area opaca the blastoderm rests immediately on the white yolk, which here forms a 1] THE BLASTODERM. 17 somewhat raised ring, often spoken of as the germinal wall ; underneath the area pellucida is a shallow space containing a nearly clear fluid, to the presence of which the central transparency seems to be due. The white spot in the middle of the area pellucida appears to be the nucleus of Pander shining through. Vertical sections of the blastoderm shew that it is formed of two layers. The upper of these two layers is com- posed, see Fig. 3, ep, of a single layer of cells, with their long axes arranged vertically, adhering together so as to form a distinct membrane, the edge of which rests upon the white yolk. After staining with silver nitrate, this membrane viewed from above shews a mosaic of uniform polygonal cells. Each cell is composed of granular protoplasm filled with highly refractive globules; in most of the cells an oval nucleus may be distinguished, and is most probably present in all. They are of a uniform size (about 9) over the opaque and the pellucid areas. The under layer (Fig. 3, 7), is composed of cells which vary considerably in diameter ; but even the smaller cells of this layer are larger than the cells of the upper layer. They are spherical, and so filled with granules and highly refractive globules, that a nucleus can rarely be seen in them: in the larger cells these globules contain a highly refractive body very similar to that present in the white yolk spheres, from the smaller kinds of which indeed they are scarcely distinguishable. The cells of this layer do not form a distinct membrane like the cells of the upper layer, but lie as a somewhat irregular network of cells between the upper layer and the bed of white yolk on which the blastoderm rests. The lowest are generally the largest; in addition we find a few still larger cells generally separated by a small interval from the remainder of the cells of the lower layer, and resting directly upon the white yolk (Fig. 3, b). These are frequently spoken of as formative cells; they are however similar in character and indeed connected by gradations with the larger cells of the lower layer. Their mode of formation during segmentation will be subsequently de- scribed. E. 2 18 THE HEN’S EGG. [CHAP. SECTION OF A BLASTODERM OF A Fow.’s EaG aT THE COMMENCEMENT OF INCUBATION. The thin but complete upper layer ep composed of columnar cells rests on the incomplete lower layer /, composed of larger and more granular bodies. The lower layer is thicker in some places than in others, and is especially thick at the periphery. The line below the under layer marks the upper surface of the white yolk, The larger so-called formative cells are seen at b, lying on the white yolk. The figure dves not take in quite the whole breadth of the blastoderm; but the reader must understand that both to the right hand and to the left ep is continued farther than J, so that at the extreme edge it rests directly on the white yolk. Over nearly the whole of the blastoderm the upper layer rests on the under layer. At the circumference however the upper layer stretches for a short distance beyond the under layer, and here consequently rests directly on the white yolk, and forms that part of the blastoderm known as the area opaca. 10. To recapitulate: —In the normal unincubated hen’s egg we recognize the blastoderm, consisting of a complete upper layer of smaller nucleated granular cells and a more or less incomplete under layer of larger cells, filled with larger granules; in these lower cells nuclei are rarely visible. The thin flat disc so formed rests, at the uppermost part of the entire yolk, on a bed of white yolk so disposed as to give rise to the appearance in the blastodermic dise it- self of an area opaca and an area pellucida. The great mass of the entire yolk consists of the so-called yellow yolk composed of gra- nular spheres. The white yolk is composed of smaller spheres of peculiar structure, and exists, in small part, as a thin coating around, and as thin concentric lamine in the sub- stance of the yellow yolk, but chiefly in the form of a flask-shaped mass in the interior of the yolk, the upper somewhat expanded top of the neck 1.] THE OVARIAN OVUM, 19 of which forms the bed on which the blastoderm rests, The whole yolk is invested with the vitellme membrane, this again with the white; and the whole is covered with two shell-membranes and a shell. 11. Such an egg has however undergone most important changes while still within the body of the hen; and in order to understand the nature of the structures which have just been described, it will be necessary to trace briefly the history of the egg from the stage when it exists as a so-called ripe ovarian ovum in the ovary of a hen up to the time when it is laid. If one of the largest capsules of the ovary of a hen which is laying regularly be opened, it will be found to contain a nearly spherical (or more correctly, ellipsoidal with but slightly unequal axes) yellow body enclosed in a delicate membrane. ‘I'his is the ovarian ovum or egg. Examined with care the ovum, which is tolerably uniform in appearance, will be found to be marked at one spot (generally facing the stalk of the capsule and forming the pole of the shorter axis of the ovum) by a small disc differing in appearance from the rest of the ovum. This disc is known as the germinal disc or discus proligerus. It consists of a lenticular mass of SECTION THROUGH THE GERMINAL Disc OF THE RIPE OVARIAN OvuUM OF A FOWL WHILE YET ENCLOSED IN ITS CAPSULE. a. Connective-tissue capsule of the ovum. Ob. epithelium of the capsule, at the surface of which nearest the ovum lies the vitelline membrane. c. granular material of the germinal disc, which becomes converted into the blastoderm. (This is not very well represented in the woodcut. In sections which have been hardened in chromic acid it consists of fine granules.) w. y, white yolk, which passes insensibly into the fine granular material of the . dise. , germinal vesicle enclosed in a distinct membrane, but shrivelled up by the action of the chromic acid. The material enclosed in the membrane of the vesicle is in the hardened specimens finely granular. y, space originally completely filled up by the germinal vesicle, before the latter was shrivelled up by the action of the chromic acid. 20 THE HEN’S EGG. [ CHAP. protoplasm (Fig. 4, c), imbedded in which is a highly refrac- tive globular or ellipsoidal body (Fig. 4, x), about 310 in diameter, called the germinal vesicle, in the interior of which again is a small body, the germinal spot. The rest of the ovum is known as the yolk. This consists of two elements, the white yolk- and the yellow yolk-spheres, which are distributed respectively very much in the same way as in the laid egg, the yellow yolk forming the mass of the ovum, and the white yolk being gathered underneath and around the disc (Fig. 4, w. y), and also forming a flask- shaped mass in the interior of the ovum. The delicate membrane surrounding the whole is the vitelline membrane. Oellacher’s (Untersuchung tiber die Furchung und Blatterbildung in Hiih- mereie. Studien aus dem Institute fiir experimentale pathologie in Wien aus dem Jahre 1869, pt. 1) account of the ovarian ovum differs considerably from that given above. He finds in the neighbourhood of the blastoderm a finely granular material, within which lies a body appearing circular when viewed trom above, but having in section a somewhat quadrilateral shape ; its side-wails, however, are curved, with their convexity turned inwards. At the bottom of it lies an oval cavity with doubly contoured walls, and at its upper surface placed somewhat excentrically a semicircular space filled with clear material. Oellacher believes that the quadrilateral body which he thus describes is the germinal vesicle which has commenced to undergo a retrogressive meta- morphosis. For the further stages in the metamorphosis, and for further par- ticulars, vide Section 13. The circular hole beneath the vesicle is probably merely filled with fluid and is due to the contractions of the germ. 12. When the ovarian ovum is ripe and about to be dis- charged from the ovary, its capsule is clasped by the dilated termination of the oviduct. The capsule then bursts, and the ovum escapes into the oviduct, its longer axis corre- sponding with the long axis of the oviduct, the germinal disc therefore being to one side. At the time of the bursting of the capsule the germinal vesicle disappears. In describing the changes which take place in the oviduct, it will be convenient, following the order previously adopted, to treat first of all of the formation of the accessory parts of the egg. These are secreted by the glandular walls of the oviduct. This organ therefore requires some descrip- tion. It may be said to consist of four parts ;—Ist. The ‘dilated proximal extremity. 2nd. A long tubular portion, opening by a narrow neck or isthmus into the 3rd portion, which is much dilated, and has been called the uterus; the 4th part is somewhat narrow; and leads from the uterus I] DESCENT OF THE OVUM. 21 into the cloaca. The whole of the mucous membrane lining the oviduct is largely ciliated. The accessory parts of the egg are entirely formed in the 2nd and 3rd portions. The layer of albumen which imme- diately surrounds the yolk is first deposited; the chalaze are next formed. Their spiral character and the less distinctly marked spiral arrangement of the whole albumen is brought about by the motion of the egg along the spiral ridges into which the interior of the second or tubular portion of the oviduct is thrown. The’ spirals of the two chalaze are in different directions. This is probably produced by their peripheral ends remaining fixed while the yolk to which their central ends are attached is caused to rotate by the contractions of the oviduct. During the formation of the chalaze the rest of the albumen is also deposited; and finally the shell-membrane is formed in the narrow neck of the 2nd portion, by the fibrillation of the most external layer of albumen. The egg passes through the 2nd portion in little more than 3 hours. In the 3rd 3 portion the shell is formed. The mucous membrane of this part is raised into numerous flattened folds, ike large villi, containing follicu- lar glands. From these a thick white fluid is poured out, which soon forms a kind of covering to the egg, in which the inorganic particles are deposited. In this portion of the oviduct the egg remains from 12 to 18 hours, during which time the shell acquires its normal consistency. At the time of laying it is expelled from the uterus by violent muscular contractions, and passes with its narrow end downwards along the remainder of the oviduct, to reach the exterior. 13. We have now to trace out the changes which take place in the germinal disc, during the passage of the egg down the oviduct. By the time when the egg becomes clasped by the expanded extremity of the oviduct the germinal vesicle has, according to Oellacier (loc cit. and also Archi. fiir Micr. Anat. Vol. vil. 1872. p. 18), undergone still further retrogressive changes. It has now become very much flattened and closely applied to the vitelline membrane. Both this and former stages, if we may judge from the analogy of osseous fishes, are preparatory to the whole germinal vesicle being bodily ejected from the germinal disc. For further particulars vide Oellacher, Archiv. fiir Micr. Anat. Vol. vu. pp. 1—26. Impregnation occurs in the upper portion of the oviduct ; the spermatozoa being found actively moving in a fluid which is there contained. 22 THE HEN'S EGG. [CHAP. It is not certain whether impregnation takes place previous to the deposition of the albumen, or whetuer the spermatozoa bore their way through the albumen. The former would appear to be the more probable view, though the fact that Oellacher has found spermatozoa in the albumen, speaks in favour of their being involved in the depositing albumen, and so being brought in contact with tue blastoderm. According to Coste, Histoire du développement des corps oryanizés, the access of the cock to the hen once in seven days is sufficient. We have no positive evidence that the spermatozoa make their way through the vitelline membrane and so gain access to the germinal disc; but, as will be seen in a later part of this work, analogy renders such an event probable. 14. At about the time when the shell is being formed round the egg, the germinal disc undergoes a remarkable change, known as segmentation. We shall have occasion to treat more fully of the nature of segmentation when we come to consider the amphibian ovum in which the various steps of the process may be more easily and satisfactorily traced. Meanwhile, inasmuch as the segmentation of the SURFACE VIEWS OF THE EARLY STAGES OF THE SEGMENTATION IN A Fow.’s Eee. (After Coste.) A represents the earliest stage. The first furrow (d) has begun to make its appearance in the centre of the germinal disc, whose periphery is marked by the line a. In B, the first furrow is completed right across the disc, and a second similar furrow at nearly right angles to the first has appeared. The disc thus becomes divided somewhat irregularly into quadrants by four (half) furrows. In a later stage (C’) the meridian furrows b have increased in number, from four, as in B, to nine, and cross furrows have also made their appearance. The dise is thus cut up into small central (c) and larger peripheral (d) segments. Several new cross furrows are seen just beginning, as ex. gr. close to the end of the line of reference d. t 1] SEGMENTATION. 23 germinal disc of a hen’s egg differs materially from the segmentation of the entire ovum of an amphibian, the former may briefly be described here. Viewed from above, a furrow is seen to make its appear- ance, running across the germinal disc and dividing it into two halves (Fig. 5, A). This primary furrow is succeeded by a second at right angles to itself, The surface thus becomes divided into four segments or quadrants (Fig. 5, B). Each of these is again bisected by radiating furrows, and thus the number of segments is increased from four to eight (it may be seven or nine). The central portion of each segment is then, by a cross furrow, cut off from the peripheral portion, giving rise to the appearance of a number of central smaller segments, surrounded by more external elongated segments (Fig. 5, C). Division of the segments now proceeds rapidly by means of furrows running apparently in all directions. And it is RGN: SURFACE VIEW OF THE GERMINAL Disc oF A HeN’s Ecc DURING THE LATER Sraces or SEGMENTATION. (Chromic Acid Preparation.) At cin the centre of the disc the segmentation masses are very small and numerous. At b, nearer the edge, they are larger and fewer; while those at the extreme margin a are largest and fewest of all. It will be noticed that the radiating furrows marking off the segments a do not as yet reach to the extreme margin e of the disc. P The drawing is completed in one quadrant only; it will of course be under- stood that the whole circle ought to be filled up in a precisely similar manner. 24 THE HEN’S EGG. [CHAP. important to note that the central segments divide more rapidly than the peripheral, and consequently become at once smaller and more numerous (Fig. 6). Meanwhile sections of the hardened blastoderm teach us that segmentation is not confined to the surface, but extends through the mass of the blastoderm; they shew us moreover that division takes place by means of not only vertical, but also horizontal furrows, ¢.e. furrows parallel to the surface of the dise (Fig. 7). Fic. 7. SECTION OF THE GERMINAL Disco oF A FowL DURING THE LATER STAGES OF SEGMENTATION. The section, which represents rather more than half the breadth of the blastoderm (the middle line being shewn at c), shews that the upper and central parts of the dise segment faster than those below and towards the periphery. At the periphery the segments are still very large. One of the larger segments is shewn at a. In the majority of segments a nucleus can be seen; and it seems probable that a nucleus is present in them all. Most of the segments are filled with highly refracting spherules, but these are more numerous in some cells (especially the larger cells near the yolk) than in others. In the central part of the blastoderm the upper cells have commenced to form a distinct layer. No segmentation-cavity is present. a. large peripheral cell. 6. larger cells of the lower parts of the blastoderm. ce. middle line of blastoderm. e. edge of the blastoderm adjoining the white yolk, w. white yolk. In this way, by repeated division, or segmentation, the original germinal disc is cut up into a large number of small rounded masses of protoplasm, which are smallest in the centre, and increase in size towards the periphery. The segments lying uppermost are moreover smaller than those beneath, and thus the establishment of the two layers of the blastoderm is foreshadowed. 1] SEGMENTATION. 25 According to Oellacher, Studien aus dem Ins. f. Exper. Pathol. Vien. 1869, p. I, sections taken through the centre of the germinal disc at the beginning of segmentation shew a somewhat uneven vertical furrow, ending below in a small triangular space, where it joins a nearly horizontal furrow which meets the surface of the egg at some little distance on either side of the vertical furrow. It seems certain that these first-formed furrows do not include the whole of the germinal disc, whose limits at this stage are however uncertain. In the later stages of segmentation not only do the first-formed segments become further divided, but segmentation also extends into the remainder of the germinal disc. Goette, Archiv. Micr. Anat. x. 145, indeed maintains that segmentation (at a later period) even involves material which is undoubtedly white yolk. He describes nuclei as making their appearance in the upper surface of the bed of white yolk, and the substance round them as rising up in the form of papille, which are subsequently constricted off and set free as supplementary segmentation masses. It is these, according to him, which give rise to the formative cells spoken of in the next paragraph. He states that they continue to be formed long after the commencement.of incubation. We shall return to this subject, when we come to discuss more fully the nature of the process of segmentation, in describing the ova of other classes of vertebrates. Between the segmented germinal disc, which we may now call the blastoderm, and the bed of white yolk on which it rests, a space containing fluid makes its appearance. This, gradually increasing in all dimensions, may be called the segmentation-cavity. 15. As development proceeds, segmentation reaches its limits in the centre, but continues at the periphery, and thus eventually the masses at the periphery become of the same size as those in the centre. The distinction however between an upper and a lower layer becomes more and more obvious. The masses of the upper layer arrange themselves, side by side, with their long axes vertical; their nuclei become very distinct. In fact they form a membrane of columnar nucleated cells. The masses of the lower layer, remaining larger than those of the upper layer, continue markedly granular and round, and form rather a close irregular network than a distinct membrane. In them nuclei are either wholly absent or at least not readily visible. It seems more probable that the nucleus is hidden than that it is really absent. In the earliest stages of segmentation which we have examined when the segments were still few in number, a very large proportion of both great and small segments contained large well-formed nuclei. These nucleated segments, which were found in both the superficial and deeper portions of the disc, were invariably those in which the granules were for some reason or other few and fine; in fact, wherever the granules were not sufficiently numerous to render the body of the segment too opaque, there a nucleus could be detected. 26 . THE HEN’S EGG. [CHAP. I. We were thus led to the conclusion that a nucleus really existed in all. It is of course quite possible that the clearer nucleated masses eventually come to the surface and leave the more granular and opaque masses to form the lower layer; but it is much more likely that they do not, and that the granular con- dition of the cells of the lower layer of the fully formed blastoderm is on the one hand the result of their being in immediate contact with the excessively granular white yolk-cells, and on the other the cause of their nuclei not being seen. We have a somewhat analogous case in the invisibility of the nucleus in the early stages of the amphibian blood-corpuscle. At the time when the segmentation- -spheres in the centre are smaller than those at the periphery, and those above are also smaller than those below, a few large spherical raasses begin to separate from the remainder (or to arise by a continued process of segmentation from the bed of white yolk), and to rest directly on the white yolk, at the bottom of the shallow segmentation-cavity. They contain either numerous small nucleated spherules, or fine granules ; the spherules precisely resembling the smaller spheres of white yolk. These loose spherical masses are the formative cells already spoken of. Thus the original germinal disc of the ovarian ovum, its germinal vesicle having disappeared, becomes, by the process of segmentation, converted into a blastoderm such as is met with in the egg when laid, into an upper layer of columnar nucleated cells, and into a lower layer of irregularly disposed rounded masses which have not yet definitely ac- quired the character of cells, accompanied by a few stray “formative” cells lying loose in the segmentation-cavity. CHAPTER II. A BRIEF SUMMARY OF THE WHOLE HISTORY OF INCUBATION. 1. Srep by step the simple two-layered blastoderm de- scribed in the previous chapter is converted into the complex organism of the chick. The details of the many changes through which this end is reached will perhaps be rendered more intelligible if we prefix to the special history of them a brief summary of the general course of events from the beginning to the end of incubation. In the first place, it is to be borne in mind that the embryo itself is formed in the area pellucida, and in the area pellucida alone. The area opaca in no part enters directly into the body of the chick; the structures to which it gives rise are to be regarded as appendages, which sooner or later disappear. 2. The blastoderm at starting consists of two layers. Very soon a third layer makes its appearance between the other two. These three layers, the establishment of which is a fact of fundamental importance in the history of the ' embryo, are called respectively the upper, middle and lower layers, or epiblast, mesoblast and hypoblast. This triple division corresponds roughly, though not exactly, to the old division into serous, vascular and mucous layers. 3. The blastoderm which at first,as we have seen, lies like a watch-glass over the segmentation-cavity, its margin resting on the circular germinal wall of white yolk, spreads, as a thin circular sheet, over the yolk, immediately under the vitelline membrane. Increasing uniformly at all points of its circumference, the blastodermic expansion covers more and more of the yolk, and at last, reaching its opposite pole 28 PRELIMINARY ACCOUNT. [CHAP. completely envelopes it. Thus the whole yolk, instead of being enclosed as formerly by the vitelle membrane alone, comes to be also enclosed in a bag formed by the blastoderm. It is not however until quite a late period that the complete closing in at the opposite pole takes place, so that the extension of the blastoderm must be thought of as going on during nearly the whole period of incubation. Both the area opaca and the area pellucida share in this enlargement, but the area opaca increases much more rapidly than the area pellucida, and plays the principal part in encompassing the yolk. 4. The mesoblast, in that part of the area opaca which is nearest to the area pellucida, becomes the seat of peculiar changes, which result in the formation of blood-vessels. Hence this part of the area opaca is called the vascular area. 5. The embryo itself may be said to be formed by a folding off of the central portion of the area pellucida from the rest of the blastoderm. At first the area pellucida is quite flat, or, inasmuch as it forms par't of the circumference of the yolk, slightly but uniformly curved. Very soon, how- ever, there appears at a certain spot a semilunar groove, at first small, but gradually increasing in depth and extent ; this groove, which is represented in section in the diagram (Fig. 8, A), breaks the uniformity of the level of the area pellucida. It may be spoken of as a tucking in of a small portion of the blastoderm in the form of a crescent. When viewed from above, it presents itself as a curved line (the hinder of the two concentric curved lines in front of A in Fig 11), which marks the hind margin of the groove, the depression itself being hidden. In a vertical longitudinal section carried through the middle line, we may recognize the following parts (Fig. 8, A, or on a larger scale Fig. 9, which also shews details which need not be considered now). Beginning at what will become the posterior extremity of the embryo, (the left-hand side of the figure in each case), and following the surface of the blastoderm forwards (to the right in the figures), the level is maintained for some distance, and then there is a sudden descent, the blastoderm bending round and pursuing a precisely opposite direction to its previous one, running backwards instead of forwards, for some distance, It soon however turns round again, and once more running I1.] THE HEAD-FOLD. 29 Fic. 8. Po eae aa Pe ue Vs: aie af _. B i att D Fig. 8, A to N forms a series of purely diagrammatic representations in- troduced to facilitate the comprehension of the manner in which the body of the embryo is formed, and of the various relations of the yolk-sac, amnion and allantois. In all vt is the vitelline membrane, placed, for convenience sake, at some distance from its contents, and represented as persisting in the later stages; in the actual egg it is in direct contact with the blastoderm (or yolk), and early ceases to have a separate existence. In all e indicates the embryo, pp the general pleuro-peritoneal space, af the folds of the amnion, a the amnion proper, ae or ac the cavity holding the liquor amnii, ai the allantois, a’ the alimentary canal, y or ys the yolk or yolk-sac. A, which may be considered as a vertical section taken longitudinally along the axis of the embryo, represents the relations of the parts of the egg at the time of the first appearance of the head-fold, seen on the right-hand side of the blastoderm e. The blastoderm is spreading both behind (to the left hand in the figure), and in front (to right hand) of the head-fold, its limits being indicated by the shading and thickening for a certain distance of the margin of the yolk y. As yet there is no fold on the left side of e correspond- ing to the head-fold on the right, while therefore the front limit of the embryo, as distinguished from the blastoderm, is marked out by the head-fold, there is at present no tail-fold, and therefore no hind limit. B is a vertical transverse section of the same period drawn for convenience sake on a larger scale (it should have been made flatter and less curved). It shews that the blastoderm (vertically shaded) is extending laterally as well as fore and aft, in fact in all directions; but there are no lateral folds, and there- 30 PRELIMINARY ACCOUNT, [CHAP. fore no lateral limits to the body of the embryo as distinguished from the blastcderm. Incidentally it shews the formation of the medullary groove by the rising up of the laminz dorsales. Beneath the section of the groove is seen the rudiment of the notochord. On either side a line indicates the cleavage of the mesoblast just commencing. This cleavage, it will be seen, does not exist in the more central parts of the embryo. In C, which represents a vertical longitudinal section of later date, both head-fold (on the right) and tail-fold (on the left) have advanced considerably. The alimentary canal is therefore closed in both, in front and behind, but is in middle still widely open to the yolk y below. Though the axial parts of the embryo have become thickened by growth, the body-walls are still thin; in them however is seen the cleavage of the mesoblast, and the divergence of the somatopleure and splanchnopleure. The splanchnopleure both at the head and at the tail is folded in to a greater extent than the somatopleure, and eo 6: i _— be K 2 11.] THE BODY-FOLDS. 31 forms the still wide splanchnic stalk. At the end of the stalk, which is as yet short, it bends outwards again and spreads over the top of the yolk. The somatopleure folded in less than the splanchnopleure to form the wider somatic stalk, sooner bends round and-runs outwards again. At a little distance from both the head and the tail it is raised up into a fold, af, af, that in front of the head being the highest. These are the amniotic foids. Descending from either fold, it speedily joins the splanchnopleure again, and the two, once more united into an uncleft membrane, extend some way downwards over the yolk, the limit or outer margin of the opaque area not being shewn. All the space between the somatopleure and the splanchnopleure is shaded with dots, pp. Close to the body this space may be called the pleuroperitoneal cavity; but outside the body it runs up into either amniotic told, and alsu extends some little way over the yolk. D represents the tail-end at about the same stage on a more enlarged scale, in order to illustrate the position of the allantois al (which was for the sake of simplicity omitted in @), shewn as a bud from the splanchnopleure, stretching downwards into the pleuroperitoneal cavity pp. The dotted area representing as before the whole space between the splanchnopleure and the somatopleure, it is evident that a way is open for tne allantois to extend from its present position into the space between the two walls of the amniotic fold af. E, also a longitudinal section, represents a stage still farther advanced. Both splanchnic and somatic stalks are much narrowed, especially the former, the cavity of the alimentary canal being now connected with the cavity of the yolk by a mere canal. ‘Tne folds of the amnion are spreading over the top of the embryo and nearly meet. Each fold consists of two walls or limbs, the space between which (dotted) is as before merely a part of the space between the somatopleure and splanchnopleure. Between these arched amniotic folds and the body of the embryo is a space not as yet entirely closed in. F represents on a different scale a transverse section of # taken through the middle of the splanchnic stalk. The black ring in the body of the embryo shews the position of the neural canal, below which is a black spot, marking the notochord. On either side of the notochord the divergence of somato- pleure and splanchnopleure is obvious. The splanchnopleure, more or less thickened, is somewhat bent in towards the middie line, but the two sides do not unite, the alimentary canal being as yet open below at this spot; after converging somewhat they diverge again and run outwards over the yolk. The somatopleure folded in to some extent at first to form the body-walls (which are here made too thick), soun bends outwards again, and almost im- mediately is raised up into the lateral folds of tle amnion uf. ‘The conti- buity of the pleuroperitoneal cavity within the body with the interior of the amniotic fold outside the body is evident; both cavities are dotted. It will of course be understood that this is a purely diagrammatic: representation, the various cavities, &c., beiny exaggerated in order to shew their relations more clearly. @ which corresponds to D at a later stage, is introduced to shew the . manner in which the allantois, now a distinctly hollow body, whose cavity is “continuous with that of the alimentary canal, becomes directed towards the amniotic fold. In # a longitudinal, and J a transverse section of later date, great changes have taken place. The several folds of the amnion have met and coalesced above the body of the embryo. The inner limbs of the several folds have united into a single membrane (a), which encloses a space (ae) round the embryo. *This membrane a is the amnion proper, and the cavity within it, z.e. between it and the embryo, is the cavity of the amnion containing the liquor amnii. 32 PRELIMINARY ACCOUNT. [CHAP. It will be seen that the amnion a now forms in every direction the termina- tion of the somatopleure; the peripheral portions of the somatopleure, the united outer or descending limbs or walls of the folds af in C, D, F, G having been cut adrift, and now forming an independent continuous membrane, the chorion, immediately underneath the vitelline membrane. In J the splanchnopleure is seen converging to complete the closure of the alimentary canal a’, even at the stalk (elsewhere the canal has of course long been closed in), and then spreading outwards as usual over the yolk. The point at which it unites with the somatopleure, marking the extreme limit of the cleavage of the mesoblast, is now much nearer the lower pole of the diminished yolk. As a result of these several changes, a great increase in the dotted space has taken place. It is now possible to pass from the actual peritoneal cavity within the body, on the one hand round a great portion of the circumference of the yolk, and on the other hand above the amnion a, in the space between it and the chorion. Into this space the allantois is seen spreading in K at al. N ys In Z the splanchnopleure has completely invested the yolk-sac, but at the lower pole of the yolk is still continuous with that peripheral remnant of the somatopleure now called the chorion. In other words, cleavage of the meso- blast has been carried all round the yolk (ys) except at the very lower pole. In & the cleavage has been carried through the pole itself; the peripheral portion of the splanchnopleure forms a complete investment of the yolk quite unconnected with the peripheral portion of the somatopleure, which now exists as a continuous membrane lining the interior of the shell. The yolk sac (ys) is therefore quite loose in the pleuroperitoneal cavity, being connected only with the alimentary canal (a’) by a solid pedicle. me” THE HEAD-FOLD. 33 Lastly, in WV the yolk sac (ys) is shewn being withdrawn into the cavity of the body of the embryo. The allantois is as before, for the sake of simplicity, omitted ; its pedicle would of course lie by the side of ys in the somatic stalk marked by the usual dotted shading. It may be repeated that the above are diagrams, the various spaces being shewn distended, whereas in many of them in the actual egg the walls have collapsed, and are in near juxtaposition. Fig. 9. D1isAcGRAMMATIO LONGITUDINAL SECTION THROUGH THE AXIS OF AN EMBRYO. The section is supposed to be made at a time when the head-fold has com- menced but the tail-fold has not yet appeared. F. So. fold of the somatopleure. . Sp. fold of the splanchnopleure. _ The line of reference F’. So. is placed in the lower bay, outside the embryo. The line of D is placed in the upper bay inside the embryo; this will remain as the alimentary canal. Both folds (Ff. So., F. Sp.) are parts of the head-fold, and are to be thought of as continually travelling onwards (to the left) as develop- ment proceeds. pp. space between somatopleure and splanchnopleure: pleuroperitoneal cavity. Am. commencing (head) fold of the amnion. A fuller explanation is given under Fig. 16. forward, with a gentle ascent, regains the original level. As seen in section, then, the blastoderm at this spot may be said to be folded up in the form of the letter @. This fold we shall always speak of as the head-fold. In it we may recognize two limbs: an upper limb in which the curve is directed for- wards, and its bay, opening backwards, is underneath the blastoderm, 7.e. as we shall see, inside the embryo (Fig. 9, J) ; and an under limb in which the curve is directed backwards, and its bay, opening forwards, is above the blastoderm, 2.e; outside the embryo. If an @ like the above, made of some elastic material, were stretched laterally, the effect would be to make both limbs longer and proportionally narrower, and E, . 3 34 PRELIMINARY ACCOUNT. [CHAP. their bays, instead of being shallow cups, would become more tubular. Such a result is in part arrived at by the growth of the blastoderm; the upper limb of the 4 is continually. growing forward (but, unlike the stretched elastic model, in- creases in all its dimensions at the same time), and the lower limb is as continually lengthening backwards ; and thus both upper and lower bays become longer and longer. This we shall hereafter speak of as the travelling backwards of the head-fold. The two bays do not however both become tubular. The section we have been speaking of is supposed to be taken vertically along a line, which will afterwards become the axis of the embryo; and the lower bay of the @ is a section of the crescentic groove mentioned above, in its middle or deepest part. On either side of the middle line the groove gradually becomes shallower. Hence in sections taken on either side of the middle line or axis of the embryo (above or below the plane of the figures), the groove would appear the less marked the farther the section from the middle line, and at a certain distance would disappear altogether. It must be remembered that the groove is at first crescent-shaped, with its concavity turned towards what will be the hind end of the embryo (Fig. 11). As the whole head-fold is carried farther and farther back, the horns of the crescent are more and more drawn in towards the middle line, the groove becoming first semi- circular, then horse-shoe-shaped. In other words, the head- fold, instead of being a simple fold running straight back- wards, becomes a curved fold with a central portion in front running backwards, and two side portions running in towards the middle line. The effect of this is that the upper bay of the @ (that within the embryo) gets closed in at the sides as well as in the front, and thus speedily becomes tubular. The under bay of the @ (that outside the embryo) remains of course open at the sides as in front, and forms a sort of horse-shoe-shaped ditch surrounding the front end of the embryo. We have dwelt thus at length on the formation of the head-fold, because, unless its characters are fairly grasped, much difficulty may be found in understanding many events in the history of the chick. The reader will perhaps find the matter easier to comprehend if he makes for himself a rough 11] THE EMBRYONIC SAC. 35 model, which he easily can do by spreading a cloth out flat to represent the blastoderm, placing one hand underneath it, to mark the axis of the embryo, and then tucking in the cloth from above under the tips of his fingers. The fingers, covered with the cloth and slightly projecting from the level of the rest of the cloth, will represent the head, in front of which will be the semicircular or horse-shoe-shaped groove of the head-fold. At its first appearance the whole @ may be spoken of as the head-fold, but later on it will be found convenient to restrict the name chiefly to the lower limb of the 2. Some time after the appearance of the head-fold, an altogether similar but less conspicuous fold makes its ap- pearance, at a point which will become the posterior end of the embryo. This fold, which travels forwards just as the head-fold travels backwards, is the tail-fold (Fig. 8, C). In addition, between the head- and the tail-fold two lateral folds appear, one on either side. These are simpler in cha- racter than either head-fold or tail-fold, inasmuch as they are nearly straight folds directed inwards towards the axis of the body (Fig. 8, F), and not complicated by being crescentic in form. Otherwise they are exactly similar. As these several folds become more and more developed, the head-fold travelling backwards, the tail-fold forwards, and the lateral folds inwards, they tend to unite in the middle point; and thus give rise more and more distinctly to the appearance of a small tubular sac seated upon, and connected, by a continually-narrowing hollow stalk, with that larger sac which is formed by the extension of the rest of the blastoderm over the whole yolk. The smaller sac we may call the “embryonic sac,” the larger one “the yolk-sac.” As incubation proceeds the smaller sac (Fig. 8), gets larger and larger at the expense of the yolk- sac (the contents of the latter being gradually assimilated by nutritive processes into the tissues forming the growing walls of the former, not directly transferred from one cavity into the other). Within a day or two of the hatching of the ehick, ata time when the yolk-sac is still of some consider- able size, or at least has not yet dwindled away altogether, and the development of the embryonic sac is nearly com- plete, the yolk-sac (Fig. 8, WV) is slipped into the body of 3—2 36 PRELIMINARY ACCOUNT. [CHAP: the embryo, so that ultimately the embryonic sac alone re- mains. 6. The embryo, then, is formed by a folding off of a portion of the blastoderm from the yolk-sac. The general outline of the embryo is due to the direction and shape of the several folds which share in its formation ; these, while preserving a nearly perfect bilateral symmetry, present marked differences at the two ends of the embryo. Hence from the very first there is no difficulty in distinguishing the end which will be the head from that which will be the tail. In addition to this, the tubular sac of the embryo, while everywhere gradually acquiring thicker and thicker walls, undergoes at various points, through local activities of growth in the form of thickenings, ridges, buds or other processes, many modifications of the outline conferred upon it by the constituent folds. Thus bud-like processes start out from the trunk to form the rudiments of the limbs, and similar thickenings and ridges give rise to the jaws and other parts of the face. By the unequal development of these outgrowths the body of the chick is gradually moulded into its proper outward shape. 7. Were the changes which take place of this class only, the result would be a tubular sac of somewhat complicated outline, but still a simple tubular sac. Such a simple sac might perhaps be roughly taken to represent the body of many an invertebrate animal; but the typical structure of a bird or other vertebrate animal is widely different. It may very briefly be described as follows. First there is, above, a canal running lengthways along the body, in which are lodged the brain and spinal cord. Below this neural tube is an axis represented by the bodies of the vertebree and their continuation forwards in the structures which form the base of the skull. Underneath this, again, is another tube closed in above by the axis, and on the sides and below by the body-walls. Enclosed in this second tube, and suspended from the axis, is a third tube, consisting of the alimentary canal with its appendages (liver, salivary glands, lungs, &c., which are fundamentally mere diverticula from one simple canal). The cavity of the outer tube, which also contains the heart and other parts of the vascular system, is the general 11.] THE NEURAL TUBE. 37 body cavity; it is divided into a thoracic or pleural, and an abdominal or peritoneal cavity; these two cavities are, how- ever, from their mode of origin, portions of one and the same tube. Thus a transverse section of a vertebrate animal always shews the same fundamental structure: above a single tube, below a double tube, the latter consisting of one tube enclosed within another, the inner being the alimentary canal, the outer the general cavity of the body. Into such a triple tube the simple tubular embryonic sac of the chick is converted by a series of changes of a remarkable character. The upper or neural tube is formed in the following way. At avery early period the upper surface of the blastoderm in the region which will become the embryo, is raised up into two ridges or folds which run parallel to each other at a short distance on either side of what will be the long axis of the embryo, and thus leave between them a shallow longitu- dinal groove (Fig. 8, B, also Figs. 11,12, m.c). As these ridges, which bear the name of medullary folds, increase in height they arch over towards each other and eventually meet and coalesce in the middle line, thus converting the groove into a canal, which at the same time becomes closed at either end (Fig. 8, & J, also Fig. 13.—.M). The cavity so formed is the cavity of the neural tube, and eventually becomes the cerebro-spinal canal. The lower double tube, that of the alimentary canal, and of the general cavity of the body, is formed in an entirely different way. It is, broadly speaking, the result of the junc- tion and coalescence of the fundamental embryonic folds, the head-fold, tail-fold, and lateral folds; in a certain sense the cavity of the body is the cavity of the tubular sac described in the last paragraph. But it is obvious that a tubular sac formed by the folding in of a single sheet of tissue, such as we have hitherto con- sidered the blastoderm to be, must be a simple tubular sac possessing a single cavity only. The blastoderm however does not long remain a single sheet, but speedily becomes a double sheet of such a kind that, when folded in, it gives rise to a double tube. Very early the blastoderm becomes thickened in the region of the embryo, the thickening being chiefly due to an increase in the middle layer or mesoblast, while at the same 38 PRELIMINARY ACCOUNT. [CHAP. time it becomes split or cleft horizontally over the greater part of its extent into two leaves, an upper leaf and a lower leaf. In the neighbourhood of the axis of the body, beneath the neural tube, this cleavage is absent (Fig. 8, B; also Figs. 13—20), in fact, it begins at some little distance on either side of the axis and spreads thence into the periphery in all directions. It is along the thickened mesoblast that the cleavage takes place, the upper part of the mesoblast uniting with epiblast to form the upper leaf, and the lower part with the hypoblast to form the lower leaf. In the fundamental folds both leaves are involved, both leaves are folded downwards and inwards, both leaves tend to meet in the middle below; but the lower leaf is folded in mere rapidly, and thus diverges from the upper leaf, a space being gradually developed between them (Fig. 8). In course of time the several folds of the lower leaf meet and unite to form an inner tube quite independently of the upper leaf, whose own folds in turn meet and unite to form an outer tube separated from the inner one by an intervening space. The inner tube is the alimentary canal which is subsequently perforated at both ends to form the mouth and anus; the walls of the outer tube are the walls of the body, and the space between the two tubes is the general “serous cavity,” which being subsequently divided into pleural and peritoneal portions, may be spoken of as the pleuroperitoneal cavity. Hence the upper (or outer) leaf of the blastoderm, from its giving rise to the body-walls, is called the somatopleure’; the lower (or inner) leaf, from its forming the alimentary canal and its tributary viscera, the splanchnopleure’. This horizontal splitting of the blastoderm into a somato- pleure and a splanchnopleure, which we shall hereafter speak of as the cleavage of the mesoblast, is not confined to the region of the embryo, but gradually extends over the whole of the yolk-sac. Hence in the later days of incubation the yolk- sac comes to have two distinct coats, an inner splanchno- pleuric and an outer somatopleuric investment, separable from each other all over the sac. We have seen that, owing to the manner of its formation, the ‘embryonic sac’ is con- nected with the ‘yolk-sac’ by a continual narrowing hollow 1 Soma, body, pleuron, side, 2 Splanchnie, viscus, pleuron, side. 11.] THE AMNION. 39 stalk; but this stalk must, like the embryonic sac itself, be a double stalk, and consist of a smaller inner stalk within a larger outer one, Fig. 8, HH. The folds of the splanchnopleure, as they tend to meet and unite in the middle line below, give rise to a continually narrowing hollow stalk of their own, a splanchnic stalk, by means ot which the walls of the alimentary canal are continuous with the splanchnopleuric investment of the yolk-sac, and the interior of that canal is continuous with the yolk inside the yolk-sac. In the same way the folds of the somatopleure form a similar stalk of their own, a somatic stalk, by means of which the body- walls of the chick are continuous (for some time; the con- tinuity, as we shall see, being eventually broken by the development of the amnion) with the somatopleuric invest- ment of the yolk-sac; and the pleuroperitoneal cavity of the body of the chick is continuous with the narrow space be- tween the two investments of the yolk-sac. At a comparatively early period the canal of the splanch- nic stalk becomes obliterated, so that the material of the yolk can no longer pass directly into the alimentary cavity, but has to find its way into the body of the chick by absorp- tion through the blood-vessels. The somatic stalk, on the other hand, remains widely open for a much longer time; but the somatic shell of the yolk-sac never undergoes that thick- ening which takes place in the somatic walls of the embryo itself; on the contrary, it remains thin and insignificant. When accordingly in the last days of incubation the greatly diminished yolk-sac with its splanchnic investment is with- drawn into the rapidly enlarging abdominal cavity of the embryo, the walls of the abdomen close in and unite, without any regard to the shrivelled, emptied somatopleuric invest- ment of the yolk-sac, which is cast off as no longer of any use. (Fig. 8. Compare the series.) 8. Very closely connected with the cleavage of the meso- blast and the division into somatopleure and splanchnopleure, is the formation of the amnion, all mention of which was, for the sake of simplicity, purposely omitted in the description just given. The amnion takes its origin from certain folds of the somatopleure, and of the somatopleure only, in the following way. 40 PRELIMINARY ACCOUNT. [CHAP. At a time when the cleavage of the mesoblast has some- what advanced, there appears, a little way in front of the semi- lunar head-fold, a second fold (Fig. 11, also Fig. 8, C.), running more or less parallel or rather concentric with the first and not unlike it in general appearance, though differing widely from it in nature. In the head-fold the whole thickness of the blastoderm is involved ; in it both somatopleure and splanch- nopleure (where they exist, 7.e. where the mesoblast is cleft,) take part. This second fold, on the contrary, is limited entirely to the somatopleure. Compare Figs. 8 and 9. In front of the head-fold, and therefore altogether in front of the body of the embryo, the somatopleure is a very thin membrane, consisting only of epiblast and a very thin layer of mesoblast; and the fold we are speaking of is, in consequence, itself thin and delicate. Rising up as a semilunar fold with its concavity directed towards the embryo (Fig. 8, C, af), as it increases in height it is gradually drawn backwards over the developing head of the embryo. The fold thus covering the head is in due time accompanied by similar folds of the somatopleure, starting at some little distance be- hind the tail, and at some little distance from the sides (Fig. 8, C, D, E, Ff). In this way the embryo becomes surrounded by a series of folds of thin somatopleure, which form a continuous wall all round it. All are drawn gradually over the body of the embryo, and at last meet and completely coalesce (Fig. 8, H, J), all traces of their junction being removed. Beneath these united folds there is therefore a cavity, within which the embryo lies (Fig. 8, H, ae). This cavity is the cavity of the amnion. The folds which we have been describing are those which form the amnion. Each fold, of course, necessarily consists of two limbs, both limbs consisting of epiblast and a very thin layer of mesoblast; but in one limb the epiblast looks towards the embryo, while in the other it looks away from it. The space between the two limbs of the fold, as can easily be seen in Fig. 8, is really part of the space between the somatopleure and splanchnopleure; it is therefore continuous with the general space, part of which afterwards becomes the pleuroperitoneal cavity of the body, shaded with dots in the figure and marked (p p). So that it is possible to pass from the cavity between the two limbs of each fold of the amnion into the cavity which surrounds 11] THE ALLANTOIS. 41 the alimentary canal. When the several folds meet and coalesce together above the embryo, they unite in such a way that all their inner limbs go to form a continuous inner membrane or sac, and all their outer limbs a similarly con- tinuous outer membrane or sac. The inner membrane thus built up forms a completely closed sac round the body of the embryo, and is called the amniotic sac, or amnion proper, (Fig. 8, H, I, &c. a.), and the fluid which it afterwards con- tains is called the amniotic fluid, or liquor amnit. The space between the inner and outer sac, being formed by the united cavities of the several folds, is, from the mode of its forma- tion, simply a part of the general cavity found everywhere between somatopleure and splanchnopleure. The outer sac over the embryo lies close under the vitelline membrane, while its periphery is gradually extended over the yolk as the somatopleuric investment of the yolk-sac described in the preceding paragraph. 9. If the mode of origin of these two sacs (the inner or true amnion, and the outer or false amnion, as Baer called it) and their relations to the embryo be borne in mind, the reader will have no difficulty in understanding the course taken in its growth by an important organ, the allantois, of which we shall hereafter have to speak more in detail. The allantois is fundamentally an appendage of the alimentary canal, and may be regarded as a bud thrown out by the splanchnopleure close to its junction with the somatopleure at the hinder end of the embryo (Fig. 8, D, al.). Krom thence it grows first into the pleuroperitoneal cavity of the embryo, and thence very rapidly pushes its way by the development of a long stalk into the space between the true and false amniotic sacs (Fig. 8, G, 1). Curving over the embryo, it comes to lie over the embryo and the amnion proper, separated from the shell (and vitelline membrane) by nothing more than the thin false amnion. In this position it performs its functions as a respiratory organ. It is evident that though now placed quite outside the embryo, the space in which it lies is a continuation of that peritoneal cavity in which it took its origin. It is only necessary to add, that the false amnion either coalesces with the vitelline membrane, in contact with which 42 PRELIMINARY ACCOUNT. [CHAP. IL. it lies, or else replaces it, and in the later days of incuba- tion is known as the chorion. In the above account we have described the somatopleure as consisting of mesoblast as well as epiblast even in its most peripheral portions. The inner limbs of the amniotic folds undoubtedly contain mesoblastic elements, since the amnion proper contains plain muscular fibres. Some authors however regard the outer limbs of the amniotic folds (giving rise to the false amnion) and the somatopleure beyond them as being composed of epiblast only. CHAPTER III. THE CHANGES WHICH TAKE PLACE DURING THE FIRST DAY OF INCUBATION. 1. Durtna the descent of the egg along the oviduct, where it is exposed to a temperature of about 40° C, the blastoderm, as we have seen, continues to undergo im- portant changes. When the egg is laid and becomes cold these changes all but entirely cease, and the blastoderm remains inactive until, under the influence of the higher tem- perature of natural or artificial incubation, the vital activities of the germ are brought back into play, the arrested changes go on again, and usher in the series of events which we have now to describe in detail. The condition of the blastoderm at the time when the ego is laid is not exactly the same in all eggs, in some the changes being farther advanced than in others, though the differences of course are slight; in some eggs, especially in warm weather, changes of the same kind as those caused by actual incubation may take place, to a certain extent, in the interval between laying and incubation; lastly, in all eggs, both under natural and especially under artificial incubation, the dates of the several changes are, within the limits of some hours, very uncertain, particularly in the first few days ; one egg being found, for example, at 36 hours in the same stage as another at 24 or 30 hours, or a third at 40 or 48 hours. When we speak therefore of any event as taking place at any given hour or part of any given day, we are to be understood as meaning that such an event will generally be found to have taken place at about that time. We introduce exact dates for the convenience of description. 44 THE FIRST DAY. [CHAP. The changes which take place during the first day will be most easily considered under three periods: from the Ist to the 12th, from the 12th to the 20th, and from the 20th to the 24th hour. 2. From the 1st to about the 12th howr—During this period the blastoderm when viewed from above is found to have increased greatly in size. The pellucid area, which at the best is but obscurely marked in the unincubated egg, becomes very distinct (the central opacity having dis- appeared), and contrasts strongly with the opaque area, which has even still more increased both in distinctness and size. For the first few hours both the pellucid and opaque areas remain circular, and the only change, besides increase in size and greater distinctness which can be observed in them, is a slight ill-defined opacity or loss of transparency, which makes its appearance in about the middle of the pellucid area, This is known as the embryonic shield. 3. Slight as are the changes which can at this stage be seen from surface views, sections taken from hardened specimens bring to light many most important changes in the nature and arrangement of the constituent cells. It will be remembered that the blastoderm in the un- incubated egg is composed of two layers, an upper (Fig. 3, ep.) and an under layer; that the upper is a coherent membrane of columnar nucleated cells, but that the lower one (Fig. 3, 1.) is formed of an irregular network of larger cells in which the nuclei, if present, are rarely visible ; and that in addition to this there are certain still larger cells, called ‘ formative cells’ (Fig. 3, 6), lymg at the bottom of the segmentation- cavity. Under the influence of incubation changes take place very rapidly, which result in the formation of the three layers of the blastoderm. The upper layer, which we shall henceforward call the epiblast (Fig. 10, A), takes but little share in these changes. In the lower layer, however, certain of the cells begin to get flattened horizontally, their granules become less numerous, and a distinct nucleus makes its appearance in them; the cells so altered cohere together and form a mem- brane. (Fig. 10, C), The membrane thus formed, which is 11. ] THE MESOBLAST. 4.5 first completed in the centre of the pellucid area, we shall henceforward speak of as the hypoblast. Between it and the epiblast many of the cells of the original lower layer are enclosed, and in addition some of the formative cells (migrating by help of amceboid movements after the fashion of white-blood corpuscles) begin to travel round the edge of the hypoblast, and to pass in between it and the epiblast. The cells, whether originally “formative” cells or cells from the lower layer, thus gathered between the epiblast and hypoblast, undergo a process of endogenous cell-formation, by which the whole of the interior of each becomes converted into a number of new cells. These new cells, spherical in form, and possessing a large nucleus with a distinct nucleolus, are first formed in the centre of the pellucid area and sub- sequently in its periphery. They constitute the third layer or mesoblast (Fig. 10, B). The epiblast is the Hornblatt (corneal layer), and the hypoblast the Darm- driisenblatt (epithelial glandular layer) of the Germans, while those parts of the mesoblast which take part in the formation of the somatopleure and splanchno- pleure correspond respectively to the Haut-muskel-platte and Darm-faser-platte. All blood-vessels arise in the mesoblast. Hence the vascular layer of the older writers fall entirely within the mesoblast. The serous layer of the same authors includes the whole of the epiblast, but also comprises a certain portion of mesoblast ; for they speak of all the organs of animal life (skin, bones, muscle, &c.) as being formed out of the serous layer, whereas the epiblast proper gives rise only to the epidermis and to certain parts of the nervous system. In the same way their mucous layer corresponds to the hypoblast with so much of the mesoblast as takes part in the formation of _ the organs of organic life. Their vascular layer therefore answers to a part only of the mesoblast, viz. that part in which blood-vessels are especially developed. It is worthy of notice that the cells of the epiblast are themselves the direct results of segmentation; but that the hypoblast and mesoblast are formed at a subsequent period, and are therefore only indirectly the results of segmentation. The true difference between the hypoblast and mesoblast lies in the mode in which each layer is formed, and not in any essential differ- ence in the segmentation-spheres from which each is derived. J At about the time when the hypoblast is completely formed as a distinct membrane, the mescblast cells form a somewhat thick mass in the centre of the blastoderm, and cause the central opacity spoken of above as the embryonic shield. 4. Soon after this, between the 8th and 12th hours, the hitherto circular pellucid area becomes oval (the opaque area remaining circular). The oval is, with remarkable regu- 4G THE FIRST DAY. [CHAP. larity, so placed that its long axis forms a right angle, or very nearly a right angle, with the long axis of the egg itself. Its narrow end corresponds with the future hind end of the embryo: and we may henceforward speak of it as the hind end. If an egg be placed with its broad end to the right hand of the observer, the head of the embryo will in nearly all cases be found pointing away from him. At about the time when the pellucid area is beginning to undergo this change of shape, there appears in surface views, along a line corresponding with the long axis of the oval, and occupying not, as might perhaps be expected, its front but its hinder two-thirds, a narrow opaque streak, much more opaque, and therefore distinct, than the em- bryonic shield, but still shadowy and ill-defined. This is known as the primitive streak. SECTION OF A BLASTODERM AT RIGHT ANGLES TO THE LONG AXIS OF THE EMBRYO AFTER EIGHT HOURS’ INCUBATION. (Taken about midway between front and hind end.) A. epiblast. B. mesoblast. C. hypoblast. pr. primitive groove. f. fold in the blastoderm, probably produced by the action of the chromic acid. m. c. mesoblast cell; the line points to one of the peripheral mesoblast cells lying between epiblast and hypoblast. 6d. formative cells. The following are the chief points represented in the section. (1) The thickening of the mesoblast underneath the primitive groove pr., even when it is hardly at all present at the sides of the groove. (2) The hypoblast, C, early formed as a single layer of spindle-shaped cells. (3) The so-called segmentation- cavity, in which coagulated albumen is present. On the floor of this are the large formative cells bd. The line of separation between the epiblast and mesoblast underneath the primitive groove is too strongly marked in the figure. The primitive streak is no sooner formed than it becomes marked on its upper surface by a delicate shallow furrow s III] THE PRIMITIVE GROOVE. 47 running along its axis. In fresh specimens viewed with transmitted light, this furrow appears as a linear trans- parency, but in hardened specimens seen under reflected light may be distinctly recognized as a furrow or narrow groove, the bottom of which being thinner than the sides appears more transparent when viewed with transmitted light. It is known as the primitive groove. The nature of the changes by which it is brought about can only be learnt by the study of vertical sections (Fig. 10). These teach us that the opacity which marks out the primitive streak is chiefly due to a thickening of the mesoblast. In the for- mation, however, both of the primitive streak, and especially of the primitive groove, the epiblast also plays an important art. FE During these twelve hours the epiblast has been spreading rapidly, much more rapidly than the other two layers. Over the white yolk in the region of the opaque area it forms a layer one cell deep, but at the same time has become two or three cells deep in the centre of the pellucid area. In the pellucid area its constituent cells have become narrower (6) and more columnar, but over the opaque area flatter and broader (12 ») than they were at first. At the 12th hour therefore we find a distinct histological difference between the epiblast cells of the pellucid and those of the opaque area. Over the thickening of the mesoblast, which forms the basis of the primitive streak, the epiblast is also thickened ; the hypoblast, however, remains here, as in the rest of the blastoderm, a flat sheet consisting of a single layer of flat- tened (seen in sections as a single row (Fig. 10, C) of spindle- shaped) cells, which become larger and more irregular at the periphery. The thickening of the mesoblast and epiblast in the region of the primitive streak causes the upper outline of the blastoderm as seen in sections to rise above the general surface in a gentle curve (Fig. 10). The primitive groove is formed almost entirely by a pushing in or depression of the epiblast at the summit of this curve. . The thickness of the epiblast remains about the same on the sides as at the bottom of the groove. The mesoblast, on the contrary, is thinner immediately beneath the bottom of the groove than at the two sides, where it is decidedly 48 THE FIRST DAY, [CHAP. thicker than in the rest of the pellucid area. It is apparently this median thinning of the mesoblast which gives rise to the linear transparency seen in specimens viewed with transmitted light. The hypoblast, it may be remarked, is generally curved downwards beneath the primitive streak and groove, though not to the same extent as the epiblast. Thus the whole blastoderm is some- what curved in this region. ~ Immediately beneath the groove a kind of fusion takes place between the epiblast and mesoblast, though on close examination the line of junction between them can generally be made out. This apparent fusion His (Ueber die Erste Anlage des Wirbeltheirleibs) regarded as an event of great importance, and gave the name of axis-cord to the part in which it occurs. In fresh specimens a narrow (opaque) streak can be seen running down the centre of the groove; but it is not represented by any structure which can be seen in sections. The chief events then which occur during the first twelve hours of incubation are the establishment of the three layers of the blastoderm, and the appearance of the embryonic shield, of the primitive streak and of the primitive groove. 5. From the 12th to the 20th hour.—During this period the pellucid area rapidly increases in size, and from being oval becomes pear-shaped. The primitive groove grows even more rapidly than the pellucid area; so that by the 16th hour it is not only absolutely, but also relatively to the pellucid area, longer than it was at the 12th hour. The interval between its end and the circumference of the pellucid area continues to be greater in front than behind. At about the 16th hour, or a little later, a thickening of the mesoblast takes place in front of the primitive groove, giving rise to an opaque streak ending abruptly in front against a semicircular fold, which appears at this time near the anterior extremity of ‘the pellucid area (Fig. 11), and is known as the head-fold. In fresh specimens ‘this streak looks like a continuation from the anterior extremity of the primitive groove; but in hardened specimens it is easy to see that the connection is only an apparent one. Along the new streak a groove (Fig. 11, m.c.) is very soon formed, ‘which, narrow in “front, but. widening very much behind, embraces between its diverging walls “the anterior extremity of the primitive groove. This new groove, by the conversion of which into a tube the medullary canal will be formed, is known as the medullary groove. On each side of it the mesoblast is thickened, and the surface of the blastoderm raised up in the form of two longi- tudinal folds, known as the lamine dorsales, or the medullary folds (Fig. 11, A). Immediately beneath the bottom of the 111.] THE NOTOCHORD. 49 SURFACE VIEW OF THE PELLUCID AREA OF A BLASTODERM OF 18 HOURS. None of the opaque area is shewn, the pear-shaped outline indicating the limits of the pellucid area. At the hinder part of the area is seen the primitive groove pr., with its nearly parallel walls, fading away behind, but curving round and meeting in front so as to form a distinct anterior termination to the groove, about half way up the pellucid area. Above the primitive groove is seen the medullary groove m. ¢., with the medullary folds 4. These diverging behind, slope away on either side of the primitive groove, while in front they curve round and meet each other close upon a curved line which represents the head-fold. The second curved line in front of and concentric with the first is the com- mencing fold of the amnion. groove, however, the mesoblast is thinned out and very soon the cells in this position, separating from the lateral masses, adhere together in the middle line, and thus form between the epiblast and the hypoblast a flattened circular rod known as the notochord, seen in section as an elliptical aggregation of cells (Fig. 12, ch.) E. 4 50 THE FIRST DAY. [CHAP. The medullary groove differs in many important particulars from the primi- tive groove. Beneath the primitive groove the mesoblast always fuses more or less with the epiblast; this is never the case under the medullary groove. Under the primitive groove the mesoblast never shews any signs of differ- entiation into any organ; under the medullary groove the notochord is formed out of the mesoblast cells. The epiblast lining the bottom of the medullary groove frequently becomes very much thinner than at its sides; this seems never to be the case with the primitive groove. The primitive groove reaches its maximum growth before the appearance of the medullary groove; and after the appearance of the latter gradually becomes less and less conspicuous, and finally disappears without leaving a trace. A curved remnant of it is to be found at the hind end of the medul- lary canal between the 30th and 4oth hours, but by the 50th not a trace of it remains. By the earlier observers the primitive groove was supposed to become con- verted into the medullary canal. Dursy (Der Primitivstreif des Hiihnchens) was the first to give a correct account of its disappearance; and the distinction between it and the medullary groove has since been fully recognized by many observers. Goette (Archiv. Micr. Anat. Vol. X. 1873, pp. 145—199) describes the medullary groove as always appearing to the left of the primitive groove, and having its floor continuous with the left wall of the latter. He states that beneath this left wall the unsymmetrically placed axis-cord is found; indeed he considers that the notochord is a forward continuation of the axis-cord, and that the latter, as the primitive groove recedes before the medullary groove, becomes continuously converted into the former. The primitive groove then is a structure which appears early, and soon disappears without entering directly into the formation of any part of the future animal. Apparently it has no function whatever. We can only sup- pose that it is the rudiment of some ancestral feature. 6. By the 20th hour the medullary groove or canal, with its medullary folds or laminee dorsales, is fully established. It then presents the appearance, towards the hinder extremity of the embryo, of a shallow groove with sloping diverging walls which embrace between them the remains of the vanishing primitive groove. Passing forwards towards what will become the head of the embryo the groove becomes narrower and deeper with steeper walls. On reaching the head-fold (Fig. 11), which continually becomes more and more prominent, the medul- lary folds curve round and meet each other in the middle line, so as to form a somewhat rounded end to the groove. In front therefore the canal does not become lost by the gradual flattening and divergence of its walls as is the case behind, but has a definite termination, the limit being marked by the head-fold. In front of the head-fold, quite out of the region of the medullary folds, there is usually another small fold which is the beginning of the amnion (Fig. 11), 111. | THE HYPOBLAST. 51 We must now go back, and say a few words about the changes which the cells of the various layers undergo from the 12—20 hours. TNE, Tee pole ODO Oo (: > OICBOS OBO) QOGOCE 6 © GoerrOOesa® O® ode po 095996990 06 COOIDEDOS eBBEsEeagepeS coe, QaBSOOOOOo TRANSVERSE SECTION OF A BLASTODERM INCUBATED FOR 18 HOURS. The section passes through the medullary groove mec., at some distance behind its extreme front, and shews some of the chief puints in which it differs from the primitive groove. The chief of these are, (1) the presence underneath it of the notochord ch., (2) the absence of any apparent adhesion between the epiblast and the meso- blast, (3) the thickening of the mesoblast underneath the medullary folds, m/f. A, epiblast. B. mesoblast. C. hypoblast. m.¢c. medullary groove. m. f. medullary fold. ch. notochord: the small group of mesoblast cells separated by a narrow gap from the thicker mass of mesoblast on either side. It is to be noticed that the cells of the hypoblast become more columnar as they approach the edge of the pellucid area, and finally pass, without any strong line of demarcation, into the white-volk spheres. Only one half of the section is represented—if completed the section would be symmetrical about the line passing through the centre of the medullary canal, me. The hypoblast (Fig. 12, C) continues to be only one cell deep; the cells being, during the whole of this period, flatter in the centre, and larger and more irregular towards the peri- phery of the blastoderm. At about the 12th hour they are very irregular in size; shewing very great variations over a very small space. This probably implies that they are rapidly undergoing division. Later, however (about the 1sth hour), they are fairly uniform over particular regions, though they vary considerably in size at different parts of the pellucid area. In no case does the hypoblast extend beyond the edge of the pellucid area. The hypoblast cells along the central axis of the pellucid area, and for some little distance on each side, are smaller than elsewhere over the blastodermn. Over a small district just outside the embryo, and at about one-third of the way from the posterior extremity of the blastoderm, they are, from the 18th—23rd hour, considerably larger than anywhere else. The remaining hypoblast cells 4—2 Aa 52 THE FIRST DAY. [ CHAP. are intermediate in size between these very large cells and the smaller cells in the centre. During the whole of this period the hypoblast cells continue to be granular and filled with highly refractive spherules, exhibiting in this respect a marked contrast to their appearance at a later time. Their mode of increase is partly by division, but the layer grows chiefly in a manner which is very different and somewhat remarkable. LEefore the 12th hour the hypoblast at its margin ended abruptly against the white-yolk cells; but after that hour its relation to the white yoik becomes altered.. As they approach the white yolk the cells of the hypoblast become more and more filled with white-yolk spherules, and at the extreme edge of the pellucid area it is very difficult to say where the white yolk ends, and’ where the hypoblast begins. This is somewhat diagrammatically shewn in Fig. 12. The white-yolk spheres near the edge of the pellucid area have generally acquired nuclei, thouzh it is frequently difficult to see them owing tv the numerous highly refractive spherules which the spheres contain. The nearer they are to the edge of the pellucid area the fewer spherules they contain, and at the very edge it is almost impossible to say whether they ought to be called white- volk spheres or hypublast cells. The chief increase of the hypoblast therefore seems to take place through the conversion, cell for cell, of the white yolk into the hypoblast. During this period the mesoblast (Fig. 12, B) cells do not undergo any marked change. The layer itself enlarges to a certain extent through the multiplication of cells by the division of old ones; but the chief increase in bulk is probably due to the formative cells, which are continually passing round from the bottom of the segmentation-cavity to the mesoblast, and there become converted, in the way described (§ 3) above, into mesoblast cells. These formative cells are more numerous at the bottom of the segmentation- cavity at the 18th hour than they were at the first hour. This accession to their number is probably due to fresh ones being formed from the floor of white yolk. They appear to grow in size by absorbing the white-yolk spherules, with which indeed they are completely filled. The epiblast cells (Fig. 12, A) probably increase entirely by division, and seem to derive their nourishment from the white yolk on which the peripheral cells rest, and perhaps also from the albuminous fluid which fills the segmentation- cavity and occupies all the interstices between the cells of the various layers. The cells near the edge of the opaque area are the largest and flattest of the epiblast cells; those in the middle of the pellucid area are smaller than those at its edge. Outside the blastoderm there are to be seen on the surface of the yolk alternating transparent and opaque white rings. These are known as the halones, and frequently appear at the commencement of incubation. It is stated by His that they are to be explained by the white-yolk spheres under- going changes of two kinds. In the one case the spherules they contain are III. | THE MEDULLARY CANAL, 53 dissolved and give place to vacuoles ; where this occurs to a large extent an Opaque ring is formed. In the other case a solution of the protoplasm of the spheres takes place, and the spherules are let loose in large numbers; where this occurs a transparent ring is formed. The chief events then of the second part of the first day, are the appearance of the medullary folds and groove, the formation of the notochord, the beginning of the head-fold and amnion, and the histological changes taking place in the several layers. 7. From the 20th to the 24th hour. The head-fold en- larges rapidly, the crescentic groove becoming deeper, while at the same time the overhanging margin of the groove (the upper limb of the 4, Chap. 1. § 5), rises up above the level of the blastoderm; in fact, the formation of the head of the embryo may now be said to have definitely begun. 8. The medullary folds, increasing in size in every di- mension, but especially in height, lean over from either side towards the middle line, and thus tend more and more to roof in the medullary canal, especially near the head. About the end of the first day they come into direct contact and completely coalesce with each other at a point which hes at some little distance behind the head-fold, in the region which will afterwards become the neck. Union, having begun at this spot, rapidly runs forward till (early in the second day) the head-part is completely closed in; and then passes more slowly backwards. The whole of the anterior portion of the groove is closed in before the union has ad- vanced more than a very short distance towards the tail. In this way a tubular canal is formed, ending blindly in front, but as yet open behind. This is the medullary or neural canal (Fig. 13, M, Fig. 20, Mc.). It is not completely closed in at the tail till a period considerably later than the one we are considering. 9. Meanwhile important changes are taking place in the axial portions of the mesoblast, which lie on each side of the notochord beneath the medullary folds. In an embryo of the middle period of this day, examined with transmitted light, the notochord is seen at the bottom of the medullary groove between the medullary folds, as a transparent line shining through the floor of the groove when the embryo is viewed from above, On either side of the 54 THE FIRST DAY. [CHAP. notochord the body of the embryo appears somewhat opaque, owing to the thickness of the medullary folds; as these folds slope away outwards on either side, so the opacity gradually fades away in the pellucid area. There is present at the sides no sharp line of demarcation between the body of the embryo and the rest of the area; nor will there be any till the lateral folds make their appearance ; and transverse vertical sections shew (Fig. 12) that there is no break in the mesoblast, from the notochord to the margin of the pellucid area, but only a gradual thinning. 10. During the latter period of the day, however, the plates of mesoblast on either side of the notochord begin to be split horizontally into two layers, the one of which attach- ing itself to the epiblast, forms with it the somatopleure (Fig. 13, compare also Fig. 20, So.), while the other, attaching itself to the hypoblast, forms with it the splanchnopleure (Fig. 13, Be, Fig. 20, sp). By the separation of these two layers from each other, a cavity (Fig. 13, pp, and Fig. 20, pp), containing fluid only, and more conspicuous in certain parts of the embryo than in others, is developed. This cavity is the be- ginning of that great serous cavity of the body which after- wards becomes divided into separate cavities, We shall speak of it as the pleuro-peritoneal cavity. 11. This cleavage into somatopleure and splanchnopleure does not extend quite up to the walls of the medullary canal. Hence there is left along either side of the canal, between it and the line along which the cleavage begins, a tract or plate of uncleft mesoblast, which receives the name of vertebral plate, the more external mesoblast being called the lateral plate. At first each vertebral plate is not only unbroken along its length but also continuous at its outer edge with the upper and lower layers of the lateral plate of the same side. Very soon, however, clear transverse lines are seen, in surface views, stretching inwards across each vertebral plate from the lateral plate towards the notochord; and not long after a transparent longitudinal line makes its appearance on either side of the notochord along the line of junction of the lateral with the vertebral plate. These transparent lines are caused by the appearance of vertical clefts, giving rise to narrow spaces containing nothing II. | THE PROTOVERTEBR&, 55 but clear fluid; and transverse sections shew that they are due to breaches of continuity in the mesoblast only, the epiblast and hypoblast having no share in the matter. The first transverse lines which appear are two in number, one a little behind the other, about opposite the spot where the medullary folds first coalesced to form the neural tube. The longitudinal lines begin at about the same place and run thence backward, parallel to the notochord, as far as the closure of the medullary canal extends. Behind the first two transverse lines other parallel transverse lines in course of time make their appearance. Thus each vertebral plate appears in surface views to be cut up into a series of square plots, bounded by transparent lines. Each square plot is the surface of a corresponding cubical mass (Fig. 18, P. v., Fig. 20, P. v.). The two such \c» ‘do \Bc ‘Pp TRANSVERSE SECTION THROUGH THE DORSAL REGION OF AN EMBRYO OF THE Seconp Day. (Copied from His), introduced here to illustrate the forma- tion of the protovertebr, and the cleavage of the mesoblast. The provertebre appear irregularly quadrate; they would have been more distinctly square, had the section been one of the first day, before the appear- ance of the primitive aortz, and of the rudiments of the Wolffian ducts: com- pare Fig. 20. M. medullary canal. Pv. protovertebra. w. rudiment of Wolffian duct. A. epiblast. C. hypoblast. Ch. notochord. Ao. aorta. BC. splanch- nopleure. cubical masses first formed, lying one on either side of the notochord beneath, and a little to the outside of the medul- lary folds, are the first pair of protovertebre. Behind this first pair, but otherwise similarly situated, a second and third pair make their appearance during the first day. 56 THE FIRST DAY. [ CHAP. The vertebral plate, while still continuous with the lateral plate (the dis- tinction between the two being indicated solely by the cleavage of the latter), consists of several layers of cells; but of these only the uppermost layer, that immediately under the epiblast, appears to be continued into the somatopleure ; the whole of the remainder, including those cells which will eventually form the so-called nucleus of the protovertebrze, seem to pass directly into the splanchno- pleure. All these changes except the formation of the pleuro-peri- toneal cavity can be seen in surface views of fresh: trans- parent specimens, but their nature is best shewn in sections, 12. Since the commencement of incubation the area opaca has been spreading outwards over the surface of the yolk, and by the end of the first day has reached about the diameter of a sixpence. It appears more or less mottled over the greater part of its extent, but this is more particu- larly the case with the portion lying next to the pellucid area ; so much so, that around the pellucid area an inner ring of the opaque area may be distinguished from the rest by the difference of its aspect. At about the 20th—24th hours an increasing number of formative cells make their way from the segmentation-cavity to the edge of the area opaca, and there, immediately under- neath the epiblast, quickly become converted into a rather thick and somewhat irregular network of mesoblast cells. The mottled appearance of the inner ring spoken of above is due to changes taking place in this mass of mesoblast, changes which eventually result in the formation of what is called the vascular area, the outer border of which marks the extreme limit to which the mesoblast extends. During the whole of this period the medullary groove has been growing rapidly backwards, so that the primitive groove appears to be pushed further and further back, and at the same time becomes smaller and less conspicuous. The amniotic fold is at the end of the first day very noticeable. 13. The changes then which occur during the first day may thus be briefly summarized : (1) The hypoblast and mesoblust are formed from the ' segmentation-spheres, so that by the 6th to the 8th hour the three layers of the germ—the epiblast, the mesoblast, and the hypoblast—are definitely established. (2) The primitive streak is formed by a thickening of the mesoblast. It. SUMMARY. oF (3) The primitive groove is formed along the centre of the primitive streak. ; (4) The pellucid area becomes pear-shaped, the broad end corresponding with the future head of the embryo. Its long axis lies at right angles to the long axis of the egg. (5) The medullary groove makes its appearance in front of the primitive groove, and below it the notochord is formed out of mesoblastic cells. (6) The development of the head-fold gives rise to the first definite appearance of the head. (7) The medullary folds rise up and coalesce in the region of the neck to form the neural tube, the primitive streak and groove disappearing. (8) One or more pair of protovertebre make their ap- pearance. (9) By the cleavage of the mesoblast, the somatopleure separates from the splanchnopleure. (10) The first trace of the amnion appears in front of the head-fold. (11) ‘The vascular area begins to be be distinguished from the rest of the opaque area. It may be well to remark, before passing on to the second day, that out of the protovertebre are formed not only the permanent vertebre, but also the superficial dorsal as well as certain other muscles and the spinal nerves; that the pair of protovertebre first formed corresponds not with the first cervical vertebra of the adult chick, but rather with the third or even fourth ; for though the majority of the protovertebrae are formed regularly behind the first pair, two or even three pair may make their appearance in front of it; and lastly, that in the part of embryo which forms the head, the meso- blast is never cut up into protovertebre, and never under- goes cleavage to form somatopleure and splanchnopleure. CHAPTER IV. THE CHANGES WHICH TAKE PLACE DURING THE SECOND DAY. 1. The First Half of the Second Day. In attempting to remove the blastoderm from an egg which has undergone from 30 to 36 hours’ incubation, the observer cannot fail to notice a marked change in the consistency of the blastodermic structures. The excessive delicacy and softness of texture which rendered the extraction of an 18 or 20 hours’ blasto- derm so difficult, has given place to a considerable amount of firmness; the outlines of the embryo and its appendages are much bolder and more distinct; and the whole blastoderm can be removed from the egg with much greater ease. In the embryo itself viewed from above one of the fea- tures which first attracts attention is the progress in the head-fold (Fig. 15). The upper limb or head has become much more prominent, while the lower groove is not only proportionately deeper, but is also being carried back beneath the body of the embryo (Chap. I1. § 5.) 2. The medullary folds are closing rapidly. In the region of the head they have quite coalesced, a slight notch in the middle line at the extreme front marking for some little time their line of junction. The open medullary groove of the first day has thus become converted into a tube, the neural canal, closed in front, but as yet open behind. For a brief period the calibre of this tube is uniform throughout; but very speedily the front end dilates into a small bulb, whose cavity remains continuous with the rest of the neural canal, and whose walls, like those of the canal, are formed of epiblast. This bulb is known as the jirst cerebral vesicle, Fig. 14, 1B, CHAP. IV. | THE CEREBRAL VESICLES. 59 An Empryo CHICK OF THE FIRST DaY (ABOUT THIRTY-SIX HOURS) VIEWED FROM BELOW AS A TRANSPARENT OBJECT. pl. Outline of the pellucid area. FB. The forebrain or first cerebral vesicle, projecting from the sides of which are seen the optic vesicles op. A definite head is now constituted, the backward limit of the somatopleure fold being indicated by the faint line S. 0. Around the head are seen the two limbs of the amniotic head- fold: one, the true amnion a, closely enveloping the head, the other, the false amnion a’, at some distance from it. The head is seen to project beyond the anterior limit of the pellucid area. The splanchnopleure fold extends as far back as sp. Along its diverging limbs are seen the conspicuous venous roots or omphalo-mesaraic veins, uniting to form the heart A, which continuing forward as the bulbus arteriosus ba, is lost in the substance of head just in front of the somatopleure fold. Lying (in this position of the embryo) under the heart is seen the broad foregut d, the wide crescentic opening into which at the hind limit of the splanchnopleure fold is very conspicuous. Beneath the foregut are faintly seen the hind brain HB. and higher up and more distinctly the mid brain WB. _ These are not yet completely differentiated, and their limits are in consequ ence very obscurely indicated. 60. THE SECOND DAY. [ CHAP. Behind the splanchnopleure fo'd, marking the hind limits of the foregut, are seen the two rows of protovertebre, the dark line between which m. c. indicates the position both of the line of junction of the medullary folds and of the notochord. The front end of the notochord is seen at ch. underneath the forebrain; its hind end is indistinct. Towards the tail the protovertebre become indistinct and give place to the vertebral plates v. pl. Still further back, at the commencing tail, all the parts become indistinct, the remains of the primitive groove pv. being as conspicuous as anything else. and makes its appearance in the early hours of the second day. Behind it a second and a third bulb, the second and third cerebral vesicles, are successively formed in a similar manner; but the consideration of these, though they begin to make their appearance soon after the formation of the first cerebral vesicle, may be conveniently reserved to a later period. 3. The number of protovertebree increases rapidly. The one or two pairs which are seen at the end of the first day have by the middle of the second day multiplied to five, or eight, or even more, Figs. 14, 15, p.v, each being formed in the same way as the first. As was mentioned previously, the chief increase takes place from before backwards, the new protovertebre appearing behind the old ones; but one pair at least is probably formed in front of that which was the very first to appear. In the early part of this day the formation of new proto- vertebra keeps pace with the closing in of the medullary folds, so that that part of the canal which is already closed in is always flanked by protovertebre; but later on the formation of protovertebre lags behind, so that for some distance to- wards the hinder extremity the closed medullary canal is unprotected by protovertebre, Fig. 15. At the extreme end the medullary folds become shallower, diverge from each other, and afterwards meet again, thus forming a lozenge- shaped open depression known as the sinus rhomboidalis, Hig. 15, ear. Behind the sinus rhomboidalis there may generally be seen a small and usually curved remnant of the primitive groove. Fig. 15, p.r. 4. Ina former chapter it was pointed out (Chap. m1. § 5) that the embryo is virtually formed by a folding or tucking in of the limited portion of the blastoderm, first at the anterior extremity, and afterwards at the posterior extremity and at Iv. | THE ALIMENTARY CANAL, 61 ni a Fig. 15. EMBRYO OF THE CHICK AT 36 HOURS VIEWED FROM ABOVE AS AN OPAQUE OBJECT. (Chromic acid preparation.) f. b. front brain. m. b. mid brain. h. 6b. hind brain. op. v. optic vesicle. au. p. auditory vesicle. o. f. omphalo-mesaraic vein. p. v. protovertebra. m. f. line of junction of the medullary folds above the medullary canal. s. 7. sinus rhomboidalis. ¢. tail-fold. p. 7. remains of primitive groove. a. p. area pellucida. The line to the side between p. v. and m. f. represents the true length of the embryo. The biscuit-shaped outline indicates the margin of the pellucid area. The head, which reaches as far back as 0.f, is distinctly marked off ; but neither the somatopleuric nor splanchnopleuric folds are shewn in the figure; the latter diverge at the level of o.f, the former considerably nearer the front, somewhere between the lines m. b. and h. 6. The optic vesicles op. v. are seen bulging out beneath the superficial epiblast. The heart lying underneath the opaque body cannot be seen. The tail-fold, ¢., is just indicated ; no distinct lateral folds are as yet visible in the region midway between head and tail. At m. f. the line of junction between the medullary folds is still visible, being lost forwards over the cerebral vesicles, while behind the folds diverge to enclose the narrowing sinus rhomboidalis, s. r. 62 THE SECOND DAY. [CHAP. the sides. One of the results of this doubling up of the blasto-_ derm to form the head is the appearance, below the anterior extremity of the medullary tube, of a short canal, ending blindly in front, but open widely behind (Fig. 16, )),a cul de sac in fact, lmed with hypoblast reaching from the extreme front of the embryo to the point where the splanchnopleuric leaf of the head-fold (Fig. 16, # Sp) turns back on itself. This cul de sac, which of course becomes longer and longer the farther back the head-fold is carried, is the rudiment of the front end of the alimentary canal, the foregut, as it might be called. In transverse section it appears to be flattened horizontally, and also bent, so as to have its convex surface looking downwards, (Fig. 18 al). At first the anterior end is quite blind, there being no mouth at all; the formation of this at a subsequent date will be described later on. At the end of the first half of the second day the head- Fic. 16. DIAGRAMMATIC LONGITUDINAL SECTION THROUGH THE AXIS OF AN EMBRYO. The section is supposed to be made at a time when the head-fold has com- menced but the tail-fold has not yet appeared. N.C. neural canal, closed in front but as yet open behind. Ch. notochord, not reaching to the extreme front, and not as yet fully formed behind. The section being taken in the middle line, the protovertebre are of course not shewn. In front of the notochord is seen a mass of uncleft mesoblast, which will eventually form part of the skull. D. the commencing fore- gut or front part of the alimentary canal. /. So. Somatopleure, raised up in its peripheral portion into the amniotic fold Am. Sp. Splanchno- pleure. At Sp. it forms the under wall of the foregut; at F. Sp. itis turning round and about to run forward. Just at its turning point the cavity of the heart Ht is being developed in its mesoblast. pp. pleuroperitoneal cavity. A epiblast, B mesoblast, C hypoblast, indicated in the rest of the figure by differences in the shading. At the part where these three lines ef reference end the mesoblast is as yet uncleft. Iv.] THE HEART. 65 fold has not proceeded very far backwards, and its limits can easily be seen in the fresh embryo both from above and from below. 5. It is in the head-fold that the formation of the heart takes place, its mode of origin being connected with that cleavage of the mesoblast and consequent formation of splanch- nopleure and somatopleure of which we have already spoken. At the extreme end of the embryo (Fig. 16), where the ‘blastoderm begins to be folded back, the mesoblast is never cleft, and here consequently there is neither somatopleure nor -splanchnopleure; but at a point a very little further back, close under the blind end of the foregut, the cleavage (at the stage of which we are speaking) begins, and the somato- pleure, /. So, and splanchnopleure, / Sp. diverge from each other. They thus enclose between them a cavity, pp, which rapidly increases behind by reason of the fact that the fold of the splanchnopleure is carried on towards the hinder extremity of the embryo considerably in advance of that of the somatopleure. Both folds, after running a certain distance towards the hind end of the embryo, are turned round again, and then course once more forwards over the yolk-sac. As they thus return (the somatopleure having meanwhile given off the fold of the ammion, Am.), they are united again to form the uncleft blastodermic investment of the yolk-sac. In this way the cavity arising from their sepa- ration is closed below. It is in this cavity, which from its mode of formation the reader will recognise as a part (and indeed at this epoch it constitutes the greater part) of the general pleuroperitoneal cavity, that the heart is formed. It makes its appearance at the under surface and hind end of the foregut just where the splanchnopleure folds turn round to pursue a forward course, (Fig. 16, Ht); and by the end of the first half of the second day (Fig. 14, h) has ac- quired somewhat the form of a flask with a slight bend to the right. At its anterior end a slight swelling marks the future bulbus arteriosus ; and a bulging behind indicates the position of the auricles. It is hollow, and its cavity opens below and behind into two vessels called the omphalo-mesaraic veins (Figs. 14, 15,0.f), which pass outwards in the folds of the splanchnopleure at nearly right angles to the axis of the 64 THE SECOND DAY. [CHAP. embryo. The anterior extremity of the heart is connected with the two aorte. The muscular portion of the walls of the heart are derived in the chick (as in all other vertebrates in which the point has been worked out) from the mesoblast of the sp!anchnopleure. Although thus much may be asserted with tolerable certainty for all verte- brates; yet the exact mode of development appears, according to our present knowledge, to be very different in different cases; and it seems probable that these differences are in part the result of variations in the mode of formation and time of closure of the alimentary canal. In the chick the investigation of the earlier stages of the heart is beset with considerable difficulties; and accordingly various inquirers have arrived at very different results, though the majority are agreed as to its formation from the mesoblast of the splanchnopleure. Exact information concerning the epithe- lium lining the heart may be said to be almost completely wanting. Von Baer described the heart as consisting in its earliest stage of two solid aggregations of the mesoblast cells of the splanchnopleure, converging in front at the end of the foregut where they are loosely united together by a thin band, but diverging behind along the diverging folds of the splanchnopleure. As the foregut lengthens, the two masses coalesce more and more completely in front, © until the whole structure assumes the shape of a fusiform mass, attached to the under wall of the foregut, with prolongations stretching like the limbs of an inverted x along the folds of the splanchnopleure on either side. At first solid throughout, the A-shape mass subsequently becomes hollowed out and filled with fluid by the solution of its central cells. The account given by Remak (Entwickelung der Wirbelthiere, 1855) is some- what similar. According to His the heart is formed by the separation of a layer of the splanchnopleure and its coalescence with a similar layer from the somatopleure. It is therefore from the beginning hollow. Its cavity is also from the be- ginning continuous with the canals of the aorte and omphalo-mesaraic veins, the roots of which are formed in a precisely similar manner as itself. It is through these that the epithelial (endothelial) elements, derived from the white yolk, make their way into the heart to form its epithelial (endothelial) lining. According to Afanassieff (Bull. Acad. St. Pétershourg., Tom. X11. 1869, Ppp. 321—335) the heart is formed by the longitudinal separation of a thick layer of the mesoblast of the splanchnopleure along the under wall of the fore- gut. At either side, so much of the mesoblast is detached, that only a single layer of cells (seen spindle-shaped in a transverse section of the embryo) remains united with the hypoblast to form the wall of the gut. Along the middle line the separation is not complete, the detached layer of mesoblast being here still connected with the wall of the gut by a few cells. The single layer of spindle- shaped cells breaks loose in turn from the wall of the gut on either side of the middle line, but still remains attached along the middle line itself. We have thus in transverse section a thinner and a thicker layer of mesoblast, hanging down in a double festoon from the (hypoblastic) under-wall of the gut. Both layers become more and more separated from the gut, and bulge out into the pleuroperitoneal space, thus creating between themselves and the gut a cavity, which is at first double, but, by the disappearance of the cells along the middle line, subsequently becomes single. This is the cavity of the heart, the thick layer representing its muscular walls, and the thin its epithelial lining. The two ends are open, the hinder end being connected with the omphalo-mesaraic veins, and the front with the aorte. At first the heart is not a tube with Iv.] THE HEART. 65 complete walls of its own, but rather a cavity, closed in below and at the sides by its mesoblastic walls, and roofed over by the bare hypoblastic under-wall of the foregut. Very shortly, however, the side walls close in above, and thus pinch off the heart as a complete and distinct tube, which becomes quite detached along the greater part of its length from the wall of the gut, though it still remains connected with it, both at the venous and arterial ends. Klein (Wien. Sitzungsbericht, LXul. U., 1871) considers that the heart is formed from the cells of the mesoblast of the splanchnopleure as a body which, at first solid, subsequently becomes hollow by the conversion of its central cells inte blood-corpuscles. The layer of cells immediately surrounding the blood-corpuscles forms the epithelial lining, and subsequently becomes connected with that of the great arteries and veins. The following view, which our own observations have led us to adopt, agrees with that of Klein in regarding the heart as being at first a solid thicken- ing of the mesoblast of the splanchnopleure ; but its accordance with the earlier statements of Von Baer is much more complete. In order to understand the formation of the heart it must be distinctly borne in mind that in the region where the heart is about to appear, the splanch- nopleure is continually being folded in on either side, and that these lateral folds are progressively meeting and uniting in the middle line to form the under-wall of the foregut (that which im the adult chick will be the anterior wall of some portion of the alimentary canal). (Compare Chap. It. § 5.) At any given moment these folds will be found to have completely united in the middle line along a certain distance measured from the point in front where the cleavage of the mesoblast (i.e. the separation into svumatopleure and splanchnopleure) begins, to a particular puint tarther back. At this particular point the folds will have met, but not united, Fig. 17, 4. Further back still they will have not even met, but will appear simply inclined towards each other, Fig. 17, B. Or, to put it in another way, they will here be found to be diverging from the point where they were united, and not only diverg- ing laterally each from the middle line, but also both turning so as to run in a forward direction to regain the surface of the yolk and rejoin the somato- pleure, Fig. 16. In a transverse section taken bebind this extreme point of union, or point of divergence, as we may call it, the splanchnopleure on either side when traced downwards from the axis of the embryo may be seen to bend in towards the middle so as to approach its fellow, and then to run rapidly outwards, Fig. 17, B. A longitudinal section shews that it runs forwards also at the same time, Fig. 16. A section through the very point of divergence shews the two tolds meeting in the middle line and then separating again, so as to form something like the letter X, with the upper limbs con- verging, and the lower limbs diverging. In a section taken in front of the point of divergence, Fig. 18, the lower diverging limbs of the X have disappeared altogether; nothing is left but the upper limbs, which, completely united in the middie line, form the under-wall of the toregut. As development proceeds, what we have called the point of divergence is continually being carried farther and farther back, so that the distance between it and the point where the somatopleure and splanchnopleure separate from each other in front, 7.e. the length of the foregut, is continually increasing. When the heart is about to be tormed, thickenings are observed in the mesoblast of the splanchnopleure, along the diverging folds, 7.e. along the lower limbs of the X just behind the puint of divergence. ‘Lhese thickenings are continued into each other by a similar thickening of the mesoblast ex- tending through the point of divergence itself. At first there is no thickening of the mesoblast in front of the point of divergence, i.¢. along the under-wall of the foregut. As the pvuint of diverg- - i, 9) 66 THE SECOND DAY. [CHAP. ence however is in the course of events carried farther back, though the lower diverging folds (the lower limbs of the X) disappear, the thickening at the point remains and increases. In a short time, consequently, we do find a thickening of the mesoblast in the under-wall of the foregut just in front of the point of divergence, which thickening is continuous like an inverted X, with two thickenings reaching down the diverging folds behind the point of divergence. This x-shaped thickening becomes hollow by a transformation of its central cells; the single cavity in front is the cavity of the heart, and the two diverging cavities behind, with which it is continuous, are the canals of the omphalo-mesaraic veins. As development proceeds, and the point of divergence is carried still farther and farther back, the heart increases in length step by step at the expense of the continually coalescing omphalo-mesaraic veins. The coalescence of the mesoblastic thickening which forms the walls of the veins precedes that of their canals, consequently in sections taken at parti- cular points we meet with two cavities invested by one wall. This is probably what was seen by the observers who have described the heart as being formed as a double tube which afterwards became single. The front end of the cavity of the heart is continuous with canals similarly formed in the mesoblast of the foregut by the solution of certain cells. These are the canals of the aortz. At first the substance of the heart is along its whole length adherent to and indeed a part of the underwall of the foregut. Sub-equently it becomes free in its middle portion, the arterial and venous ends alone remaining attached. Soon after its formation the heart begins to beat, its at first slow and rare pulsations beginning at the venous and passing on to the arterial end. It is of some interest to note that its functional activity commences long before the cells of which it is composed shew any distinct differentiation into muscular or nervous elements. 6. To provide channels for the fluid thus pressed by the contractions of the heart, a system of tubes has made its appearance in the mesoblast both of the embryo itself and of the vascular and pellucid areas. In front the single tube of the heart bifurcates into two primitive aorte, each of which bending round the front end of the foregut, passes from its under to its upper side, the two forming together a sort of incomplete arterial collar imbedded in the mesoblast of the gut. Arrived at the upper side of the gut, they turn sharply round, and run separate but parallel to each other backwards towards the tail, in the mesoblast on each side of the notochord immediately under the protovertebre (Figs. 18, Ao, 20, Ao). About half way to the hinder extremity each gives off at right angles to the axis of the embryo a large branch, the omphalo- mesaraic artery (Fig. 23, Of, A.), which, passing outwards, is Iv.] THE BLOOD-VESSELS. 67 distributed over the pellucid and vascular areas, the main trunk of each aorta passing on with greatly diminished calibre towards the tail, in which it becomes lost. Fie. 17. A. Two cONSECUTIVE SECTIONS OF A 36 HOURS’ EMBRYO ILLUSTRATING THE FORMATION OF THE HEART. A IS THE MOST ANTERIOR OF THE TWO. h.b. hind brain. nc. notochord. Z. epiblast. so. somatopleure. sp. splanchnopleure. d. alimentary canal. hy. hypoblast. hz. (in A) heart. of. omphalo-mesaraic vein. The heart is seen from the sections to be formed from the mesoblast of the splanchnopleure. It is not however split off from a portion of the mesoblast which forms the muscular wall of the alimentary canal, but the mesoblast, where it turns round to run outwards again over the yolk-sack, becomes thickened, and in each of the thickenings (one on each side) so formed, a cavity appears 5—2 68 ‘THE SECOND DAY. [CHAP. forming immediately behind the heart the omphalo-mesaraic veins (section B), (of). As however the folding of the splanchnopleure becomes more complete, and the digestive canal becomes completely closed (instead of remaining partially open as in section B), these two cavities unite; and an appearance is produced similar to that represented in figure A, where there is the single cavity of the heart (fz). In the interior of the heart is seen a lining of flattened cells. The shading, .as will be seen, is purely diagrammatic. The epiblast, whether superficial as at #, or involuted as part of the neural canal hd, is shaded of one tint. The mesoblast, whether uncleft, or diverging into soma- topleure and splanchnopleure, is of another tint. In the hypoblast a distinction has been drawn between the thickened portion which lmes the alimentary canal, and the thinner portion which belongs to the more peripheral part of the splanchnopleure, the two being at first continuous as in B, and afterwards separated as in A. It will be understood that the two figures, though actually two consecutive sections of the same embryo, may be taken to represent two phases of the formation of the heart. B in the process of development will become 4, and A ashort time previously was in the condition of B. Fic. 18. TRANSVERSE SECTION OF AN EMBRYO AT THE END OF THE SECOND DAY PASSING THROUGH THE REGION OF BULBUS ARTERIOSUS. (Copied from His.) YY. medullary canal in the region of the hind brain. JV. anterior cardinal or superior vertebral vein. Ao. Aorta. Ch. Notochord. al. alimentary canal. H. Heart (bulbus arteriosus). Pp. Pleuroperitoneal cavity. am. amnion, On comparing this with Fig. 17, it will be seen that the mesoblast (muscular) wall of the heart H has now become quite separate from the rest of the mesoblast of the splanchnopleure, which forms, in the section, an independent line below the heart, the section of branches of the omphalo-mesaraic veins being seen on either side. The bridle of mesoblast represented in the drawing as passing from the splanchnopleure below to the somatopleure above, reaching the latter just inside the fold of the amnion, has been described by His, but has never been seen by ourselves. In the vascular and pellucid areas, the formation of vascular channels with a subseqnent differentiation into arteries, capillaries and veius, is proceeding rapidly. Blood- Iv.] THE BLOOD-VESSELS. 69 corpuscles too are being formed in considerable numbers. The mottled yellow vascular area becomes covered with red patches consisting of aggregations of blood-corpuscles, often spoken of as blood- islands. Round the extreme margin of the vascular area and nearly completely encircling it, is seen a thin red line, the sinus or vena terminalis (Fig. 23, Sv.). This will soon increase in size and importance. From the vascular and pellucid area several large channels are seen to unite and form two large trunks, one on either side, which running along the splanchnopleure folds at nearly right angles to the axis of the embryo, unite at the “ point of divergence” to join the venous end of the heart. These are the omphalo-mesaraic veins (Figs. 14, 0. f., 23, 0. f.) spoken of above. Both vessels and corpuscles are formed entirely from the cells of the mesoblast; and in the regions where the meso- blast is cleft, are at first observed exclusively in the splanch- nopleure. Ultimately of course they are found in the meso- blast everywhere. The mode of formation of the blood-vessels and corpuscles has been much and long debated. The observations of one of us have led us to believe the following to be the true account. In the pellucid area, where the formation of blood-vessels may be most easily observed, a number of mesoblastic cells are seen to send out processes. These processes unite, and by their union a protoplasmic network is formed containing nuclei at the points from which the processes started. The nuclei, which as a rule are much elongated and contain large oval nucleoli, increase very rapidly by division, and thus form groups of nuclei at the, so to speak, nodal points of the network. Several nuclei may also be seen here and there in the processes themselves. The network being completed, these groups, by con- tinued division of the nuclei, increase rapidly in size; the majority of the nuclei composing them acquire a red colour and become converted into blood-corpuscles (Fig. 19, 6.c.); but a few, generally on the outside of the group, remain un- unaltered, (Fig. 19, a). The protoplasm in which the central reddened nuclei are imbedded becomes liquefied, while that on the outside of each group, as well as that of the uniting processes, remains granular, and increasing in quantity, forms an investment for the unaltered nuclei which are embedded in it. Each nodal point is thus transformed into a more or less rounded mass of blood-corpuscles floating in plasma but enveloped by a layer of nucleated proto- plasm, the several groups being united by strands of nucleated protoplasm. These uniting strands rapidly increase in thickness; new processes are also continually being formed ; and thus the network is kept close and thickset while the area is increasing in size. By a transformation of nuclei similar to that which took place in the nodal points, blood-corpuscles make their appearance in the processes also, the central portions of which become at the same time liquefied. The uncoloured nuclei 70 THE SECOND DAY. [CHAP. situate in the envelopes of the nodal groups, as well as those lying on the exterior of the connecting processes, appropriate a quantity of the granular protoplasm surrounding each, and thus become converted into spindle-shaped cells. Each nodal group and each connecting process thus gets a distinct wall of nucleated cells. By the continued widening of the connecting processes and solution of their central portions, accompanied by a corresponding increase in the enveloping nucleated cells, the original protoplasmic network is converted into a system of communicating tubes, the canals of which contain blood-corpuscles and plasma, and the walls of which are formed of spindle-shaped nucleated cells. SURFACE VIEW FROM BELOW OF A SMALL PORTION OF THE POSTERIOR END OF THE PELLUCID AREA OF A 36 HOURS’ Cuick. To illustrate the formation of the blood-capillaries and blood-eorpuscles, magnified 400 diameters. 6. c. Blood-corpuscles at a nodal point, already beginning to acquire a red colour. They are enclosed in masses of protoplasm in the outermost layer of which are found nuclei, a, some of which contain two nucleoli. These nuclei subsequently become the nuclei of the cells forming the walls of the vessels. The nodal groups are united by protoplasmic processes (p.pr), also containing nuclei with large nucleoli (x). These nuclei increase in number by division, and become converted in part into the nuclei of the cells forming the walls of the vessels, and in part into blood-corpuscles, The blood-corpuscles pass freely from the nodal points into the hollow pro- cesses, and thus the network of protoplasm becomes a network of blood-vessels ; the corpuscles and the nuclei of the walls of which have been by separate paths of development derived from the nuclei of the original protoplasm. The formation of the corpuscles does not proceed equally rapidly or to the’ IV. | THE BLOOD-VESSELS. Tit same extent in all parts of the blastoderm. By far the greater part are formed in the vascular area, but some arise in the pellucid area, especially in the hinder part. In the front of the pellucid area the processes are longer and the network accordingly more open; the corpuscles also are both later in appearing and less numerous when formed. The omphalo-mesaraic arteries and veins, and the sinus terminalis which from the first has a distinct wall, seem to take origin in a manner altogether similar to that of the smaller vessels ; and the description of the formation of the heart which we gave above shews that it too is nothing but a gigantic nodal point. Assuming the truth of the above account, it is evident that the blood-vessels of the chick do not arise as spaces or channels between adjacent cells of the mesobiast, but are hollowed out in the communicating protoplasmic substance of the cells themselves, It is also perhaps worthy of note that the red-blood corpuscles are not cells, but nuclei. The red-blood corpuscles when removed from the vessels exhibit energetic amceboid movements. They seem to increase at this stage chiefly by division. The above is the view which we deduce from our own observations. The following may serve as a brief summary of the history of the matter. Von Baer and the older embryologists regarded the blood-vessels as being at first mere gaps or spaces betweeu the cellular elements of the mesoblast, hollowed out so to speak by the flow of blood from the heart. The first steps in the right direction were taken by Remak and Kdlliker, who described the formation of solid bands or cylinders composed of cells and arranged in a close-set network. Tlese bands, becoming hollowed by solution while their central cells were converted into blood-corpuscles, gradually put on the appearance of blood-vessels, the aggregation of the red corpuscles at various points, through arrest of the circulation, giving rise to the blood-islands of Wolff and Pander. According to Afanassieff (Wien. Sitz, Bericht. Bd. 53, 1866) there appear in the mesoblast vesicles of variable size, with protoplasmic envelopes and contents. These vesicles, which are at first clear and homogeneous, subse- quently become traversed with strands of nucleated protoplasm, forming often a close net-work within the vesicle. The space intervening between the nume- rous vesicles is cut up into a network of canals by protoplasmic processes stretching from one vesicie to another. These canals are the rudimentary blood-vesse!ls. From the outside of the vesicles, forming the inner wall of the adjacent vessels, nucleated masses of protoplasm are budded off as blood-cor- puscles and fall into the current of the circulation. His (op. cit.), following out his peculiar theory of development, derived both blood and blood-vessels from the white yolk or parablast. According to him while certain of the white-yolk masses become converted into conglomerations of cells, which acquiring a yellow colour stand out in surface yiews as blood- islands, other white-yolk masses, metamorphosed into anvular cells, form a network of thick lines permeating the mass of true blastodermic (archiblastic) cells of the mesoblast. These lines, at the first solid, subsequently become hollow. The meshwork of canals, or rudimentary blood-vessels, thus developed first in the vascular and pellucid areas and spreading thence into the embryo, contains for a certain time clear fluid only, the blood-islands being imbedded in or attached to the walls of the canal and surrounded by protoplasmic envelopes, so that the blood-corpuscles are shut out from the cavities of the vessels. Later on, however, the envelopes of the islands are broken through, and the blood-corpuscles emerging from their nests fall into the current of the circulation. His therefore regarded the blood-corpuscles as formed in greater part at least 72 THE SECOND DAY. [ CHAP. separately from the blood-vessels ; their entrance into the vascular spaces being an after event. The parablastic cells (derived trom the white yolk), in his view, give rise to the epithelium (endothelium) and connective tissue elements only of the blond vessels, the muscular elements being derived from genuine (blasto- dermic) mesoblastic cells. Klein (Wien. Sitz. Bericht. uxmt. 1871) describes the blood-vessels as taking their origin from certain cells of the mesoblast in which a vacuole, appearing and rapidly increasing in size, pushes the nucleus on one side, leaving only a thin layer of protoplasm round the periphery of the cell. In this thin layer nuclei appear; and, multiplying, form a complete nucleated invest- ment. to the vacuole, which meanwhile continues to increase in size. From the inside of this protoplasmic investment cells are budded off, and fall into the vacuole. Here they soon acquire a red colour and become converted into blood-corpuscles. From the exterior of these vacuolated cells nucleated processes are thrown out, which end freely or join with similar processes from other cells. A protoplasmic network is thus formed, the lines of which become vacuolated, and hollow, and ultimately communicate with the original central: vacuoles now crowded with corpuscles. By these means a system of com- municating tubes is established. Klein also describes two other forms of cells somewhat differing from the above, but also taking part in the formation of the blood-vessels. One of these forms is found chiefly in the vascular area, and he believes that these latter are simply the formative cells of which we have already so often spoken. It wil thus be seen that Klein’s view, from which our own differs chiefly in reference to the matter of vacuolation, is a return, with some modifications and extensions, to the earlier view of Remak, and that the accounts of both Afanas- sieff and His, which in turn agree in many respects, have proved to be uncorro- borated divergences from the older track. Suil more recently Goette (Archiv fiir Micro, Anat. Vol. X. 1873, pp. 145—199) has given an entirely different account of the origin of the blood- vessels and blood-corpuscles in the vascular area. He believes that in the thick mass of cells immediately outside the ‘pellucid area’ (vide Chap. 111. § 12) a quantity of fluid collects and causes the cells to separate into a network with large spaces filled with fluid. Into these spaces the formative cells travel, and undergoing a species of endogenous cell-formation, form masses of bl.od- corpuscles—the blqod-islands of the earier authors. This view differs, it will be seen, from all the later views, and goes back to that of Von Baer in rezard- jing the blood-vessels as primitively mere gaps between the cellular elements. In the investigation of such a point as this, sections (which apparently Goette has alone employed) are very untrustworthy. 7. The cells of the epiblast and hypoblast as well as of the mesoblast undergo considerable changes between the 24th and the 36th hour. Up to the 24th hour the cells of both layers, but more especially of the hypoblast, were filled with fine granules aud also contained many highly refractive spherules. By the 36th hour, however, they have become much more transparent. Each cell nuw consists of a clear protoplasm with hardly any granules or spherules, and a large oval nucleus together with one or more vacuoles is distinctly visible. The cells of the hypublast still pass insensibly into the white-yolk cells ; and it is still by the conversion of the while yolk into hypoblast that the peripheral extension of the latter is chiefly carried on. The hypoblast cells beneath and at the sides of the embryo are markedly smaller than those at the periphery of the pellucid area, The epiblast cells exhibit considerable variation in size in different parts of IV.] THE WOLFFIAN DUCT. 73 the embryo; but all are considerably smaller and also somewhat more columnar than the more peripheral cells of the pellucid area. The largest epiblastic cells are to be found in the region of the vascular area, but here they are much flattened. At the extreme outer edge of the opaque area the cells are smaller again, shewing in this respect a marked contrast to their con- dition during the previous stage. 8: About this period there may be seen in transverse sections, taken through the embryo in the region of the proto- vertebra, a small group of cells (Fig. 20, W. d) projecting on either side from the mass of uncleft mesoblast on the outside of the protovertebra, into the somewhat triangular space formed by the epiblast above, the upper and outer angle of the protovertebra on the inside, and the mesoblast on the outside. This group of cells is the section of a longitudinal ridge, the rudiment of the Wolfian duct. We shall return to it immediately. 9. The most important changes then which take place during the first half of the second day, are the closure of the medullary folds, especially in the anterior part, and the dilatation of the canal so formed into the first cerebral vesicle ; the establishment of a certain number of protoverte- bre; the elevation of the head from the plane of the blastoderm ; the tormation of the tubular heart and of the great blood-vessels; and the appearance of the rudiment of the Wolffian duct. It is important to remember that the embryo of which we are now speaking is simply a part of the whole germinal membrane, which is gradually spreading over the surface of the yolk. It is important also to bear in mind that all that part of the embryo which is in front of the most anterior protovertebrz corresponds to the future head, and the rest to the neck, body and tail. At this period the head occupies nearly a third of the whole length of the embryo. 10. The changes which take place from the 36th to the 45th hour will best form the next stage, since those which occur during the last few hours of the second day will be more conveniently described with the third day. One important feature of the stage is the rapid increase in the process of the folding off of the embryo from the plane of the germ, and its consequent conversion into a distinct tubular cavity. At the beginning of the day, the head alone projected from the rest of the germ, the remainder of the [CHAP. THE SECOND DAY. Fig. 20. TRANSVERSE SECTION THROUGH THE DorSAL REGION OF AN EMBRYO OF 45 HOURS. duct. S. 0. Somatopleure. S. p. Splanchnopleure. 233 ee O38 Es ae allies se on a Py deem = oe & Gye) ee 8 ae wo S £8 mE ane a, SS BR x3 ote fe)! BE an Cad 4 Cz b=} ss ae ae gS Ex a oO ee 23 of “a8 oO x Iv.] THE TAIL-FOLD. 75 cavity. c.h. notochord. a.o. dorsal aorta. v. blood-vessels of the yolk-sac. 0. p. line of junction between opaque and pellucid areas; at this point the hypoblast cells are seen ta pass without any strong line of demarcation into the white-yolk spheres. w. white-yolk spheres, some of which near the edge of the pellucid area contain a body very like a nucleus. Only one half of the section is represented in the figure—if completed it would be bilaterally symmetrical about the line of the medullary canal. Fig. 21. op EMBRYO OF THE CHICK AT 36 HOURS VIEWED FROM ABOVE AS AN OPAQUE OxssEcT. (Chromic acid preparation. ) f. b. front brain. m.b. mid brain. h. 6. hind brain. op. v. optic vesicle. au. p. auditory vesicle, o. f. omphalo-mesaraic vein. jp. v. protovertebra. m. f. line of junction of the medullary folds above the medullary canal. 8. r. sinus rhomboidalis, ¢. tail-fold. jp. 7. remains of primitive groove. a. p. area pellucida. embryo being simply a part of a flat blastoderm, nearly completely level from the front protovertebre to the hind edge of the pellucid area. At this epoch, however, a tatl-fold 76 THE SECOND DAY. [CHAP.. (Fig. 21, t) makes its appearance, elevating the tail above the level of the blastoderm in the same way that the head was elevated. Lateral folds also, one on either side, soon begin to be very obvious. By the progress of these, together with the rapid backward extension of the head-fold and the slower forward extension of the tail-fold, the body of the embryo becomes more and more distinctly raised up and marked off from the rest of the blastoderm. 11. The medullary canal closes up rapidly. The wide sinus rhomboidalis becomes a narrow fusiform space (Fig. 21, s.r.), and at the end of this period is entirely roofed over. The conversion of the original medullary groove into a closed tube is thus completed. 12. In the region of the head most important changes now take place. We saw that at the beginning of this day the front end of the medullary canal was dilated into a buib, the first cerebral vesicle. This, from the very first broader than long, now increases so much in breadth as to give the embryo a hammer-headed appearance. The lateral portions, continuing to enlarge, become after a while separated by constrictions from the central portion. The single vesicle is thus converted into three vesicles: a median one connected by short hollow stalks with a lateral one on either side. The lateral vesicles are known as the optic vesicles (Fig. 21, op. v, Fig. 22, a), and will afterwards become converted into parts Fie. 22, HEAD oF A CHICK AT THE END OF THE SECOND DAY VIEWED FROM BELOW AS A TRANSPARENT OBJECT. (Copied trom Huxley). I. first cerebral vesicle. a. optic vesicle. d. infundibulum. The specimen shews the formation of the optic vesicles (a), as outgrowths from the rst cerebral vesicle or vesicle of the 3rd ventricle, so that the optic vesicles and vesicle of the 3rd ventricle at first freely communicated with each other, and also the growth of the lower wall of the vesicle of the 3rd ventricle into a process which becomes the infundibulum-(d). Iv.] THE OPTIC VESICLES. 77 of the eyes; the median one still retains the name of the first cerebral vesicle. The constriction takes place chiefly from above downwards, so that the optic vesicles soon appear to spring from the under portions of the cerebral vesicle. The original vesicle being primarily an involution of the epiblast, the walls of all three vesicles are formed of epiblast ; all three vesicles are likewise covered over with the common epiblastic investment which will eventually become the epidermis of the skin of the head. Between this superficial epiblast and the involuted epiblast of the vesicles, there exists a certain quantity of mesoblast to serve as the material out of which will be formed the dermis of the scalp, the skull, and other parts of the head. At this epoch, however, the mesoblast is found chiefly underneath the several vesicles. A small quantity may in section be seen at the sides; but at the top the epidermic epiblast is either in close contact with the involuted epiblast of the cerebral and optic vesicles or separated from it by fluid alone, there being as yet in this region no cellular elements between the two representing the mesoblast. The constrictions marking off the optic vesicles take place of course beneath the common epiblastic investment, which is not involved in them. As a consequence, though easily seen in the transparent fresh embryo (Fig, 22), they are but slightly indicated in hardened specimens (Fig. 21). In sections they are very clearly seen. 13. When an embryo of the early part of the second day is examined as a transparent object, that portion of the medullary canal which lies immediately behind the first cerebral vesicle is seen to be conical in shape, with its walls thrown into a number of wrinkles. These wrinkles may vary a good deal in appearance, and shift from time to time, but eventually, before the close of the second day, after the formation of the optical vesicles, settle down into two con- strictions, one separating the first cerebral vesicle from that part of the medullary canal which is immediately behind it, and the other separating that second portion from a third. So, instead of there being one cerebral vesicle only, as at the commencement of the second day, there is now, in addition to the optic vesicles, a series of three, one behind the other; a second and third cerzbral vesicle have been added to the 78 THE SECOND DAY. [CHAP. first (Fig. 21, mb, hb). They may be also called the “fore brain,” the “mid brain,” and the “hind brain,” for into these parts will they eventually be developed. 14. The optic vesicles, lying underneath the epiblast, towards the end of the day are turned back and pressed somewhat backwards and downwards against the sides of the first cerebral vesicle or fore brain, an elongation of their stalks permitting this movement to take place. The whole head becomes in consequence somewhat thicker and rounder. 15. Before the end of the .day the fore brain, by a pro- cess similar to that whereby the optic vesicles were formed, viz. undue growth followed by constriction, has begun to bud off two small vesicles in front; these are the vesicles of the cerebral hemispheres, which subsequently become the most conspicuous part of the brain, but up to the end of the day are still very small and inconspicuous. 16. The notochord, whose origin was described in the account of the first day (Chap. m1. § 5), is during the whole of the second day a very conspicuous object. It is seen as a trans- parent rod, somewhat elliptical in section (see Fig. 20, ch), lying immediately underneath the medullary canal for the greater part of its length, and reaching forward in front as tar as below the centre of the second cerebral vesicle, where it ends either in a point (Remak), or in a rounded knob (Baer, Dursy, Entwickelungsgeschichte des Kopfes). The ex- act relations of its termination will be discussed later on. Round the anterior termination of the notochord, the medullary canal, which up to the present time has remained perfectly straight, towards the end of the day begins to curve. The front portion of the canal, 7.e. the fore-brain with its optic and cerebral vesicles, becomes slightly bent downwards, so as to form a rounded obtuse angle with the rest of the embryo. This is the commencement of the so-called cranial flexure. 17. Lastly, as far as the head is concerned, the rudiment of the ear appears about this time on the dorsal surface as a small depression or pitting of the epiblast on either side of the hind-brain (Fig. 21, aw. p). 18. We left the heart as a fusiform body slightly bent to the right, attached to the under wall of the foregut by the - Iv.] THE CIRCULATION. one aorta and by its venous end, but with its intermediate portion quite free. The curvature now increases so much that the heart becomes almost m-shaped, the venous portion being drawn up towards the head so as to lie somewhat above (dorsal to) and behind the arterial portion. (It would perhaps be more correct to say that the free intermediate portion is by its own growth bent downwards, backwards, and somewhat to the right, while the venous root of the heart is at the same time continually being lengthened by the carrying back of that “point of divergence ” of the splanchnopleure folds which marks the union of the omphalo-mesaraic veins into a single venous trunk). The heart then has at this time two bends, the one, the venous bend, the right-hand curve of the ; the other, the arterial bend, the left-hand curve of the ». The venous bend which, as we have said, is placed above and somewhat behind the arterial bend, becomes marked by two bulgings, one on either side. These are the rudiments of the auricles, or rather of the auricular appendages. The ascending limb of the arterial bend soon becomes conspicuous as the bulbus arteriosus, while the rounded point of the bend itself will hereafter grow into the ventricles. 19. The blood-vessels, whose origin during the first half of this day has been already described, become during the latter part of the day so connected as to form a complete system, through which a definite circulation of the blood is now for the first time (consequently some little while after the commencement of the heart’s pulsation) carried on. The two primitive aortw have already been described as encircling the foregut, and then passing along the body of the embryo immediately beneath the protovertebre on either side of the notochord. They are shewn in Fig. 20 a.o in section as two large rounded spaces lined with spindle-shaped cells. At first they run as two distinct canals along the whole length of the embryo; but, after a short time, unite at some little distance behind the head into a single trunk, which lies in the middle line of the body immediately below the notochord (Fig. 39). Lower down, nearer the tail, this single primitive trunk again divides into two aortz, which, getting smaller and smaller, are finally lost in the small blood-vessels of the tail. At this epoch, therefore, there are two aortic arches springing from the bulbus arteriosus, and 80 THE SECOND DAY. [ CHAP. uniting above the alimentary canal in the back of the embryo to form the single dorsal aorta, which travelling backwards in the median line divides near the tail into two main branches. From each of the two primitive aorte, or from each of the two branches into which the single aorta divides, there is given off on either side a large branch. These have been already spoken of as the omphalo-mesaraic arteries. At this stage they are so large that by far the greater part of the blood passing down the aorta finds its way into them, and a small remnant only pursues a straight course into the continuations of the aorta towards the tail Each omphalo-mesarai‘c artery leaving the aorta at nearly right angles (at a point some little way behind the backward limit of the splanchnopleure fold which is forming the ali- mentary canal), runs outwards beneath the protovertebre in the lower range of the mesoblast, close to the hypoblast. Consequently, when in its course outwards it reaches the point where the mesoblast is cleft to form the somatopleure and splanchnopleure, it attaches itself to the latter. Travel- ling along this, and dividing rapidly into branches, it reaches the vascular area in whose network of small vessels (and also to a certain extent in the similar small vessels of the pellucid area) it finally loses itself. The terminations of the omphalo-mesaraic arteries in the vascular and pellucid areas are further connected with the heart in two different ways. From the network of capillaries, as we may call them, a number of veins take their origin, and finally unite into two main trunks, the omphalo-mesaraie veins. ‘These have already been described as running along the folds of the splanchnopleure to form the venous roots of the heart. Their course is consequently more or less parallel to that of the omphalo-mesaraic arteries, but at some little dis- tance nearer the head, inasmuch as the arteries run in that part of the splanchnopleure which has not yet been folded in to form the alimentary canal. Besides forming the direct roots of the omphalo-mesaraic veins, the terminations of the omphalo-mesaraic arteries in the vascular area are also connected with the sinus terminalis spoken of above as run- ning almost completely round, and forming the outer margin of the vascular area. This (Fig. 23, 8.v), may be best de- scribed as composed of two semicircular canals, which nearly IV.] THE AORTIC ARCHES. $1 meet at points opposite the head and opposite the tail, thus all but encircling the vascular area between them. At the point opposite the head the end of each semi- circle is connected with vessels (Fig. 23), which run straight in towards the heart along the fold of the splanchnopleure, and join the right and left omphalo-mesaraic veins. At the point opposite the tail there is at this stage no such definite connection. At the two sides, midway between their head and tail ends, the two semicircles’ are espe- cially connected with the omphalo-mesaraic arteries. The circulation of the blood then during the latter half of the second day may be described as follows. The blood brought by the omphalo-mesaraic veins falls into the twisted cavity of the heart, and is driven thence through the bulbus arteriosus and aortic arches into the aortic trunk. From the aorta, by far the greater part of the blood Hows into the omphalo-mesaraic arteries, only a small remnant passing on into the caudal terminations. From the capillary net-work of the vascular and pellucid area into which the omphalo- mesaraic arteries discharge their contents, part of the blood is gathered up at once into the lateral or direct trunks of the omphalo-mesaraic veins. Part however goes into the middle region of each lateral half of the sinus termi- nalis, and there divides on each side into two streams. One stream, and that the larger one, flows in a forward direction until it reaches the point opposite the head, thence it returns by the veins spoken of above, straight to the omphalo-mesaraic trunks. The other stream flows backward, and becomes lost at the point opposite to the tail. This is the condition of things during the second day; it becomes considerably changed on the succeeding day. At the time that the heart first begins to beat the capillary system of the vascular and pellucid areas is not yet completed ; and the fluid which is at first driven by the heart contains, according to most observers, very few corpuscles. 20, At the close of the second day the single pair of aortic arches into which the bulbus arteriosus divides is found to be accompanied by a second pair, formed in the same way as the first, and occupying a position a little behind it. Sometimes even a third pair is added. Of these aortic arches we shall have to speak more fully later on. E, 6 82 THE SECOND DAY. [ CHAP. 21. At the latter end of this day, the ridge which we have already spoken of as the rudiment of the Wolffian duct, has become distinctly hollow, is in fact no longer a ridge but a canal. Sections now shew not an irregular group of ordi- nary mesoblastic cells, but a small cavity surrounded by a wall of cells; and these cells are beginning to put on a columnar character, and thus appear to radiate from the cen- tral cavity. The canal or duct so formed, the anterior termination of which is closed, and the posterior not as yet completely formed, reaches from about the fifth pair of protovertebree backwards towards the hind end of the embryo. The conversion of the ridge into a canal appears to take place by the eells acquiring a radiating arrange- ment, and a small hole appearing at the centre where the points of the cells meet; this rapidly grows larger till it reaches the final size of the cavity of the duct. The exact mode of development of the Wolffian duct is still a matter of some doubt, although its origin has been investigated by numerous embryologists. Remak, and after him Kolliker, described it as taking its origin from the mesoblast of the somatopleure, and appearing about the middle of the second day, at the external border of the protovertebre immediately under the epi- blast, in the form of a solid cord which subsequently became hollow. Dursy (Zeitsch. f. Rat. Med. 1865) gave a very similar account, except that he regarded it as being derived from the substance of the protovertebre, instead from the somatopleure. Hensen (Archiv. Microscop. Anat. Bd. 111. 1867), and for some time His, believed that the duct took origin as a longitudinal involution of the epiblast between the protovertebre and the lateral mesoblast, in the form of a groove, which subsequently became closed in and detached from the superficial epiblast, in a manner very similar to the way in which the lens is formed. Subsequently His tovk up the view that it was a product of the proto- vertebre, the central cells of these bodies, according to him, protruding as a ridge along their upper and external angles. He states that at first a distinct connection is visible between the Wolffian duct and the central cells of the protovertebre. Waldeyer (Hierstock u. Ei, 1870) has given a totally different account. Between the external border of the protovertebre, and the point where the mesoblast splits into somatopleure and splanchnopleure, there lies a mass of cells, which we shall have occasion to speak of hereafter as the intermediate cell- mass. According to Waldeyer, the upper surface of this mass grows up into a narrow ridge, seen in sections as a tongue-shaped process projecting into the vacant space (z.e. the space filled with fluid only) which exists below the epiblast at this point. Later on, this tongue-shaped process is seen to curve out- wards, and thus to become hook-shaped ; and the point of the hook subsequent:y unites with a similar smaller process derived from the more external portions of the same cell-mass. The small cavity thus seen to be enclosed by a larger and smaller process, is of course the sectional view of a canal enclosed by a larger and smaller ridge. This canal is the Wolffian duct. Waldeyer further believes that the cells which thus form the walls of the duct are primarily epi-. Iv.] THE AMNION. 83 blastic in origin, having been separated from the epiblast at the epoch of the apparent fusion of the. epiblast and mesoblast in the region of the primitive streak or axis-cord of His, This view, prompted as it evidently i is by theoretical considerations, must be regarded as untenable, since the primitive streak has nothing whatever to do with the permanent body of the embryo. Quite recently Romiti (Archiv. Microscop: Anat. x. 1874) has described the Wolffian duct as being formed by an involution of the epithelium of the pleuro- peritoneal cavity, in the form of a longitudinal groove which is thrust up into the superior portions of the intermediate cell-mass, and the communication of which with the pleuroperitoneal cavity is speedily obliterated. Such a mode of origin recommends itself to the embryologist, inasmuch as it is certainly the way in which, as we shall see, the Wolffian duct is formed in Amphibia and Osseous Fishes. For that very reason however it should be received with caution; all the more since the sections drawn by Romiti, and described as supporting his views, evidently belong to a stage considerably later than that at which the duct first distinctly appears. We hope to be able to shew, in the second part of this work, that the mode of development of the Wolffian duct described above, and which we believe to be the real one, is not so abnormal as it might at first sight be supposed to be. 22. The amnion, especially the anterior or head-fold, advances in growth very rapidly during the second day, and at its close completely covers the head and neck of the embryo; so much so that it is necessary to tear or remove it when the head has to be examined in hardened opaque speci- mens. The tail and lateral folds of the amnion, though still progressing, lag considerably behind the head-fold. 23. The chief events then which occur during the second half of the second day are as follows :— 1. The second and third cerebral vesicles make their appearance behind the first. 2. The optic vesicles spring as hollow buds from the lateral, and the vesicles of the cerebral hemispheres from the front portions of the first cerebral vesicle. 3. The first rudiment of the ear is formed as an involu- tion of the epiblast on the side of the hind-brain or third cerebral vesicle. 4. The first indications of the cranial flexure become visible. 5. The head-fold, and especially the splanchnopleure moiety, advances rapidly backwards ; the head of the embryo is In consequence more definitely formed, The tail-fold also becomes distinct. 6. The curvature of the heart increases; the first rudi« ments of the auricles appear. 7. The circulation of the yolk-sac is completed. 8. The amnion grows rapidly. 6—2 CHAPTER V, THE CHANGES WHICH TAKE PLACE DURING THE THIRD DAY, 1. OF all days in the history of the chick within the egg this perhaps is the most eventful ; the rudiments of so many important organs now first make their appearance. On opening an egg on the third day, the first thing which attracts notice is the diminution of the white of the egg. This seems to be one of the consequences of the functional activity of the newly-established vascular area whose blood- vessels are engaged either in directly absorbing the white or, as is more probable, in absorbing the yolk, which is in turn replenished at the expense of the white. The absorption, once begun, goes on so actively that, by the end of the day, the decrease of the white is very striking. 2. The blastoderm has now spread over about half the yolk, the extreme margin of the opaque area reaching about half-way towards the pole of the yolk opposite to the embryo. The vascular area, though still increasing, is much smaller than the total opaque area, being in average-sized eggs about as large as a florin. Still smaller than the vascular area is the pellucid area in the centre of which lies the rapidly growing embryo. 3. During the third day the vascular area is not only a means for providing the embryo with nourishment from the yolk, but also, inasmuch as by the diminutiun of the white it is brought close under the shell and therefore fully exposed to the influence of the atmosphere, serves as the chief organ ef respiration. This in fact is the period at which the vascular area may be said to be in the stage of its most complete development ; CH. V.] THE VASCULAR AREA. 85 for though it will afterwards become larger it will at the same time become less definite and less important. We may therefore, before we proceed, add a few words to the description of it given in the last chapter. DIAGRAM OF THE CIRCULATION OF THE YOLK-SACK AT THE END OF THE H. THirp Day or INCUBATION. heart. AA. the second, third and fourth aortic arches; the first has become obliterated in its median portion, but is continued at its proximal end as the external carotid, and at its distal end as the internal carotid. AQ. dorsal aorta. LZ. Of. A. left omphalo-mesaraic artery. R. Of. A. right omphalo-mesaraic artery. S. 7. sinus terminalis. L. Of. left omphalo-mesaraic vein. 2. Of. right omphalo-mesaraic vein. S. V. sinus venosus. D.C. ductus Cuvieri. S. Ca. V. superior cardinal vein. . V.Ca, inferior cardinal vein. The veins are marked in outline and the arteries are made black. The whole blastoderm has been removed from the egg and is supposed to be viewed from below, Hence the left is seen on the right, and vice versa. 86 _| THE THIRD DAY. [CHAP. The blood leaving the body of the embryo by the omphalo- mesaraic arteries (Fig. 23, R. Of. A., L. Of. A.), is carried to the small vessels and capillaries of the vascular area, a small portion only being appropriated by the pellucid area. From the vascular area part of the blood returns directly to the heart by the main lateral trunks of the omphalo- mesaraic veins, R. Of. L. Of. During the second day these venous trunks joined the body of the embryo considerably in front of, that is, nearer the head than, the corresponding arterial ones. Towards the end of the third day, owing to the continued lengthening of the heart, the veins and arteries run not only parallel to each other, but almost in the same line, the points at which they respectively join and leave the body being nearly at the same distance from the head. According to Von Baer and other observers the veins in the vascular area are placed nearer the surface than are the arteries. Close to the body the reverse is the case; near the body therefore they cross over each other. The rest of the blood brought by the omphalo-mesaraic arteries finds its way into the lateral portions of the sinus terminalis, §.7., and there divides on each side into two streams, Of these, the two which, one on either side, flow backward, meet at a point about opposite to the tail of the embryo, and are conveyed along a distinct vein which, run- ning straight forward parallel to the axis of the embryo, empties itself into the left omphalo-mesaraic vein. The two forward streams reaching the gap in the front part of the sinus terminalis fall into either one, or in some cases two veins, which run straight backward parallel to the axis of the embryo, and so reach the roots of the heart. When one such vein only is present it joins the left omphalo- mesaraic trunk; where there are two they join the left and right omphalo-mesaraic trunks respectively. The left vein is always considerably larger than the right; and the latter when present rapidly gets smaller and speedily disappears. The chief differences then between the peripheral cir- culation of the second and of the third day are due to the greater prominence of the sinus terminalis and the more complete arrangements for returning the blood from it to the heart. After this day, although the vascular area will go on increasing in size until it finally all but encompasses the v.] CHANGE OF POSITION, 87 yolk, the prominence of the sinus terminalis will become less and less in proportion as the respiratcry work of the vascular area is shifted on to the allantois, and its activities confined to absorbing nutritive matter from the yolk. 4. The folding in of the embryo makes great progress during this day. Both head and tail have become most distinct, and the side folds which are to constitute the lateral walls have advanced so rapidly that the embryo is now a bona fide tubular sac, connected with the rest of the yolk by a broad stalk. This stalk, as was explained in Chap. 1, is double, and consists of an inner splanchnic stalk continuous with the alimentary canal, which is now’ a tube closed at both ends and open to the stalk along its middle third only, and an outer somatic stalk continuous with the body-walls of the embryo, which have not closed nearly to the same extent as the walls of the alimentary canal. (Com- pare Fig. 8. A and B, which may be taken as diagrammatic representations of longitudinal and transverse sections of an embryo of this period.) 5. The embryo is almost completely covered by the amnion. Before the close of the day the several amniotic folds will have met along a line over the back of the embryo. Their complete coalescence, and the obliteration of their line of junction, will however not take place till the following day. 6. During this day a most remarkable change takes place in the position of the embryo. Up to this time it has been lying symmetrically upon the yolk with the part which will be its mouth directed straight downwards. It now turns round so as to lie on its left side. This important change of position is almost invariably completed on the third day. At the same time the left omphalo-mesaraic vein, the one on the side on which the embryo comes to lie, grows very much larger than the right, which henceforward gradually dwindles and finally dis- appears. 7. Coincidently with the change of position the whole embryo begins to be curved on itself. This curvature of the body, Fig. 46, becomes still more marked on the fourth day. 8. In the head very important changes take place. One of these is the cranial flexure, Figs. 24,25. This (which 38 THE THIRD DAY, [CHAP. must not be confounded with the curvature of the body just referred to) we have already seen was commenced in the course of the second day, by the bending downwards of the head round a point which may be considered as the extreme end either of the notochord or of the alimentary canal. Fig. 24. al a Ne : i 3. = Ae ae =) / Né CHICK OF THE THIRD Day (34 HOURS) VIEWED FROM UNDERNEATH AS A TRANSPARENT OBJECT. a’. the outer amniotic fold or false amnion. This is very conspicuous around the head, but may also be seen at the tail. a. the true amnion, very closely enveloping the head, and here seen only between the projections of the several cerebral vesicles. It may also be traced at the tail, ¢. In the embryo of which this is a drawing the head-fold of the amnion reached a little farther backward than the reference u, but its limit could not be v.] THE CRANIAL FLEXURE. 89 distinctly seen through the body of the embryo. The prominence of the false amnion at the head is apt to puzzle the student; but if he bears in mind the fact, which could not well be shewn in Fig. 8, that the whole amniotic fold, both the true and the false limb, is tucked in underneath the head, the matter will on reflection become intelligible. C. H. cerebral hemisphere. /. B. fore-brain or vesicle of the third ventricle. M. B. mid-brain. H. B.hind-brain. Op. optic vesicle. Ot. otic vesicle. OfV. omphalo-mesaraic veins forming the venous roots of the heart. The trunk on the right hand (left trunk when the embryo is viewed in its natural position from above) receives a large branch, shewn by dotted lines, coming from the anterior portion of the sinus terminalis Ht. the heart, now com- pletely twisted on itself. Ao. the bulbus arteriosus, the three aortic arches being dimly seen stretching from it across the throat, and uniting into the aorta, stil more dimly seen as a curved dark line running along the body. The other curved dark line by its side, ending near the reference y, is the notochord ch. About oppusite the line of reference x the aorta divides into two trunks, which running in the line of the somewhat opaque protovertebre on either side, are not clearly seen. Their branches however, Ofa, the omphalo-mesaraic arteries, are conspicuous and are seen to curve round the commencing side folds. Py, protovertebre. Below the level of the omphalo-mesaraic arteries the vertebral plates are but imperfectly cut up into protovertebre, and lower down still, not at all. x is placed at the ‘‘point of divergence” of the splanchnopleure folds. The blind foregut begins here and extends about up to y, the more transparent space marked by that letter being partly due to the presence there of the cavity of the alimentary canal. « therefore marks the present hind limit of the splanchnopleure folds. The limit of the more transparent somato- pleure folds cannot be seen. Té will be of course understood that all the body of the embryo above the level of the reference x, is seen through the portion of the yolk-sac (vascular and pellucid area), which has been removed with the embryo from the egg, as well as through the double amniotic fold. We may repeat that, the view being from below, whatever is described in the natural position as being to the right here appears to be left, and vice versa. The flexure progresses rapidly, the front-brain being more and more folded down till, at the end of the third day, it is no longer the first vesicle or fore-brain, but the second cerebral vesicle or mid-brain, which occupies the extreme front of the long axis of the embryo. In fact a straight line through the long axis of the embryo would now pass through the mid-brain instead of, as at the beginning of the second day, through the fore-brain, so completely has the front end of the neural canal been folded over the end of the notochord. The commencenient of this cranial flexure gives the body of an embryo of the third day somewhat the appearance of a retort, the head of the embryo corresponding 90 THE THIRD DAY. [CHAP. to the bulb. On the fourth day the flexure is still greater than on the third, but on the fifth and succeeding days it becomes less obvious owing to the filling up of the parts of the skull. Fic. 25. J Assi 3 Si! 4 ‘| FE HEAD oF A CHICK OF THE THIRD DAY VIEWED SIDEWAYS AS A TRANSPARENT OxpsEoT. (From Huxley.) I. a. the vesicle of the cerebral hemisphere. I. }. the vesicle of the third ventricle (the original fore-brain); at its summit is seen the projection of the pineal gland e. Below this portion of the brain is seen, in optical section, the optic vesicle a already involuted with its thick inner and thinner outer wall (the letter a is placed on the junction of the two, the primary cavity being aimost obliterated). In the centre of the vesicle lies the lens, the shaded portion (represented too large), being the expression of its cavity. Below the lens between the two limbs of the horse-shoe is the choroidal fissure. II. the mid-brain, now, owing to the cranial flexure, opposite the end of the alimentary canal. III. the hind-brain. V. the rudiments of the fifth cranial nerve, VII. of the seventh. Below the seventh nerve is seen the auditory vesicle 6. The head having been subjected to pressure, the vesicle appears somewhat distorted as if squeezed out of place. The orifice is not yet quite closed up. 1. the inferior maxillary process of the first visceral fold. Below, and to the right of this, is seen the first visceral cleft, below that again the second visceral fold (2), and lower down the third (3) and fourth (4) visceral folds. In front of the folds (i.e. to the left) is seen the arterial end of the heart, the aortic arches being buried in their respective visceral folds. J: represents at its lowest part the cavity of the alimentary canal as seen through the transparent body of the embryo; at the upper part below the brain it is difficult to distinguish between the transparency due to the presence of the cavity of the alimentary canal, and that caused by the character of the meso- blast at the base of the skull, which, being formed of stellate cells with largely developed clear spaces or vacuoles, allows the light to pass readily through it. Near its upper end below the mid-brain is seen a small conical process, the rudiment of the infundibulum. 9. The two vesicles of the cerebral hemispheres which v.] THE FORE-BRAIN. 91 on the second day began to grow out from the front of the fore-brain, increase rapidly in size during the third day, so much so that by the end of the day each of them (Fig. 24, CH, Fig. 25, Ia) is as large or larger than the original fore-brain from which they both sprang, and they form together a most conspicuous part of the brain. In their growth they push aside the optic vesicles, and thus contribute largely to the roundness which the head is now acquiring. Each vesicle possesses a cavity, known afterwards as a lateral ventricle, which, though quite separate from its fellow, is continuous with the cavity of the fore-brain. Owing to the development of these cerebral hemispheres, the original fore-brain no longer occupies the front position (Fig. 24, FB, Fig. 25, Id), and ceases to be the conspicuous object that it was. Inasmuch as its walls will hereafter be de- veloped into the parts surrounding the so-called third ventricle of the brain, we shall henceforward speak of it as the vesicle of the third ventricle, or, inasmuch as it soon comes to lie between the expanded posterior ends of the cerebral hemi- spheres, as the tween brain. On the summit of the tween brain there may now be seen a small conical projection, the rudiment of the pineal gland (Fig. 25, e), while the ceutre of the floor is produced into a funnel-shaped process, the infundibulum (Fig. 22, d), which, stretching towards the extreme end of the alimentary canal, joins the pituitary body. The development of the pituitary body or hypophysis cerebri has been the subject of considerable controversy amongst embryologists. Von Baer (loc. cit.) and Smidt (Zeitschrift fiir Wiss. Zoologie, 1862, B. X1, p. 43) believed that the base of the fore-brain, or vesicle of the third ventricle, became produced into a downward process, the ‘infundibulum,’ which subsequently became expanded at its termination to form the pituitary body. Rathke (Archiv fiir Anatomie und Physiologie 1838, Bd. v.) states that very early a diverticulum is produced from the upper end of the alimentary canal, which passes backwards and meets the process of the brain called the infundibulum. This diverticulum subsequently loses all connection with the epithelium of the digestive canal, and, uniting with the infundibulum, forms the pituitary body. Dursy (Entwicklungsgeschichte des Kopfes, Tiibingen, 1869) states that both the end of the notochord and the epithelium of the alimentary canal take part in the formation of the pituitary body. The apparent diverticulum of the ali- mentary canal is not so much a true diverticulum, as a part of the alimentary canal constricted off from the remainder by the cranial flexure. Reichert (Entwicklungsleben im Wirbelthierreich. Berlin, 1840) states that the pituitary body is formed from the remains of the front end of notochord. Se THE THIRD DAY. [CHAP. Subsequently however (Der Bau des menschlichen Gehirns) he supposed that it was formed from the pia mater. Rathke also subsequently (Entwicklunsgeschichte der Wirbelthiere, Leipzig, 1861) gave up bis former view, and believed that the diverticulum of the alimentary canal disappeared, but that the pituitary body was formed from the mesoblast in front of the clinoid process. Wilhelm Miiller (Ueber die Entwicklung und Bau der Hypophysis und des Processus infundibuli cerebri. Jenaische Zeitschrift, Bd. vt. 1871) has recently written an elaburate memoir on the deve‘opment and anatomy of the pituitary body and infundibulum in all the orders of Vertebrates, of which the following is an abstract. In order to understand the formation of the diverticulum from the ali- mentary canal which forms the pituitary body, we must remember that at first the hypoblast of the throat closely underlies the notochord, and beyond the end of the notochord is almost in contact with the base of the vesicle of the third ventricle. When the cranial flexure occurs, which it will be remembered takes place about an axis coinciding with the end of the notochord, the hypoblast, which closely underlies the base of the brain, becomes at the same time bent; and as the angle of the flexure becomes an acute angle, a wedge- shaped space lined by hypoblast is as it were constricted off from the alimen- tary canal. In this way there is formed a diverticulum of hypoblast which passes forwards from the alimentary canal to the base of the fore-brain— a diverticulum not produced by a torward growth from the alimentary canal, but solely due to the cranial flexure constricting off a wedge-shaped portion of the alimentary canal. This we may call the pituitary diverticulum. When the cranial flexure commences the end of the notochord becomes bent down- ward, and, ending in a somewhat enlarged extremity, comes in contact with the termination of the pituitary diverticulum. The mesoblast around and at the front of the end of the notochord increases and grows up, in frout of the notochord and behind tle vesicle of the third ventricle, to form the posterior clinoid process. The base of the vesicle of the third ventricle at the same time grows downwards towards the pituitary diverticulum and forms what is kuown as the infundibulum. This state of things may be observed on the third day. On the fourth day the mesoblast tissue around the notochord increases in quantity, and the end of the nutochord, though still bent down- wards, recedes a little from the termination of the pituitary diverticulum, which is still a triangular space with a wide opening into the alimentary canal. On the fifth day, the opening of the pituitary diverticulum into the alimentary canal has become narrowed, and around the whole diverticulum a formation of mesoblast-cells has commenced. Behind it the clinoid process has become cartilaginous, while to the side and in front it is enclosed by the trabecule. At this stage, in fact, we have a diverticulum from the alimentary canal passing through the base of skull to the infundibulum. The end of the notochord has at this stage become atrophied, so that it is separated by a considerable interval from the pituitary body. On the seventh day the mesoblast around the pituitary diverticulum has grown into a complete investment of spindle-shaped cells, and the communi- cation between the cavity of the diverticulum and that of the throat has become stillnarrower. The diverticulum is all but converted into a vesicle, and its hypo- blast walls have commenced to send out into the mesoblastic investment solid processes, which form the first commencement of the true pituitary body. The infundibulum now appears as a narrow process from the base of the vesicle of the third ventricle, which approaches, but doves not unite with the pituitary vesicle. This latter lies in the space between the basi- and the presphenoid, and is v.] THE MID-BRAIN AND HIND-BRAIN. 93 completely surrounded by a ring of cartilage. The mesoblast-cells immediately around it do not, however, exhibit any signs of becoming cartilaginous. By the tenth day the opening of the pituitary vesicle into the threat becomes almost obliterated, and the lumen of the vesicle itself very much diminished. The body itself consists of anastomosing cords of hyp ‘blast-cells, the meso- blast between which has already commenced to become vascular. The cords or masses of hypoblast cells are surrounded by a delicate membrana propria, ’ and a few of them possess a small lumen. The infundibulum has increased in length. ‘On the twelfth day the communication between the pituitary vesicle and the throat is entirely obliterated, but a solid cord of cells still connects the two. The vessels of the pia mater of the vesicle of the third ventricle have become connected with the pituitary body, and the infundibulum has grown down along its posterior border. In the tater stages, all connection is lust between the pituitary body and the throat, and the former becomes connected with the elongated processus infundibuli. Such is Wilhelm Miiller’s account, Goette, however (Archiv. Micr. Anat. IX. p. 397), has recently given reasons for thinking that the pituitary diverti- culum arises not from the closed foregut, lined with hypoblast, but from the buceal cavity lined with epiblast. He states that in its earlier stages it may be seen to start on the oral side of the partition, which for some time divides the secondarily formed buccal cavity from the primarily formed foregut, and therefore, belonging to the former, cannot be regarded as the natural anterior termination of the latter. Beyond an increase in size, which it shares with nearly all parts of the embryo, and the change of position to which we have already referred, the mid-brain undergoes no great alteration during the third day. Jts roof will ultimately become developed into the corpora bigemina or optic lobes (quadrigemina in mammals), its floor will form the crura cerebri, and its cavity will be reduced to the narrow canal known as the tter a tertio ad quartum ventriculum. In the hind-brain, or third cerebral vesicle, that part which lies nearest to the mid-brain, becomes during the third day marked off from the rest by a slight constric- tion. This distinction, which becomes much more evident later on by a thickening of the walls and roof of the front portion, separates the hind-brain into the cerebellum in front, and the medulla oblongata behind. While the walls of the cerebellar portion of the hind-brain become very much thickened as well at the roof as at the floor and sides, the roof of the posterior or medulla oblongata portion thins out into a mere membrane, forming a delicate covering to the cavity of the vesicle (Fig. 26. Iv), which here becoming broad and shallow with greatly thickened floor and sides, is 94 THE THIRD DAY. [CHAP. known as the fourth ventricle, subsequently overhung by the largely developed posterior portion of the cerebellum. The third day, therefore, marks the distinct differentiation of the brain into its fundamental parts: the cerebral hemi- spheres, the central masses round the third ventricle, the corpora bigemina, the cerebellum and the medulla oblongata; the original cavity of the neural canal at the same time, passing from its temporary division of three single cavities into the permanent arrangement of a series of connected ventricles, viz. the lateral ventricles, the third ventricle, the iter (with a prolongation into the optic lobe on each side), and the fourth ventricle. 10. At the same time that the outward external shape of the brain is thus being moulded, internal changes are taking place in the whole neural canal. These are best seen in sections. At its first formation, the section of the cavity of the neural canal is round or nearly so. About this time, however, the lining of involuted epiblast along the length of the whole spinal cord becomes very much thickened at either side, while increasing but little at the mid-points above and below. The result of this is that the cavity as seen in section (Fig. 44), instead of being circular, has become a narrow vertical slit, almost completely filled in on either side. In the region of the brain the thickening of the lining epiblast follows a somewhat different course. While almost everywhere the sides and floor of the canal are greatly thick- ened, the roof in the region of the various ventricles, not of the fourth only, but of the others as well, becomes excessively thin, so as to form a membrane reduced to almost a single layer of cells. (Fig. 26. Iv.) 11. In the preceding chapter we saw how the first cere- bral vesicle, by means of lateral outgrowths followed by constrictions, gave rise to the optic vesicles. These and the parts surrounding them undergo on the third day changes which result in the formation of the eyeball. At their first appearance the optic vesicles stand out at nearly right angles to the long axis of the embryo (Fig. 15), and the stalks which connect them with the fore-brain are short and wide. We have already said (p. 77) that the con- v.] THE OPTIC VESICLES. 95 strictions which give rise to the stalks take place chiefly from above downwards, and also somewhat inwards and backwards, Thus from the first the vesicles appear to spring from the under part of the fore-brain. Fic. 26. SECTION THROUGH THE HIND-BRAIN OF A CHICK AT THE END OF THE THIRD Day oF INCUBATION. IV. Fourth ventricle. The section shews the very thin roof and thicker sides of the ventricle. Ch. Notochord—(diagrammatic shading). CV. Anterior cardinal vein. CG. Involuted auditory vesicle. CC points to the end which will form the cochlear canal. RZ. Recessus labyrinthi. hy. hypoblast lining the alimen- tary canal. hy is itself placed in the cavity of the alimentary canal, in that part of the canal which will become the throat. The lower (anterior) wall of the canal is not shewn in the section, but on each side are seen portions of a pair of visceral arches. In each arch is seen the section of the aortic arch AOA belonging to the visceral arch. The vessel thus cut through is running upwards towards the head, being about to join the dorsal aorta AO. Had the section been nearer the head, and carried through the plane at which the aortic arch curves round the alimentary canal to reach the mesoblast above it, AOA and AO would have formed one continuous curved space. In sections lower down in the back the-two aorta, AQ, one on either side would be found fused into one median canal. The shading of the mesoblast is diagrammatic; it is here a uniform mass of spindle-shaped cells, there being in this region no differentiation into proto-. vertebra. 96 THE THIRD DAY. [CHAP, As the vesicles of the cerebral hemispheres grow out rapidly from the front and under portions of the first cerebral vesicle, they seem to thrust the optic vesicles apart and to the sides. Thus these, instead of standing out from the extreme front, come to be placed at the sides of the head, the stalks, which are correspondingly lengthened and narrowed, running obliquely downwards and inwards from the vesicles to open into the cavity of the brain at its base. Their openings are at first placed close to each other at the junction of the cerebral hemispheres with the remnant of the fore-brain (now called the vesicle of the third ventricle), so that the cavities of the two optic vesicles may be said to communicate directly both with each other and with the cavities of the cerebral hemispheres. The later connection is however soon lost, and the stalks of the optic vesicles then open exclusively into the third ventricle. At the same time the floor of the third ven- tricle, during the occurrence of the cranial flexure, grows down and thrusts apart the openings of the two optic stalks. At a later date the stalks shift their position backwards, and thus become connected chiefly with the base of the mid- brain. While these changes have been going on in the optic stalks, development has also proceeded in the region of the vesicles themselves, and given rise to the rudiments of the retina, lens, vitreous humour, and other parts of the eye. The changes through which these are formed are of a somewhat complicated character, and not a few points in reference to them are still involved in some doubt. Towards the end of the second day, the external or super- ficial epiblast which covers, and is in all but immediate contact with the most projecting portion of the optic vesicle, becomes thickened. This thickened portion is then driven inwards in the form of a shallow open pit with thick walls (Fig. 27 A. x), carrying before it the front wall (r) of the optic vesicle. To such an extent does this involution of the super- ficial epiblast take place, that the front wall of the optic vesicle is pushed close up to the hind wall, and the cavity of the vesicle becomes almost obliterated (Fig. 27, B). The bulb of the optic vesicle is thus converted into a cup with double walls, containing in its cavity the portion of v.] THE OPTIC CUP. oF involuted epiblast. This cup, in order to distinguish its cavity from that of the original optic vesicle, is generally called the secondary optic vesicle. We may, for the sake of brevity, speak of it as the optic cup; in reality it never is a vesicle, since it always remains widely open in front. Of its double walls the inner or anterior (Fig. 27 B, r) is formed from the front portion, the outer or posterior (Fig. 27 £B, u) from the hind portion of the wall of the primary optic vesicle. The inner or anterior (7), which very speedily becomes thicker than the other, is converted into the retina; in the outer or posterior (w), which remains thin, pigment is eventually deposited, and it ultimately becomes the tesselated pigment-layer of the choroid. Fie. 27. As B. DIAGRAMMATIC SECTIONS ILLUSTRATING THE FORMATION OF THE EYE. (After Remak.) In A, the thin superficial epiblast 4 is seen to be thickened at x, in front of the optic vesicle, and involuted so as to form a pit 0, the mouth of which has already begun to close in. Owing to this involution, which forms the 1udiment of the lens, the optic vesicle is doubled in, its front portion 7 being pushed against the back portion u, and the original cavity of the vesicle thus reduced in size. The stalk of the vesicle is shewn as still broad. In B, the optic vesicle is still further doubled in so as to form a cup with a posterior wall w and an anterior wall 7. In the hollow of this cup lies the lens /, now completely detached from the superficial epiblast «, hk. Its cavity is still shewn. ‘The cavity of the stalk of the optic vesicle is already much narrowed. By the closure of its mouth the pit of involuted epiblast becomes a completely closed sac with thick walls and a small central cavity. (Fig. 27 Bl). At the same time it breaks away from the external epiblast, which forms a continuous layer in front of it, all traces of the original opening being lost. There is thus left lying in the cup of the secondary optic vesicle, an isolated elliptical mass of epiblast. This is E. 4 98 THE THIRD DAY. [CHAP. the rudiment of the lens. The small cavity within it speedily becomes still less by the thickening of the walls, especially of the hinder one. At its first appearance the lens is in immediate contact with the anterior wall of the secondary optic vesicle (Fig. 27 B). In a short time however, the lens is seen to lie in the mouth of the cup (Fig. 30 D), a space (vh) (which is subsequently occupied by the vitreous humour) making its appearance between the lens and anterior wall of the vesicle. In order to understand how this space is developed, the position of the optic vesicle and the relations of its stalk must be borne in mind. Fie. 28, Gry Sf D DIAGRAMMATIC SECTION OF THE EYE AND THE Optic NERVE AT AN EARLY STAGE (from Lieberkiihn), to shew the lens 7 occupying the whole hollow of the optic cup, the inclination of the stalk s to the optic cup, and the continuity of the cavity of the stalk s’ with that of the primary vesiclec; 1, anterior, w posterior wall of the optic cup. The vesicle lies at the side of the head, and its stalk is directed downwards, inwards and backwards. The stalk in fact slants away from the vesicle. Hence when the involu- tion of the lens takes place, the direction in which the front wall of the vesicle is pushed in is not in a line with the axis ot the stalk as for simplicity’s sake has been represented in the diagram Fig. 27, but forms an obtuse angle with that axis, after the manner of Fig. 28, where s’ represents the cavity of the stalk leading away from the almost obliterated cavity of the primary vesicle. Fig. 28 represents the early stage at which the lens fills the whole cup of the secondary vesicle. The subsequent state of affairs is brought about through the growth of the v.] THE CHOROIDAL FISSURE. 99 walls of the cup taking place more rapidly than that of the lens, or in other words to the cavity of the cup dilating. But this growth or this dilatation does not take place equally im all parts of the cup. The walls of the cup rise up all round except that poimt of the circumference of the cup which is opposite the middle (from side to side) of the stalk. While elsewhere the walls increase rapidly in height, carrying so to speak the lens with them, at this spot, which in the natural position of the eye is on its under surface, there is no growth: the wall is here imperfect, and a gap is left. Through this gap, which afterwards receives the name of the choroidal fissure, 2 way is open from the mesoblastic tissue surrounding the optic vesicle and stalk into the interior of the cavity of the cup. DIAGRAMMATIC REPRESENTATION OF THE EYE OF THE (HICK OF ABOUT THE THIRD DAY AS SEEN WHEN THE HEAD IS VIEWED FROM UNDERNEATH AS A TRANSPARENT OBJECT. 1 the lens, /’ the cavity of the lens, lying in the hollow of the optic cup. v the anterior, ~ the posterior wall of the optic cup, ¢ the cavity of the primary optic vesicle, now nearly obliterated. By inadvertence w has been drawn in some places thicker than 7, it should have been thinner through- out. $ the stalk of the optic cup with s’ its cavity, at a lower level than the cup itself and therefore out of focus; the dotted line indicates the continuity of the cavity of the stalk with that of the primary vesicle. The line z, z, through which the section shewn in Fig. 30 F is supposed to be taken, passes through the choroidal fissure. From the manner of its formation the gap or fissure is evidently in a line with the axis of the optic stalk, and in id —_ . = 100 THE THIRD DAY. . [CHAP. order to be seen must be looked for on the under surface of the optic vesicle. In this position it is readily recog- nized in the transparent embryo of the third day, Figs. 25 and 29. ; Bearing in mind these relations of the gap to the optic stalk, the reader will understand how sections of the optic vesicle at this stage present very different appearances according to the plane in which the sections are taken. When the head of the chick is viewed from underneath as a transparent object the eye presents very much the ap- pearance represented in the diagram Fig. 29. D. Diagrammatic section taken perpendicular to the plane of the paper, along the line y, y, Fig. 29. ‘The stalk is not seen, the section falling quite out of its region. vh, hollow of optic cup filled with vitreous humour ; other letters as in Fig. 27 B. E. Section taken parallel to the plave of paper through Fig. 29, so far behind the front surface of the eye as to shave off a small portion of the posterior surface of the lens J, but so far in front as not to be carried at all through the stalk. Letters as before; f, the choroidal fissure. F. Section along the line z, z, perpendicular to the plane of the paper, to shew the choroidal fissure f, and the continuity of the cavity of the optic stalk with that of the primary optic vesicle. Had this section been taken a little to one side of the line z, z, the wall of the optic cup would have extended up to the lens below as well as above. Letters as before. A section of such an eye taken along the line y, per- pendicular to the plane of the paper, would give a figure corresponding to that of Fig, 30 D. The lens, the cavity and double walls of the secondary vesicle, the remains of the primary cavity, would all be represented (the superficial epiblast of the head would also be shewn); but there would be nothing seen of either the stalk or the fissure. If on the other hand the section were taken in a plane parallel to the plane of the paper, at some distance above the v.] THE. CHOROIDAL FISSURE. 101 level of the stalk, some such figure would be gained as that shewn in Fig. 30 E. Here the fissure f is obvious, and the communication of the cavity wh of the secondary vesicle with the outside of the eye evident; the section of course would not go through the superficial epiblast. Lastly, a section, taken perpendicular to the plane of the paper along the line z, v.é. through the fissure itself, would present the ap- pearances of Fig. 30 F, where the wall of the vesicle is entirely wanting in the region of the fissure marked by the position of the letter fA The fissure such as we have described it exists for a short time only. Its lips come into contact, and unite (in the neighbourhood of the lens, directly, but in the neighbourhood of the stalk, by the intervention of a structure which we shall describe presently), and thus the cup-like cavity of the secondary optic vesicle is furnished with a complete wall all round. The interior of the cavity is filled by the vitreous humour, a clear fluid in which are a few scat- tered cells. In the foregoing account of the formation of the secondary optic vesicle, and of the fissure, as the results of a process of unequal growth, we have fol- lowed the account of Lieberkiihn (Uber das Auge des Wirbelthierembryos, Schriften der Geseilschaft zur Betérderung der gesammten Naturwissenschaiten zu Marburg. Bd. to, 1872). Their origin is more generally described as being due to a doubling up of the primary vesicle from the side along the line of the fissure at the same time that the lens is being thrust inin front. In mammalia, the doubling up is said to involve the optic stalk, which becomes flattened (whereby its original cavity is obliterated) and then folded in on itself, so as to embrace a new ceutral cavity continuous with the cavity of the vitreous humour. According to Lieberkiihn the optic stalk in birds is never so folded up, but is converted into the optic nerve by the gradual obliteration of its primary central cavity through increased thickening of the walls. The optic nerve of the bird, moreover, contains no arteria centralis retin, while the involu- tion of the optic stalk into the optic nerve was supposed to have for its purpose the introduction of a quantity of mesoblast into the interior of nerve, in order to form the artery. According to Remak and the majority of observers after him, no mesoblast whatever exists between the external epiblast and the optic vesicle, at the point where the former is thrust inwards to form the lens, and hence this organ carries with it in its involution no mesoblast whatever to serve as a rudiment of either the vitreous humour or the capsule of the lens. They described the vitreous humour as being formed entirely out of the meso- blast which was intruded from the exterior of the eye through the choroidal fissure, and Kdlliker considered the capsule of the lens as a sort of cuticular excretion from the surface of the lens itself. Lieberkiibn on the other hand states that shortly after the commencement of the involution of the lens there 102 THE THIRD DAY. [ CHAP. may be already found a thin layer of mesoblast, interposed between it and the optic vesicle. This layer is carried inward during the involution, and from it both the vitreous humour and the capsule of the lens take their origin. Jn birds it is very difficult to be sure of the existence of this layer, though Lieberkiihn says that in mammals it is conspicuous ; and even if its existence be admitted, it still remains doubtful whether it gives rise to the whole vi- treous humour, or to the capsule of the lens only; though the latter view is most probable. During the changes in the optic vesicle just described, the surrounding mesoblast takes on the characters of a distinct investment, whereby the outline of the eyeball is definitely formed. The internal portions of this investment, nearest to the retina, become the choroid (i.e. the chorio-capillaris, and the lamina fusca, the pigment epithelium, as we have seen, being derived from the epiblastic optic cup), and pig- ment is subsequently deposited in it. The remaining external portion of the investment forms the sclerotic. The complete differentiation of these two coats of the eye does not however take place till a late period. Along the line of the choroidal fissure the pigment is wanting. Con- sequently in embryos of an age when the pigment has be- come generally deposited in the choroid, a colourless streak marking out the position of the choroidal fissure is very con- spicuous. In front of the optic cup the mesoblastic investment grows forwards, between the lens and the superficial epi- blast, aud so gives rise to the substance of the cornea; the epiblast supplying only the anterior epithelium. At first the whole space between the lens and the super- ficial epiblast is occupied by undifferentiated mesoblast; but on the sixth day a layer of epithelium makes its ap- pearance in midst of the mass, and thus divides it into an anterior and a posterior portion. The anterior portion, in- creasing in solidity, becomes the cornea, and remains con- tinuous with the sclerotic; the epithelium in question per- sisting as the posterior epithelium of the membrane of Descemet. The posterior portion is reduced to a mere membrane forming, according to Lieberkiihn, the front limb of the capsule (and the suspensory ligament) of the lens, the space between it and the cornea becoming filled with aqueous humour. We left the original cavity of the primary optic vesicle as | THE OPTIC CUP. 105 a nearly obliterated space between the two walls of the optic cup. By the end of the third day the obliteration is com- plete, and the two walls are in immediate contact. The inner or anterior wall is, from the first, thicker than the outer or posterior; and over the greater part of the cup this contrast increases with the growth of the eye, the anterior wall becoming markedly thicker and undergoing changes of which we shall have to speak directly (Fig. 31). In the front portion however, along, so to speak, the lip of the cup, anterior to a line which afterwards becomes the ora serrata, both layers not only cease to take part in the increased thickening, accompanied by peculiar histo- logical changes, which the rest of the cup is undergoing, but also completely coalesce together. Thus a hind portion or true retina is marked off from a front portion. The front portion, accompanied by the choroid which immediately overlays it, is behind the lens thrown into folds, the ciliary ridges; while further forward it bends in between the lens and the cornea to form the iris. The original wide opening of the optic cup is thus narrowed to a smaller orifice, the pupil; and the lens, which before lay in the open mouth of, is now inclosed in the cavity of the cup. While in the hind portion of the cup or retina proper, no deposit of black pigment takes place in the layer formed out of the inner or anterior wall of the vesicle, in the front portion we are speaking of, pigment is largely deposited throughout both layers, so that eventually this portion seems to become nothing more than a forward prolongation of the pigment- epithelium of the choroid. Thus while the hind moiety of the optic cup becomes the retina proper, including the choroid-pigment in which the rods and cones are imbedded, the front moiety is converted into the ciliary portion of the retina, covering the ciliary processes, and into the uvea of the iris; the bodies of the ciliary processes and the substance of the iris, their vessels, muscles, connective tissue and ramified pigment, being derived from the mesoblastic choroid. The margin of the pupil marks the extreme lip of the optic vesicle, where the outer or poste- rior wall turns round to join the inner or anterior. We have stiil to speak of the choroidal fissure. In mammals the slit remains open for a short time only. After 104 THE THIRD DAY. [CHAP, Fie. 31. 2 \ SECTION OF THE EYE OF CHICK AT THE FourtH Day, e. p. superficial epiblast of the side of the head. R. true retina: anterior wall of the optic eup. p. Ch. pigment-epithelium of the choroid: posterior wall of the optic cup. 6 is placed at the extreme lip of the optic cup at what will become the margin of the pupil. l, the lens. The hind wall, the nuclei of whose elongated cells are shewn at nl, now forms nearly the whole mass of the lens, the front wall being reduced to a layer of flattened cells el. m, the mesoblast surrounding the optic cup and about to form the choroid and sclerotic. It is seen to pass forward between the lip of the optic cup and the superficial epiblast. Filling up a large part of the hollow of the optic cup is seen a hyaline mass vh, possibly the rudiment of the vitreous humour. It has fallen away from the retina at #, and is also (apparently accidentally) wanting at y. In the neighbourhood of the lens it seems to be continuous as at cl with the tissue a, which in turn is continuous with the mesoblast m, and appears to be the rudiment of the capsule of the lens and suspensory ligament, v.] THE RETINA. 105 the formation of the vitreous humour within the cup, the edges of the slit grow completely together, and all traces of the seam disappear. In birds the course of events is some- what different. In addition to such amount of mesoblast as may pass through the slit to form the vitreous humour, two special processes of mesoblast grow in, one in the neighbourhood of the optic stalk, in the region of the true retina, and a second, which speedily becomes highly vascular, in that portion of the slit which corresponds to the ciliary part of the retina. The former process remains as the pecten so characteristic of the avian eye, while the latter vascular process serves to supply the pecten with blood. By the twelfth day the fissure completely closes up and disappears between these two processes and also in front of the vascular one; but both the pecten and the vascular process are left projecting into the interior of the eye. Hence in the adult eye, the pecten seems to perforate the retina close to the entrance of the optic nerve, the nervous fibres of the retina spreading away in a radiate manner from it. The optic stalk, which, as we have said, by an obliteration of its central canal becomes converted into the optic nerve, is at first equally continuous with the inner and with the outer wall of the retina, This must of necessity be the case, since the interval which primarily exists between the two walls is continuous with the cavity of the stalk (vide Figs. 28 and 30 F, s’). When the fibres however make their appear- ance in the nerve, they are found to be connected with the inner wall, or functional retina, only. The histological condition of the eye in its earliest stages is very simple.. Both the epiblast forming the walls of the optic vesicle, and the superficial layer which is thickened to become the lens, are composed of several layers of simple columnar cells. The surrounding: mesoblast is made up of cells whose protoplasm is more or less branched and irregu- lar. These simpie elements are gradually modified into the complicated tissues of the adult eye, the changes undergone being most marked in the cases of the retina, the optic nerve, and the lens with its appendages. The retina. At first the two walls of the optic cup do not greatly ditfer in thickness. On the third day the outer or 106 THE THIRD DAY. [ CHAP. posterior becomes much thinner than the inner or anterior, and by the middle of the fourth day is reduced to a single layer of flattened cells (Fig. 31, p. Ch.). At about the 80th hour its cells commence to receive a deposit of pigment, and eventually form the so-called pigmentary epithelium of the choroid; from them no part of the true retina (or no other part of the retina, if the pigment-layer in question be sup- posed to belong more truly to the retina than to the choroid) is derived. On the fourth day, the inner (anterior) wall of the optic cup (Fig. 31, &) is perfectly uniform in structure, being composed of elongated somewhat spindle-shaped cells, with distinct nuclei. On its external (posterior) surface a distinct cuticular membrane, the membrana limitans externa, early appears. As the wall increases in thickness, these cells multiply rapidly, so that the wall becomes several cells thick. The first indications of a division into layers are noticed on the seventh day; and on the eighth day a layer of ‘granules’ is very obvious. The granules, which are appar- ently nuclei of cells, become on the tenth day distinctly arranged into an inner and an outer layer; and at about the same time small processes, apparently outgrowths from the outer granular layer, make their appearance on the external surface of the membrana limitans externa. These processes are the rudiments of the rods and cones. From the first they may be roughly divided into two categories, (1) those of smaller, (2) those of larger diameter. Both kinds grow rapidly and in the tips of both small highly refractive globules soon appear. The thinner processes are the cones, the thicker the rods. The cones remain for a long time thinner than the rods, but shortly before the exclusion of the chick they increase rapidly in diameter and soon after that occurrence are found to surpass the rods in thickness. On the 18th day some of the globules in the cones become red, on the roth others become yellow, and very soon all the globules in the cones acquire a distinct colour. The globules in the rods remain uncoloured. The rods and cones then are outgrowths through the membrana limitans externa, from the inner wall of the optic cup or retina into the outer wall or pigment- epithelium of the choroid. : Remak and some other investigators were of opinion that the outer wall of the optic cup gave rise to the rods and cones as well as to the pigment-epi- thelium. The observations however of Max Schultze, Archiv Micros. Anat Iv. p- 239, supported by Babuchin, Wiirz. Nat. Zeitsch. tv. (1863) p. 71, and others, have clearly shewn that Remak’s views were erroneous. On the thirteenth day the molecular layer and the gan- v.] THE OPTIC NERVE. 107 elionic layer are distinguishable. Very early the substance of certain of the cells takes on the appearance of fibres, arranged vertically, 7.e. radiating from the inner or anterior surface of the retina to the membrana limitans externa. These are the rudiments of the fibres of Miiller. Thus of the cells of the inner wall of the cup, some become ganglionic cells, and others the fibres of Miiller, while the nuclei of yet others remain as the inner and outer granules. The rods and cones are outgrowths of the cells to which the outer granules belong. All parts of the retina, in fact, whether simply connective, or really nervous in nature, seem to be derived from epiblastic cells. The changes described above are confined to that portion of the retina which lies behind the ora serrata. In front of this both walls of the cup coalesce as we have said into a cellular layer in which a deposit of pigment takes place. The optic nerve. Histological changes are first observable in the optic stalk at about the time when its cavity loses all connection with the cerebral hemisphere and opens ex- clusively into the third ventricle. It is then that fibres first make their appearance in its walls, nuclei being still abun- dantly present. The stalk though much elongated is still hollow and its cavity is circular in section. According to Lieberkiihn at no time does it (in the bird) undergo any involution tending to obliterate its cavity. Soon after the deposition of pigment in the outer wall of the optic cup, while the optic stalks are as yet still hollow, the rudiments of the optic chiasma appear. The fibres of the one stalk grow over into the attachment of the other. About the same time the fibres at the neck of the optic cup grow forwardand become connected with the retina,over whose internal surface they spread. The entrance of the optic nerve into the eyeball is closely connected with that of the pecten, its fibres passing in at the lower end of that body, coursing along its sides to its upper end and radiating from it as from a centre to all parts of the retina. Before the exclusion of the chick the optic nerve becomes solid by the gradual filling up of its central cavity. The lens when first formed is somewhat elliptical in section with a small central cavity of a similar shape, the front and hind walls being of nearly equal thickness, 108 THE THIRD DAY. [CHAP. each consisting of a single layer of elongated columnar cells. In the subsequent growth of the lens, the development of the hind wall is of a precisely opposite character to that of the front wall. The hind wall becomes much thicker, and tends to obliterate the central cavity by becoming convex on its front surface. At the same time its cells, still remaining as a single layer, become elongated and fibre-like. The front wall on the contrary becomes thinner and thinner and its cells more and more flattened and pavement-like. These modes of growth continue until at the end of the fourth day, as shewn in Fig. 31, the central hind wall / is in absolute contact with the front wall ed and the cavity thus becomes entirely obliterated. The cells of the hind wall have by this time become veritable fibres which when seen in section appear to be arranged nearly parallel to the optic axis, their nuclei J being seen in a row along their middle. The front wall, somewhat thickened at either side where it becomes continuous with the hind wall, is now a single layer of flattened cells separating the hind wall of the lens, or as we mav now say the lens itself, from the front limb of the lens-capsule ; of this it becomes the epithelium. The subsequent changes undergone consist chiefly in the continued elongation and multiplication of the lens-fibres, with the partial disappearance of their nuclei. During their multiplication they become arranged in the manner so characteristic of the adult lens. The lens-capsule and its adjuncts. In spite of the numerous investigations which have been made in reference to the development of the lens-capsule, its precise mode of origin can hardly even yet be said to be certainly known. Remak was led from analogy to regard it as a product of the mesoblast, though he did not succeed in satisfactorily demon- strating the fact. Kolliker looked upon it as a cuticular membrane thrown off by the superficial cells of the lens, and his view has been very generally adopted. Lieberkiihn has given a different account of its origin. According to him the involution of the lens, as we have already stated, carries inwards with it a very thin layer of meso- blast. This remains continuous with the mesoblast surround- ing the eyeball, so that when subsequently the mesoblast v.] THE LENS-CAPSULE. 109 grows forward over the front of the lens, the latter receives a complete mesoblastic investment. Of this mesoblast a very thin layer next to the lens both in front. and behind becomes separated from the rest, and forms the lens-capsule and suspensory ligament. The remainder of the mesoblast behind the lens becomes converted into the vitreous humour, the layer immediately in contact with the retina giving rise to the hyaloid membrane. That the hyaloid is really a product of the mesoblast and not a cuticular outgrowth from the epiblastic cells of the retina is indicated by the fact that it is con- tinuous over the pecten, where of course the retina is absent. At its first appearance the vitreous humour is a mass of stellate cells; while however it is rapidly enlarging to fill up the growing optic cup, a large portion of it becomes entirely fluid, the cellular elements being more and more restricted to the immediate neighbourhood of the posterior surface of the lens, where a few stellate cells may be seen even in the adult. Briefly to recapitulate. The eye commences as a lateral outgrowth of the fore-brain, in the form of a stalked vesicle. The stalk becoming narrowed and subsequently solid, is converted into the optic nerve. An involution of the superficial epiblast over the front of the optic vesicle, in the form first of a pit, then of a closed sac with thick walls, and lastly, of a solid rounded mass (the small central cavity being entirely obliterated by the thickening of the hind wall), gives rise to the lens. Owing to this involu- tion of the lens, the optic vesicle is doubled up on itself, and its eavity obliterated ; thus a secondary optic vesicle or optic cup with a thick anterior and a thin posterior wall is produced. As a result of the manner in which the doubling up takes place, or of the mode of growth afterwards, the cup of the secondary optic vesicle is at first imperfect along its under surface, where a gap, the choroidal fissure, exists for some little time, but subsequently closes up. The mesoblast in which the eye is imbedded gathers itself together around the optic cup into a distinct invest- ment of which the internal layers become the choroid, the external the sclerotic. An ingrowth of this investment between the front surface of the lens and the superficial 110 THE THIRD DAY. [CHAP. epiblast furnishes the body of the cornea, the epiblast itself remaining as the anterior corneal epithelium. A portion of mesoblast, carried in from the front by the lens during its involution, gives rise to the capsule of the lens and suspensory ligament, while some mesoblast entering on the under side through the choroidal fissure becomes (in birds) the pecten, and probably also contributes to the vitreous humour. Of the walls of the optic cup, the thinner outer (posterior) wall becomes, behind the line of the ora serrata, the pigment- epithelium of the choroid, while the thicker inner (anterior) wall supplies all the elements of the retina, including the rods and cones which grow out from it into the pigment- epithelium. Fic. 32. AOA SECTION THROUGH THE HIND-BRAIN OF A CHICK AT THE END OF THE THIRD Day or IxcuBation. IV. Fourth ventricle. The section shews the very thin roof and thicker sides of the ventricle. Ch. Notochord—.(diagrammatic shading). OV. Anterior cardinal vein. CG. Involuted auditory vesicle. CC points to the end which will form the cochlear canal. RZ. Recessus labyrinthi. fy. Hypoblast lining the alimentary canal. AO, AOA. Aorta, and aortic arch, v.] THE EAR. 111 In front of the line of the ora serrata, both walls of the optic cup, quite thin and wholly fused together, give rise to the pigment-epithelium of the ciliary processes and iris, the bodies of both these organs being formed from the meso- blastic investment. 12. During the second day the ear first made its appear- ance on either side of the hind-brain as an involution of the external epiblast, thrust down into the mass of mesoblast rapidly developing between the epiblast of the skin and that of the neural canal (Fig. 15, aw.p.). It then had the form of a shailow pit with a widely open mouth. Before the end of the third day, its mouth closes up and all signs of the opening are obliterated. The pit thus becomes converted into a closed vesicle lined with epiblast and surrounded by mesoblast. This vesicle is the otic vesicle, whose cavity rapidly enlarges while its walls become thickened (Fig. 32, CC). The changes by which this simple otic vesicle is converted into the complicated system of parts known as the internal ear, have been much more completely worked out for mammals than for birds. We shall therefore reserve a full account of them for a later portion of this work. Meanwhile a brief statement of the main course of events in the chick may be useful; and will be most conveniently introduced here, although we shall have, in doing so, to speak of changes which do not occur till much later than the third day. The internal ear consists essentially of an inner mem- bransus labyrinth lying loosely in and only partially attached to an outer osseous labyrinth. The membranous labyrinth (Fig. 33) consists of two parts: (1) the vestibule, with which are connected three pairs of semicircular canals, pag’, fr’, hor’, and a long narrow hollow process, the aqueductus or recessus vestibuli, and (2) the ductus cochlearis, which in birds is a flask-shaped cavity slightly bent on itself. The several parts of each of these cavities freely communicate, and the two are joined together by a narrow canal, the canalis reuniens, cr. The osseous labyrinth has a corresponding form, and may be similarly divided into parts: into a bony vestibule. with its bony semicircular canals and recessus vestibuli, and into a bony cochlea ; but the junction between the cochlea and the bony vestibule is much wider than the membranous canalis reuniens, 112 THE THIRD DAY. [CHAP. A Fic. 33- B PAG” 5 ike : sn ae ) hor s mr by La Loy—_——- Two VIEWS OF THE MEMBRANOUS LABYRINTH OF COLUMBA DOMESTICA (copied from Hasse). A from the exterior, B from the interior. hor’ horizontal semicircular canal, hor ampulla of ditto, pag’ posterior vertical semicircular canal, pag ampulla of ditto, fr’ anterior vertical semicircular canal, fr ampulla of ditto, w utriculus, vw recessus utriculi, v the connect- ing tube between the ampulla of the anterior vertical semicircular canal and the utriculus, de ductus endolymphaticus (recessus vestibuli), s sacculus hemisphericus (this is smaller in birds than in any other vertebrate), cr canalis reuniens, lag lagena (the dilated extremity of the cochlea), mr membrane of Reissner, which forms the boundary between the scala vestibuli and scala media, pl Basilar membrane, which forms the boundary between the scala tympani and the scala media. The cochlea of a bird consists (1) of a scala vestibuli with a very small lumen, which opens at one end into the perilymphatic cavity of the vestibule, and at the other into the lagena (the dilated extremity of the cochlea corresponding with the cupola of mammals) ; (2) of a scala tympani, also opening into the lagena at one end, and into the foramen rotundum at the other; (3) of a scala media ending blindly at one end, but in communication with the mem- branous vestibule at the other, through the membranous canalis reuniens (c/’). As in mammals, the cavity of the osseous cochlea is divided lengthways by the ductus cochlearis into a scala tympani ending in a foramen rotundum, and a scala vestibuli ending in the cavity of the osseous vestibule, which in its turn is connected with the foramen ovale. The auditory nerve, piercing the osseous labyrinth in various points, is distributed in the walls of the membranous labyrinth. All these complicated structures are derived from the simple primary otic vesicle by changes in its form and differentiation of its walls. All the epiblast of the vesicle ~ — 132 THE THIRD DAY. [CHAP. hollow processes, lined with hypoblast. Each diverticulum becomes in this way surrounded by a thick mass composed partly of solid cylinders, and to a less extent of hollow processes, continuous with the cylinders on the one hand, and the main diverticulum on the other, all knit together with commencing blood-vessels and unchanged mesoblastic tissue. Between the two masses runs the meatus venosus, with the bulgings on which, referred to above, the blood-vessels in each mass are connected. Early on the fourth day each mass sends out underneath the meatus venosus a solid projection of hypoblastic cylinders ‘towards its fellow, that from the left side being much the longest. The two projections unite and form a long solid wedge, which passes obliquely down from the right (or from the embryo lying on its left side, the upper) mass to the left (or lower) one. In this new wedge may be seen the same arrangement of a network of hypoblastic cylinders filled in with vascular mesoblast as in the rest of the liver. The two original diverticula with their investing masses represent respectively the right and left lobes of the liver, and the wedge-like bridge connecting them is the middle lobe. During the fourth and fifth days the growth of the solid, lobed liver thus formed is very considerable ; the hypoblastic cylinders multiply rapidly, and the network formed by them becomes very close, the meshes containing little more than blood-vessels. The hollow processes of the diverticula also ramify widely, each branch being composed of a lining of hypoblast enveloped in a coating of spindle-shaped meso- blastic cells. The blood-vessels are in direct connection with the meatus venosus—have become, in fact, branches of it. It may soon be observed, that in those vessels which are connected with the posterior part of the liver (Fig. 53), the stream of blood is directed from the meatus venosus into the network of the liver. In those connected with the anterior part the reverse is the case; here the blood flows from the liver into the meatus venosus. The thick network of solid cylinders represents the hepatic parenchyma of the adult liver, while the hollow processes of the diverticula are the rudiments of the biliary ducts. The exact morphological significance of these anastomosing cylinders, and v.] THE PANCREAS. 133 the manner of their ultimate metamorphosis into the ordinary hepatic tissue, is not as yet quite clear. If we suppose each solid cylinder to represent a duct with its lumen almost, but not quite, completely obliterated, we should gain a view agreeing very closely with that put forward by Hering on the structure of the adult liver. During the fifth day, a special sac or pouch is developed from the right primary diverticulum. This pouch, consisting of an inner coat of hypoblast, and an outer of mesoblast, is the rudiment of the gall-bladder. 20. About the middle of the third day, the pancreas (Fig. 42, p.) also appears, but its exact mode of origin is still somewhat doubtful. According to Gétte (Beit. z. entwick. des Darmcanals des Hiihnchens) it commences as a thickening of both the hypoblast and mesoblast of a portion of the wall of the digestive canal on the same level as the left diverticulum of the liver. The hypeblast in the centre of this thickening becomes hollow, forming a cavity connected with the inside of the digestive canal by a narrow opening. Around this cavity processes of hypoblast are seen on the fourth day stretching into the surrounding mesoblast. These processes, which are at first solid but afterwards become hollow and ultimately branched, are in the early stages so completely covered up by mesoblast that they are not visible on the exterior. The primary cavity elongates into the duct, the hollow processes representing its branches. On the sixth day a new similar outgrowth takes place between the primary one and the stomach. This, which ultimately coalesces with its predecessor, gives rise to the second duct, and forms a considerable part of the adult pancreas. A third duct is formed at a much later period. According to this view, with which those put forward by Kdlliker and Remak in the main agree, the so-called ‘secreting’ cells of the pancreas as well as the epithelial lining of the ducts are derived from hypoblast. Schenk (Die Bauchspeicheldriise des Embryos. Anat. u. Physiol. Untersuch- ung. Wien. S. I.) however is of opinion that the former originate in a transformation of the mesoblast, the hypoblast giving rise to the epithelium of the ducts only. Shortly after the first appearance of the pancreas, the spleen appears as a thickening of the mesentery of the stomach (mesogastricum) and is therefore entirely a mesoblastic struc- ture. Its development has been recently investigated by Peremeschko (Sitz. der k. Akad. in Wien, Bd. 56, 1867) and by W. Miiller (Stricker’s Histology). According to these investigators, the mass of mesoblast which forms the spleen becomes early separated by a groove on the one side from the pancreas and on the other from the mesentery. Some of its cells become elongated, and send out processes which, uniting with like processes from other cells, form the trabe- cular system. From the remainder of the tissue are derived the cells of the spleen pulp, which frequently contain more than one nucleus. Especial accumulations of these take place at a later period to form the so-called Malpighian corpuscles of the spleen. 21. The thyroid body is also formed towards the end of the third day, in connection with the alimentary canal. 134 THE THIRD DAY. [CHAP. According to Miiller (Ueber die Entwickelung der Schilddriise. Jenaische Zeitschrift, 1871) who has recently studied its development with great care, the thyroid body arises on the third day as an involution from the hypoblast of the throat opposite the point of origin of the second arterial arch. ‘This involution becomes by the fourth day a solid mass of cells, and by the fifth ceases to be connected with the epithelium of the throat, becoming at the same time bilobed. By the seventh day it has travelled somewhat backwards, and the two lobes have completely separated from each other. By the ninth day the whole is invested by a capsule of connective tissue, which sends in septa dividing it into a number of lobes or solid masses of cells, and by the sixteenth day it isa paired body composed of a number of follicles, each with a ‘membrana propria,’ and separated from each other by septa of connective tissue, much as in the adult. 22. Coincidently with the appearance of these several rudiments of important organs in the more or less modified splanchnopleure-folds, the solid trunk of the embryo is undergoing marked changes. When we compare a transverse section taken through say the middle of the trunk at the end of the third day (Fig. 44), with a similar one of the second day (Fig. 20), or even the commencement of the third day (Fig. 41), we are struck with the great increase of depth (from dorsal to ventral surface) in proportion to breadth. This is partly due to the slope of the side walls of the body having become much steeper as a direct result of the rapidly progressing folding off of the embryo from the yolk-sac. But it is also brought about by the great changes both of shape and structure which are taking place in the protovertebre as well as by the development of a mass of tissue between the notochord and the hypoblast of the alimentary canal. 23. The protovertebrae on the second day, as seen in a transverse section, Fig. 20, P.v., are somewhat quadrilateral in form but broader than high. Each at that time consists of a somewhat thick cortex of radiating rather granular columnar cells, enclosing a small kernel of spherical more transparent cells. Remak and after him Kélliker have described the centre of the proto- vertebra as being simply fluid without structural elements. His explicitly denies this in the case of the protovertebre of the neck, and it seems probable that the centre is in all cases really occupied by transparent spherical cells, Towards the end of the second and the beginning of the third day, the central cells increase rapidly in number (Fig. 41), and towards the end of the latter day (Fig. 44), as it were lift wp and push out the columnar cortex above and at the outer side. In this way the portions forming the v.] THE MUSCLE-PLATES. 135 SECTION THROUGH THE DorsaL REGION OF AN EMBRYO AT THE END OF THE THIRD Day. Am. amnion. m. p. muscle-plate. C. V. cardinal vein, Ao. dorsal aorta. The section passes through the point which the dorsal aorta is just commencing to divide into two branches. Ch. notochord. W.d. Wolffian duct. W. 0. commencing differentiation of the mesoblast cells to form the Wolffian body. ep. epiblast. SO. somatopleure. Sp. splanchnopleure. hy. hypoblast. The section passes through the point where the digestive canal communicates with the yo!k-sac, and is consequently still open below. This section should be compared with the section through the dorsal region of an embryo at the commencement of the third day (Fig. 41). The chief differences between them arise from the great increase in the space (now filled with meso- blast-cells) between the notochord and the hypoblast. In addition to this we have in the later section the completely formed amnion, the separation of the muscle-plate from the protovertebre, the formation of the Wolffian body, ete. The mesoblast including the Wolffian body and the muscle-plate (m.p.) is represented in a purely diagrammatic manner. The amnion, of which only the inner limb or true amnion is represented in the figure, is seen to be composed of epiblast and a layer of mesoblast ; though in contact with the body above the top of the medullary canal, it does not in any way coalesce with it, as might be con- cluded from the figure. 136 THE THIRD DAY. [CHAP. upper and outer border of the protovertebra become separated from the rest of the cortex, the columnar cells of the latter at the same time losing their distinctive characters and ceasing to be distinguishable from the central cells. As a consequence of this the whole protovertebra, while thus increasing in breadth out of proportion to its height, becomes split up into two portions which lie one above the other. Of these the upper one, which is from the first the most flattened and longest, follows the curvature of the body-wall and thus from being nearly horizontal comes to slope at a considerable angle. It now receives the name of muscle-plate, Fig. 44, m.p. Of its subsequent changes we shall have to speak in a succeeding chapter. The remaining portion of the original protovertebra is still called protovertebra and begins to extend inwards over the neural canal above and towards the notochord below. 24. Meanwhile the breadth or rather depth of the trunk is also being increased by the development of mesoblastic cells between the notochord and hypoblast. In a transverse section of a 45 hours’ embryo a consider- able mass of cells may be seen collected between the pro- tovertebra and the point where the divergence into somato- pleure and splanchnopleure begins (Fig. 20 just below W-d., also Fig. 41, the diagrammatically shaded part lying between p.v. and g.e). This mass of cells, which we may speak of as the intermediate cell-mass, now passes without any very sharp line of demarcation into the protovertebra itself; and as the folding in of the side wall progresses, increases in size and grows in between the notochord and the hypoblast, but does not accumulate to a sufficient extent to separate them widely until the end of the third or beginning of the fourth day. "The fusion between the intermediate cell-mass and the outer and under portions of the altered protovertebree becomes so complete on the third day that it is almost impossible to say which of the cells in the immediate neighbourhood of the notochord are derived from the proto- vertebree and which from the intermediate cell-mass. It seems probable however that the cells which form the immediate investment of the notochord really belong to the protovertebree. ae v.] THE CRANIAL NERVES. 137 Schenk (Wien. Sitz. Bericht. 1868) describes all the cells which invest the hypoblast of the digestive tract, as primarily derived from the proto- vertebrz, with the exception of the peritoneal epithelium, which alone, he con- siders, is the representative of the original mesoblast of the splanchnopleure. According to this view, the muscles of the walls of the alimentary canal, and the ‘hypaxial’ muscles, are derived from the original protovertebre, quite as much as those muscles which spring out of the muscle-plate. In the absence of any satisfactory means of distinguishing the cells of the intermediate eell-mass from those of the protovertebre, this view must be considered as at least very doubtful. 25. In the mesoblast, which lies by the side of the hind brain, and which though not divided into protover- tebre is the prolongation forwards of the same column of mesoblast out of which in the trunk the protovertebree are formed, there appear on either side in the course of the third day a series of four small opaque masses, somewhat pearshaped with the stalk directed away from the middle line. These are the rudiments of four cranial nerves, of which two lie in front of and two behind the auditory vesicle. The most anterior of these is the rudiment of the fifth nerve (Figs. 25, V. 45, V). Its narrowed outer portion or Fic, 45. HEAD oF AN Empryo CHICK OF THE THIRD Day (seventy-five hours) VIEWED SIDEWAYS AS A TRANSPARENT OBJECT (from Huxley). Ia. cerebral hemispheres. Id. vesicle of the third ventricle. II. mid-brain. III. hind-brain. g. nasal pit. a. optic vesicle. b. otic vesicle. d. infundi- bulum. e. pineal body. h. notochord. V. fifth nerve. VII. seventh nerve. VIII. united glossopharyngeal and pneumogastric nerves. 1, 2, 3, 4, 5 the five visceral folds. The stage here represented is a little later than that shewn in Fig. 25, with which it should be compared. 138 THE THIRD DAY. [ CHAP. stalk divides into two bands or nerves. Of these one passing towards the eye terminates at present in the immediate neighbourhood of that organ. Compare Fig. 46. The other branch (the rudiment of the inferior maxillary branch of the fifth uerve) is distributed to the first visceral arch (Fig. 46). The second mass (Fig. 25, VII. 45, VII.) is the rudiment of the seventh, or facial nerve. It is the nerve of the second visceral arch. The two masses behind the auditory vesicle represent the glossopharyngeal and pneumogastric nerves (Fig. 45, VIIL., Fig. 46, G. Ph. and Pg.). At first united, they subsequently become separate. The glossopharyngeal supplies the third arch, and the pneumogastric the fourth arch. These four masses, representing four important mixed cranial nerves, seem to be derived directly from the mesoblast surrounding the hind-brain. It is worthy of notice that they are mixed, sensory and motor, nerves; for, restricted as are the sensory functions of the seventh, and the motor func- tions of the pneumogastric in the adult mammal, the study of their com- parative physiology leaves no doubt as to the essentially mixed nature of each. It is also worthy of note that of the third, fourth and sixth nerves, no such early rudiments appear; and there are reasons for thinking that these are in reality intercranial branches, the third and fourth of the fifth, and the sixth of the seventh nerves. The purely sensory nerve or rather sense-nerve, the auditory, seems to have a different origin altogether from all the above, though it may perhaps be looked upon as the dorsal branch of the seventh, while the erratic hypoglossal appears to be distinctly a spinal nerve. Of the interesting relations of these cranial nerves to the visceral arches, we shall have to speak more fully in the second part of this work, when we describe the more primitive forms in the lower vertebrata. . At the same time that these ganglia make their appearance, or a little earlier, near the beginning of the third or end of the second day, there may be seen in ~ the region of the hind-brain, lines which appear to divide off the mesoblast on either side into masses somewhat resembling protovertebre. Of these masses there are four or five on each side, generally three in front of, and two behind the optic vesicle. They were first noticed by Remak, and are easily dis- tinguished from the rudiments of the cranial nerves. They at first sight suggest the idea of an initial and transitory segmentation of the cranial mesoblast into protovertebre. It seems possible that they are, in reality, appearances pro- duced by a series of very characteristic transverse wrinkles into which the walls of the hind-brain are at this time thrown, and which subsequently disap- pearing altogether, as the walls increase in thickness, may perhaps be viewed as indications of an aborted segmentation of the hind-brain into a series of vesicles. The true nature of these quadrate masses is still very problematical. 26. On the second day the newly formed Wolffian duct extended along the greater part of the length of the embryo as a tube resting on the mass of celis which we have already called the intermediate cell-mass. v.] THE WOLFFIAN DUCT. 139 On the third day, in consequence of the continually folding in of the somatopleure and especially of the splanch- nopleure, as well as owing to the changes taking place in the protovertebre, the Wolffian duct undergoes a remarkable change of position. Instead of lying as on the second day immediately under the epiblast (Fig. 20, W.d.), it is soon found to have apparently descended into the middle of the intermediate cell-mass (Fig. 41, w.d.) and at the end of the third day occupies a still lower position and even pro- jects somewhat into the pleuroperitoneal cavity. (Fig. 44, W.d.) Towards the end of the day the rudiments of the Wolffian bodies (Fig. 44, W.b.) begin to make their appearance in con- nection with the ducts, but the consideration of these may conveniently be reserved to the next chapter. 27. The chief events then which take place on the third day are as follows: 1. ‘The turning over of the embryo so that it now lies on its left side. 2. The cranial flexure round the anterior extremity of the notochord. 3. The completion of the circulation of the yolk-sac; the increased curvature of the heart, and the demarcation of its several parts; the appearance of new aortic arches, and of the cardinal veins. ¢ 4. The formation of four visceral clefts and five visceral arches, 5. The involution to form the lens, and the formation of the secondary optic vesicle. 6. The closing in of the otic vesicle. 7. The formation of the nasal pits. 8. The appearance of the vesicles of the cerebral hemi- spheres; the separation of hind-brain into cerebellum and medulla oblongata. 9. The completion of the fore-gut and of the hind-gut ; the division of the former into cesophagus, stomach and duodenum, of the latter into large intestine and cloaca. 10. The formation of the lungs as two diverticula from the alimentary canal immediately in front of the stomach. 11. The formation of the liver and pancreas: the former as two diverticula from the duodenum, which subsequently 140 THE THIRD DAY. [cHAP. Vv. become united by solid outgrowths; the latter as a single diverticulum also from the duodenum. 12, The changes in the protovertebre and the appear- ance of the muscle-plates. 13. The appearance of the cranial nerves in the meso- blast adjoiing the hind brain. 14, The change in position of the Wolffian duct. he CHAPTER VI. THE CHANGES WHICH TAKE PLACE DURING THE FOURTH DAY. 1. ON opening an egg in the middle or towards the end of the fourth day, a number of points in which progress has been made since the third day are at once apparent. In the first place, the general growth of the embryo has been very rapid, so that its size is very much greater than on the previous day. In the second place, the white of the egg has still further diminished, the embryo lying almost in immediate contact with the shell membrane. _ The germinal membrane embraces more than half the yolk, and the vascular area is about as large as a halfpenny. Corresponding to the increased size of the embryo, there is a great increase in the quantity of blood circulating in the vascular area as a whole, though the sinus terminalis is already less distinct than it was. 2. The amnion becomes increasingly conspicuous. It is now seen as a distinct covering obscuring to a certain extent the view of the body of the chick beneath, and all traces of the junction of its folds are by this time lost. As yet there is very little fluid in the amniotic sac proper, so that the true amnion lies close upon the embryo. 3. The folding off of the embryo from the yolk sac has made great progress. The splanchnic stalk, which on the third day was still tolerably wide, inasmuch as about one third of the total length of the alimentary canal was as yet quite open to the yolk sac below, now becomes so much con- stricted by the progressive closing in of the splanchnopleure 142 THE FOURTH DAY. [CHAP. Fie. 46. EMBRYO AT THE END OF THE FourTH DayY SEEN AS A TRANSPARENT OBJECT. The amnion has been completely removed, the cut end of the somatic stalk is shewn at S.S. with the allantois (Al.) protruding from it. C.H. cerebral hemisphere. /. B. fore brain or vesicle of the third ventricle with the pineal gland (Pn.) projecting from its summit. J4.8. mid brain. Cb. cerebellum. JV. V. fourth ventricle. JZ. lens. ch.s. choroid slit. Owing to the growth of the optic cup the two layers of which it is composed cannot any longer be seen from the surface, but the posterior surface of the choroid layer alone is visible. Cen. V. auditory vesicle. s.m. superior maxillary process. 1f. 27. etc. first, second, third and fourth visceral folds. V. fifth nerve sending one branch to the eye, the ophthalmic branch, and another to the first visceral arch. VJJ. seventh nerve passing to the second visceral arch. G. Ph. glossopharyngeal nerve passing towards the third visceral arch. Pg. pneumogastric nerve passing towards the fourth visceral arch. i. investing mass. No attempt has been made in the figure to indicate the position of the dorsal wall of the throat, which cannot be easily made out in the living embryo. ch. notochord. The front end of this cannot be seen in the living embryo. It does not end however as shewn in the figure, but takes a sudden bend downwards and then terminates in a point. H¢. heart seen through the walls of the chest. JM. P. muscle-plates. W. wing. H. LZ. hind limb. Beneath the hind limb is seen the curved tail. “| VI.] THE LIMBS. 143 folds, that the alimentary canal may be said to be connected with the yolk sac by a very narrow neck only. This rem- nant of the splanchnic stalk we may now call the wmbilical duct; though narrow, it is as yet quite open, affording still free communication between the inside of the yolk sac and the interior of the alimentary canal. The somatic stalk, though narrowing somewhat, is much wider than the splanchnic stalk, so that a considerable ring-shaped space exists between the two. 4. Another very prominent feature is the increase in the cranial flexure. During the third day, the axis of the front part of the head was about at right angles to the long axis of the body; the whole embryo being still somewhat retort- shaped. On this day, however, the flexure has so much increased that the angle between the long axis of the body and that of the front segment of the head is an acute one, and the mouth is turned so as completely to face the chest. The tail-fold, which commenced to be noticeable during the third day, has during this day increased very much, and the somewhat curved tail (Fig. 46) forms quite a conspicuous feature of the embryo. The general curvature of the body has also gone on increasing, and as the result of these various flexures, the embryo has very much the appearance of being curled up on itself (Fig. 46). ; 5. The distinct appearance of the limbs must be reckoned as one of the most important events of the fourth day. Owing to the continued greater increase of depth than of breadth, the body of the embryo appears in section (Fig. 47) higher and relatively narrower than even on the third day, and the muscle-plates, instead of simply slanting downwards, come to be nearly vertical in position. Not far from the line which marks their lower ends, the somatopleure, almost immediately after it diverges from the splanchnopleure, is raised up (Fig. 47, W.R.) into a low rounded ridge which runs along nearly the whole length of the embryo from the neck to the tail. It is on this ridge, which is known as the Wolffian ridge, that the limbs first appear as flattened conical buds project- ing outwards. They seem to be local developments of the ridge, the rest of which becomes less and less prominent as they increase in size. Each bud, roughly triangular in sec- 144 THE FOURTH DAY. [CHAP Fia. 47. eee ne. Sep) \ > SECTION THROUGH THE LUMBAR REGION OF AN EMBRYO AT 'THE END OF THE FourtH Day. n.c.neural canal. p.7. posterior root of spinal nerve with ganglion. a. 7. anterior root of spinal nerve. A. G. C. anterior grey column of spinal cord. A. W.C. anterior white column of spinal cord just commencing to be formed, and not very distinctly marked in the figure. m. p. muscle plate. ch. notochord. W. R. Wolffian ridge. A. O. dorsal aorta. V.c. a. posterior cardinal vein. W.d. Wolffian duct; its section is not circular, owing to its being cut through at a point where it is being joined by one of the tubules. W. 6, Wolffian body, consisting of tubules and Malpighian corpuscles. One of the latter is represented on each side, that on the left hand having its glomerulus entirely filled with blood-corpuscles. g. e. germinal epithelium. M.d. commencing involution of germinal epithelium to form the duct of Miiller. d. alimentary canal. M. commencing mesentery. SS. 0. somato- pleure. S.P. splanchnopleure. V. blood-vessels. pp. pleuroperitoneal cavity. VIL] THE NASAL PITS. 145 tion, consists of somewhat dense mesoblast covered by epiblast which on the summit is thickened into a sort of cap. The front hmbs or wings (Fig. 46) arise just behind the level of the heart, and the hind limbs in the immediate vicinity of the tail. The first traces of them can be seen towards the end of the third, but they do not become conspicuous till the fourth day, by the end of which the two pairs may be already distinguished by their different shapes. The front limbs are the narrowest and longest, the hind ones being comparatively short and broad. Both are flattened from above downwards and become more so as their growth continues. 6. In the head, the vesicles of the cerebral hemispheres are rapidly increasing in size, overlapping the insignificant olfactory vesicles in front, and encroaching on the ‘tween- brain or vesicle of the third ventricle behind. The mid-brain is now, relatively to the other parts of the brain, larger than at any other epoch, and an indistinct median furrow on its upper surface indicates its division into two lateral halves. The great increase of the mesoblastic contents of the second- ary optic vesicle or involuted retinal cup causes the two eye- balls to project largely from the sides of the head (Fig. 48, Up). The mass of mesoblast which invests all the various parts of the brain, is not only growing rapidly below and at the sides, but is also undergoing developments which result in the forma- tion of the primitive skull, and of which we shall speak in detail in a subsequent separate chapter. All these events, added to the cranial flexure spoken of above, give to the anterior extremity of the embryo a shape which it becomes more and more easy to recognize as that of a head. 7. Meanwhile the face is also being changed. The two nasal pits were on the third day shallow depressions with thickened borders complete all round. As the pits deepen on the fourth day by the growth upwards of their rims, a break is observed in each rim in the form of a groove (Fig. 48, NV) directed obliquely downwards towards the cavity of the mouth, The fronto-nasal process or median ridge (Fig. 48, nf), which on the third day rose up between the superficial projections caused by the bulging anterior extremities of the vesicles of the cerebral hemispheres, and on the fourth day becomes increasingly prominent, separates the two grooves from each E. 10 146 THE FOURTH DAY. [CHAP. Fic. 48. FB caf “ii m “a Ma ‘TR i A. Heap oF AN Empryo CHIck oF THE FourtH DAY VIEWED FROM BELOW AS AN OPAQUE OBJECT. (Chromic acid preparation). CH. cerebral hemispheres. /'B. vesicle of the third ventricle. Op. eyeball. nf. naso-frontal process. M. cavity of mouth. S. M. superior maxillary process of /’. 1, the first visceral fold (inferior maxillary process). /. 2, F. 3. second and third visceral folds. NV. nasal pit. In order to gain the view here given the neck was cut across between the third and fourth visceral folds. In the section e thus made, are seen the alimentary canal a/ with its collapsed walls, the neural canal n.c., the notochord ch., the dorsal aorta AO., and the vertebral veins V. The incision has been carried just below the upper limit of the pleuroperi- toneal cavity, consequently a portion of the somatopleure appears at the angle between the two third visceral folds. Almost embraced by the piece of somato- pleure is seen the end of the bulbus arteriosus Ao. In the drawing the nasal groove has been rather exaggerated in its upper part. On the other hand the lower part of the groove where it runs between the superior maxillary process S.M/. and the broad naso-frontal process was, in this particular embryo, extremely shallow and indeed hardly visible. Hence the end of the superior maxillary process seems to join the inner and not, as described in the text, the outer margin of the nasal groove. A few hours later the separation of the two would have been very visible. B. The same seen sideways, to shew the visceral folds. Letters as before. other, and helps to form the inner wall of each of them. Abutting on the outer side of each groove and so helping to form the outer wall of each, lie the ends of the superior maxillary processes of the first visceral arch (Fig. 48 B, sm), which like the fronto-nasal process are increasing in size. By their continued growth, the groove is more and more deepened, and leading as it does from the nasal pit to the cavity of the mouth, may already be recognized as the rudiment of the passage of the posterior nares. VI.] THE CRANIAL NERVES. 147 8. During the latter half of the fourth day there appears at the bottom of the deep lozenge-shaped cavity of the mouth, in the now thin wall dividing it from the alimentary canal, a longitudinal, or according to Gotte a vertical slit which, soon becoming a wide opening, places the two cavities in complete communication. The cavity of the mouth, being, it will be remembered, formed partly by depression, partly by the growth of the sur- rounding folds, is lined entirely with epiblast, from which the epithelium of its surface and of its various glands is derived. In this respect, as Remak pointed out, it differs from all the rest of the alimentary canal, whose whole epithelium is formed out of hypoblast. 9. By the side of the hind-brain, in which the cerebellum, through the increased thickening of its upper walls, is be- coming more and more distinct from the medulla oblongata, both in front and behind the auditory vesicle, in which the rudiments of the cochlea and recessus vestibuli are already visible, the cranial ganglia and nerves are acquiring increased distinctness and size. They may be very plainly seen when the head of the fresh embryo 1s subjected to pressure. The foremost is the fifth cranial nerve (Fig. 46, V.) with its Gasserian ganglion ; it lies a little way behind the anterior extremity of the notochord immediately below the cerebellum. Next to this comes the seventh (Fig. 46, VZZ) nerve, starting just in front of the ear-vesicle, and extending far downwards towards the second visceral arch. The two nerves which lie behind the ear-vesicle are now obviously separate from each other; the front one is the glossopharyngeal (Fig. 46, G. Ph.), and the hinder one already shews itself to be the pneumo- gastric (Fig. 46, Pg.). 10. Besides the progressive changes of the alimentary canal and its surroundings, which we incidentally described in the last chapter, and the closure of the mid-gut to form the umbilical duct, of which we have also already spoken, a totally new and most important appendage of the digestive tract, the allantois, becomes for the first time conspicuous on this day, though the first rudiments of it appeared on the third. Soon after its appearance the allantois may easily be recog- nized as a pear-shaped vesicle lying in the hinder district of the pleuroperitoneal space, and connected with the under 10—2 148 THE FOURTH DAY. [ CHAP. surface of the cloaca by a long hollow stalk, which places its cavity in communication with that of the alimentary canal. Both vesicle and stalk have an outer coat of mesoblast and an inner lining of what apparently is hypoblast. So much any observer may readily determine for himself; but of the earliest stages of the development of this organ different embryologists have given very different accounts. Von Baer believed that, soon after the cloaca was formed by the enlargement of the cecal hind end of the alimentary canal, the allantois arose from it as a spherical diverticulum, generally visible about the middle of the third day, in whose formation both of the coats of the alimentary canal took part. This spherical diverticulum gradually lengthened out into a pear-shaped vesicle, connected with the cloaca by a hollow stalk which rapidly narrowed and lengthened, until the allantois formed an independent hollow body, composed of an outer coat of mesoblast and a lining of hypoblast, and communicating with the cloaca by a narrow tube of the same construction. Reichert (Entwicklungsgeschichte, s. 186) on the other hand stated that the allantois was formed of two solid outgrowths from the mesoblast of the somato- pleure, which subsequently coalesced and became hollow; but believed that it was primarily connected with the Wolffian ducts and not with the cloaca. According to Remak (Entwicklung, § 57, 58) it is formed by two solid vascular outgrowths of the mesoblast of the body-wall, one on each side of the middle line, which project in the pleuroperitoneal cavity near to the cloaca. These two outgrowths coalesce, and then grow up, till they come in contact with Fic. 49. LONGITUDINAL SECTION OF THE TAIL-END OF AN EMBRYO CHICK AT THE COMMENCEMENT OF THE THIRD Day (Dobrynin), t. the tail, m. the axial mesoblast of the body, about to form the protovertebre. 2x’. the roof of «’’. the neural canal. Dd. the hind end of the hind-gut. SO. somatopleure. Spl. splanchnopleure. wu. the mesoblast of the splanchno- pleure carrying the vessels of the yolk-sac. pp. pleuroperitoneal cavity. Df. the epithelium lining the pleuroperitoneal cavity. All. the commencing allantois. w. and y. the bypoblast thickened and projecting on either side of the opening of the allantois. VI. ] THE ALLANTOIS. 149 the wall of the cloaca: with this they unite, and form together a solid spherical body, bearing on its external surface a median furrow, indicating its double origin. A narrow diverticulum of hypoblast now passes into the mass, and forms within it a cavity, which is at first small and, corresponding to the ex- ternal contour of the body, to a certain extent double. The hypoblast diverticulum grows rapidly, while its mesoblastic covering remains nearly stationary, so that the mesoblast finally comes to form a thin coating only over the hypoblast. His (op. cit.) gives a somewhat elaborate and complicated account of the development of the allantois; which is accepted by Waldeyer (Kierstock und Ei) and Bornhaupt (Untersuchung tiber die Entwickelung des Urino-genitalsystems beim Hiihnchen, Riga, 1867). It appears to be nearly the same as the fuller account given by Dobrynin (Ueber die erste Anlage der Allantois. Sitz. der k, Akad. Wien, Bu. 64, 1871), ot which the following is an abstract. Whea the first commencement of the hind fold takes place, immediately beyond the point where the hypoblast turns back to assume its normal direction over the yolk-sac, a narrow diverticulum which points backwards and some- what upwards is formed by a special flexure of the splanchnopleure. The open end of the diverticulum, Fig. 49, All., looks forwards towards the wide opening connecting the digestive tract with the yolk-sac; its blind end points directly towards the pleuroperitoneal cavity. This diverticulum is the commencing allantois. It is lined by hypoblast, while its exterior is composed of the mesoblast of the splanchnopleure. As the folding in to form the digestive tract increases, the diverticulum alters LONGITUDINAL SECTION OF THE TAIL-END OF AN Empryo CHICK AT THE MIDDLE OF THE THIRD Day (Dobrynin). t. the tail; the line of reference points to the axial mesoblast at the tail. zx’. epiblast. SO. somatopleure. m. mesoblast to form the body wall. V. commencing amniotic fold. Hp. space between the true and the false amnion. pp. Pleuroperitoneal cavity. Spl. splanchnopleure. (G. Cloacal enlargement of the alimentary canal. Dd. dorsal wall of the alimentary canal, All. vesicle of the allantois having a wide opening into the alimen- tary canal. ; 150 THE FOURTH DAY. [ CHAP. its position and becomes quite parallel with the commencing digestive tract. Its cavity is separated from that of the digestive canal by a projection of mesoblast covered by hypoblast ; but both open freely in front into the common splanchnic stalk. In the next stage it still further alters its position, and forms, Fig. 50, arather wide vesicle lying immediately below the hind end of the digestive canal, with which it communicates freely by a still broad opening; its blind end projects freely into the pleuroperitoneal cavity below. It was in this condition when Von Baer first observed it. At the time when these changes are taking place, the somatopleure is being folded in to form the walls of the body; and as the folds, one on either side, are carried forward from the extreme end of the tail, they present them- selves, when seen from within or in sections, as two ridges projecting towards the sides of the allantois: Reaching the allantois these ridges fuse with its wall, and in this way reduce the pleuroperitoneal cavity immediately below the allan- tois to quite a narrow space, which is seen in section as a mere chink. Remak apparently mistook these infoldings of the somatopleure, and the consequent projections into the pleuroperitoneal cavity, for the first formation of the allantois, although they have in fact little or no connection with it. We may therefore probably consider the allantois as a portion of the cloaca, which grows forward and becomes an independent spherical vesicle, still however remaining con- nected with the cloaca by a narrow canal which forms its neck or stalk. The opening of the allantois into the cloaca is on the under side of the latter. Both the neck and vesicle of the allantois are lined by hypoblast, while its exterior is com- posed of the mesoblast of the splanchnopleure. From the first the allantois lies in the pleuroperitoneal cavity. In this cavity it grows forwards till it reaches the front limit of the hind-gut, where the splanchnopleure turns back to reach the yolk-sac. It does not during the third day project be- yond this point; but on the fourth day begins to pass out beyond the body of the chick along the as yet wide space between the splanchnic and somatic stalks of the embryo on its way to that space between the external and internal folds of the amnion, which it will be remembered is directly continuous with the pleuroperitoneal cavity (Fig foe ci) See he this space it eventually spreads out over the whole body of the chick. On the first half of the day the vesicle is still very small, and its growth is not very rapid. Its mesoblast wall still remains very thick. In the later half of the day its growth becomes very rapid and it forms a very conspicuous object i in a chick of that date (Fig. 46, Al). At the same time its blood-vessels become important. To these we shall presently return. VI.] THE PROTOVERTEBR, 151 11. The protovertebre, which by the continued differ- entiation of the axial mesoblast at the tail end of the embryo have increased in number from thirty to forty, undergo during this day changes of great importance. Since these changes are intimately connected with the subsequent development of the vertebral column, it will perhaps be more convenient to describe briefly here the whole series of events through which the protovertebree become converted into the per- manent structures to which they give rise, though many of the changes do not take place till a much later date than the fourth day. The separation of the muscle-plates (Chap. v. § 23) left the remainder of each protovertebra as a somewhat tri- angular mass lying between the neural canal and notochord on the inside, and the muscle-plate and intermediate cell-mass on the outside (Fig. 44). Already on the third day the upper angle of this triangle grows upwards, between its muscle- plate and the neural canal, and meeting its fellow in the middle line above, forms a roof of mesoblast over the neural canal, between it and the superficial epiblast. At about the same time, the inner and lower angle of the triangle grows inwards towards the notochord, and passing both below it (between it and the aorta) and above it (be- tween it and the neural canal), meets a similar growth from its fellow protovertebra of the other side, and thus com- pletely invests the notochord with a coat of mesoblast, which, as seen in Fig. 47, is at first much thicker on the under than on the upper side. While the inner portion of each protovertebra is thus extending inwards around both notochord and neural canal, the remaining outer portion is undergving a remarkable change. It becomes divided into an anterior or preaxial, and a posterior or postaxial segment. The anterior, which is the larger and more transparent of the two, is the rudiment of the spinal ganglion and nerve, while the posterior, which remains more particularly connected with the extensions round the neural canal and notochord, goes to form part of the permanent vertebra. In this way, each protovertebra, having given rise to a muscle-plate, is further divided into a ganglionic rudiment, and into a mass which we may speak of as a “primary” 152 THE FOURTH DAY. [CHAP. vertebra, consisting as it does of a body or mass investing the notochord, from which springs an arch covering in the neural canal. Both body and arch consist at this epoch of but slightly differentiated mesoblast, and the arch springs, to a certain extent, not only from the posterior segment of the protover- tebra, but also from the anterior or ganglionic segment: though, as seen in Fig. 47, it is far less conspicuous at the level of the latter than of the former. Both neural canal and notochord are thus furnished from neck to tail with a complete investment of mesoblast, still marked, however, by the transparent lines indicating the fore and aft limits of the several protovertebre. This is sometimes spoken of as the “ membranous” vertebral column. The ganglionic rudiment, placed anteriorly to its corre- sponding primary vertebra, consists in chief of a large oval swelling, the ganglion of the posterior root (Fig. 47, pr). At a little distance beyond its ganglion, the posterior root is joined by the anterior root (ar); and the two form together the common nerve-trunk, which is at first very short. Com- pared with either root or with the nerve-trunk the ganglion, at this epoch and for some time afterwards, seems dispropor- tionably large. At first, neither root is connected with the involuted epiblast of the neural canal. Very speedily, how- ever, they both come to be united with that portion of the neural tube which, as we shall presently state, gives origin to the grey matter of the spinal cord. It is, however, easier to trace the fibres of the anterior root into the cord; than those of the posterior, and they can be followed in it for a greater distance. On the fourth day the nerves are composed of cells whose protoplasm is beginning to become converted into fibres. Amongst these fibres, the nuclei of the cells with distinct nucleoli are thickly scattered. On the sixth day and still more on the seventh the fibrillated structure of the nerves is much more distinct and the nuclei are far less numerous. The ganglia on the fourth day are composed of numerous nuclei surrounded by protoplasm, between which the fibres of the nerves pass. Covering this mass of nucleated cells is a layer of mesoblast (also derived from the tissue of the protovertebra) which, by the sixth day, forms a kind of sheath around them. The cells of the ganglia from the fourth to the sixth day contain round granular nuclei with distinct nucleoli very similar to the nuclei of the ordinary mesoblast-cells, The limits of the protoplasm of the individual cells are as a general rule not easily seen, but with care may be made out. The amount of protoplasm round each nucleus appears however to be very small. vI.] THE SPINAL GANGLIA. 153 The fibres of the nerve can easily be traced through the ganglion. In section they appear to have a somewhas wavy course, and by interlacing divide the ganglion into a number of elongated areas in each of which is a row of nuclei. In sections of the sixth day it is not possible to trace a connection between the nerve-fibres and the cells. The nuclei are must numerous at the lower ends of the ganglia. On the seventh day, the nuclei have become larger, and where the outline of a cell can be distinctly seen it is generally somewhat angular. In sections it is still on the seventh day not possible to trace any connection between the cells and the nerve fibres. Remak (op. cit.) speaks of the ganglia being composed of non-nucleated spheres, and Lockhart Clarke (Philosophical Transactions, 1862) also describes the ganglia-cells as ‘cells or nuclei” which are at first mere rounded masses of protoplasm, and do not acquire a nucleus till a later period. Both of these Statements are according to our observations incorrect. In the later stages according to Lockhart Clarke’s account (/oc. cit.) the cells of the ganglia send out processes which anastomose together into a fine network. The ceils also become connected with the nerve fibres, which can sometimes be seen to divide in the ganglion into a fine brush-like bundle of fibrille. At this time the cells possess a distinct nucleus and nucleolus. These changes he describes as completed by the ninth day of incubation. His believes that the spinal nerves are derived from downward prolonga- tions of the superficial epiblast descending between the protovertebre. This view has not been corroborated by subsequent observers. 12. The remaining portions of the protovertebra: form- ing the primary vertebrae or membranous vertebral column spoken of in the last paragraph, are converted into the per- manent vertebre ; but their conversion is complicated by a remarkable new or secondary segmentation of the whole vertebral column. On the fourth day, the transparent lines marking the fore and aft limits of the protovertebrz are still distinctly visible. On the fifth day, however, they disappear, so that the whole vertebral column becomes fused into a homoge- neous mass whose division into vertebrze is only indicated by the series of ganglia. This fusion, which does not extend to the muscle-plates in which the primary lines of division still remain visible, is quickly followed by a fresh segmentation, the resulting segments being the rudiments of the permanent vertebre. The new segmentation, however, does not follow the lines of the earlier division, but passes between the ganglionic and the vertebral portions, in fact, through the middle, of each protovertebra. In consequence, each spinal ganglion and nerve ceases to form the front portion of the primary vertebra, formed cut of same protovertebra as itself, . but is attached to the hind part of the permanent vertebra | 154 THE FOURTH DAY. [CHAP. immediately preceding. Similarly, the rudiment of each vertebral arch covering in the neural tube no longer springs from the hind part of the protovertebra from which it is an outgrowth, but forms the front part of the permanent ver- tebra, to which it henceforward belongs. The ganglia are still, however, the most conspicuous portions of each segment. By these changes this remarkable result is brought about, that each permanent vertebra is formed out of portions of two consecutive protovertebree. Thus, for instance, the tenth permanent vertebra is formed out of the hind portion of the tenth protovertebra, and the front portion of the eleventh protovertebra, while its arch, now attached to its front part, was attached to the hind part of the tenth protovertebra. The new segmentation is associated with or rather is caused by histological changes. At the time when the fusion takes place, the mesoblast, which in the form of processes from the protovertebral bodies surrounds and invests the notochord, has not only imcreased in mass but also has become cartilaginous, so that, as Gegenbaur (Untersuchung zur vergleichenden Anatomie der Wirbelsdule bei Amphibien und Reptilien, Leipzig, 1862) points out, we have for a short period on the fifth day a continuous and unsegmented carti- laginous investment of the notochord. This cartilaginous tube does not however long remain uni- form. At a series of points corresponding in number to the original protovertebree it becomes connected with a number of cartilaginous arches which appear in the protovertebral investment of the neural canal. These arches, which thus roof in the neural canal and each of which arises opposite to the vertebral portion of each protovertebral body, are the cartilaginous precursors of the osseous vertebral arches. We further find that the portions of the cartilaginous tube from which the arches spring come to differ histologically from the portions between them not connected with arches: they are clearer and their cells are less closely packed. There is however at this period no distinct segmentation of the cartilaginous tube, but merely a want of uniformity in its composition. The clearer portions, from which the arches spring, form the bodies of the vertebre, the segments between them the intervertebral regions of the column. VI.] SECONDARY SEGMENTATION OF VERTEBRAL COLUMN. 155 W. Schwarck (Beitriige zur Entwicklung der Wirbelsdule bei den Vogeln. Anatomische Studien, Dr Hasse, m1. Heft, 1872) states that both in the intervertebral and the vertebral segments the cartilage is divided into two layers, an inner, central, and an outer peripheral. This division is less marked in the intervertebral than in the vertebral region. The inner layer in the vertebral region he speaks of as “the body of the vertebra belonging to the notochord,” and the external layer as ‘‘the skeleton- forming layer.” On the fifth day a division takes place of each of the in- tervertebral segments into two unequal parts; a larger one appertaining to the vertebra in front and a smaller one to the vertebra behind. To the larger segment the spinal ganglia naturally remain attached, and thus comes about the altera- tion of their place in relation to the vertebree which we before spoke of. This fresh segmentation is not well marked, if indeed it takes place at all in the sacral region. Each arch at its first appearance corresponds to about the middle of a vertebral portion of a protovertebra, but after the secondary segmentation the portion of each vertebra behind its arch grows more quickly than that in front, and thus after a while the arches seem to spring from the front rather than from the middle of the vertebral segments. To recapitulate:—the original protovertebree lying side by side along the notochord, after giving off the muscle-plates, and dividing lengthways into ganglionic and vertebral portions, ow around, and by fusing together completely invest, with mesoblast of protovertebral origin, both neural canal and notochord. This investment, of which by reason of its greater growth the original bodies of the protovertebre seem to be only an out- lying part, becomes cartilaginous in such a way that while the notochord becomes surrounded with a thick tube of car tilage bearing no signs of segmentation, but having the ganglia lodged on its exterior at intervals, the neural canal is covered in with a series of cartilaginous arches, connected to each other by ordinary mesoblastic tissue only, but passing at their bases directly into the cartilaginous tube around the noto- chord. By a histological process of differentiation the cartila- ginous tube is divided into vertebral and intervertebral portions, the vertebral portions corresponding to the arches 156 THE FOURTH DAY. [ CHAP. over the neural canal. Fresh lines of segmentation then appear in the intervertebral portions, which run in such a way that each ganglion is now more closely associated with the vertebral portion in front of it than with that behind it, theugh the latter sprang in part from the same original protovertebra as itself. 13. Meanwhile from the fourth to the sixth day im- portant changes take place in the notochord itself. On its first appearance the notochord was, as we have seen, composed of somewhat radiately arranged but otherwise perfectly typical mesoblast-cells. On the third day some of the central cells become vacuo- lated, while the peripheral cells are still normal. The vacuo- lated cells exhibit around the vacuole a peripheral layer of granular protoplasm in which the nucleus lies embedded, whilst the vacuoles themselves are filled with a perfectly clear and transparent material, which in an unaltered con- dition is probably fluid. Towards the end of the day the notochord acquires a delicate structureless sheath which is no doubt a product of its peripheral cells. According to His there is a cavity in the centre of the notochord on the third day. We have never observed this, and it is denied by Miiller (Ueber den Bau der Chorda Dorsalis. Jenaische Zeitschrift. Bd. vi. 1871). On the fourth day all the cells become vacuolated with the exception of a single layer of flattened cells at the peri- phery; and the vacuoles themselves become larger. At the pot where the nucleus lies there is generally rather more protoplasm than round the remainder of the circumference of the cells. On the sixth day all the cells are vacuolated. In each cell the vacuoles have so much increased at the expense of the protoplasm that only a very thin layer of the latter is left at the circumference of the cell, at one part of which, where there is generally more protoplasm than elsewhere, the starved remains of a nucleus may generally be detected. Miiller (loc. cit.) considers that the cells have a membrane. This however is probably merely a hardened external layer of the protoplasm; and is stained by reagents. Dursy (Zur Entwicklungsgeschichte des Kopfes des Menschen und der hiheren Wirbelthiere) believes that what we have spoken of as vacuoles in the cells are really intercellular spaces. So that according to his view the notochord is composed of stellate cells with large round intercellular spaces filled with transparent intercellular matter. Superficially viewed a section of the noto- VI.] THE NOTOCHORD. 157 chord of the sixth day might be supposed to have such astructure, but the study of its development and a careful examination of its structure proves that this is not a correct account. According to the measurements of Miiller (loc. cit.) the diameter of the notochord on the third day is o‘og mm. and that of the central cells o':012— o’o18. On the fourth day the notochord is 0°16 mm. in diameter and its com- ponent cells are also larger. On the sixth day its diameter is at the maximum and reaches o°2 mm. The central cells measure 0°02 mm. From these measurements it will be seen that the vacuolation of the cells of the notochord is accompanied by a rapid growth both in the size of the cells and in the diameter of the notochord itself. 14. The notochord is on the sixth day at the maximum of its development, the change which it henceforward under- goes being of a retrograde character. From the seventh day onward, it is at various points encroached upon by its investment. Constrictions are thus produced which first make their appearance in the interverte- bral portions of the sacral region. In the cervical region, according to Gegenbaur, the intervertebral portions are not constricted till the ninth day, though as early as the seventh day constrictions are visible in the vertebral portions of the lower cervical vertebrae. By the ninth and tenth days, however, all the intervertebral portions have become distinctly constricted, and at the same time in each vertebral portion there have also appeared two constrictions giving rise to a central and to two terminal enlargements. In the space therefore corresponding to each vertebra and its appropriate intervertebral portion, there are in all four constrictions and three enlargements. On the twelfth day the ossification of the bodies com- mences. At that time, according to Schwarck (loc. cit.), the cartilaginous bodies of the vertebrze are composed of an inner layer in which the cells form lines radiating from the noto- chord, and an outer layer somewhat sharply separated from the inner one. In the inner layer, immediately around the notochord, ossification first commences. Gegenbaur (loc. cit. p. 67) considers that this layer in which ossification commences corresponds to the primordial body of the vertebre in amphibians. Schwarck is doubtful whether it corresponds to his inner layer of cartilage in the first stage. In rare cases ossification first commences as a deposit on the exterior of the vertebre. The first vertebra to ossify is the second or third cervical, and the ossification gradually extends backwards. It does 158 THE FOURTH DAY. [CHAP. not commence in the arches till somewhat later than in the bodies. For each arch there are two centres of ossification, one on each side. We may remind the reader that in the adult bird we find between each of the vertebre of a neck and back a cartilaginous disc—the meniscus—which is pierced in the centre. These discs are thick at the circumference but thin off to a fine edge round the central hole. Owing to the shape of these discs there are left between each pair of vertebra two cavities, which only communicate through the central aperture of the meniscus. Through this central aperture there passes a band connecting the two vertebre which is called the ‘ligamentum suspensorium.’ In the tail the menisci are replaced by bodies known as the ‘annuli fibrosi,’ which precisely resemble the similarly named bodies in mammals. They differ from the menisci in being attached over their whole surface to the ends of the vertebral bodies, so that the cavities between the menisci and the vertebre cease to exist. They are pierced however by a body corresponding with the ligamentum suspensorium and known as the ‘nucleus pulposus.’ In the intervertebral regions the chorda, soon after the commencement of ossification, entirely disappears. The cartilage around it however becomes converted (in the region of the neck) into the ligamentum suspensorium, which unites the two vertebra between which it is placed. In the tail the cartilage becomes the nucleus pulposus, which corresponds exactly to the ‘ligamentum suspensorium’ of the neck and. back. Shortly after the formation of the ligamentum suspensorium the remaining cartilage of the intervertebral segments is converted in the neck and back into the meniscus between each two vertebre, and in the tail into the annulus fibrosus. Both are absent in the sacrum. These points together with the anatomy of these parts in the adult were first made out by Jager (Wirbelkérper- gelenk der Véyel. Sitz. der k. Akad. Wien, vol. xxxiI. 1859). In the bodies of the vertebre the notochord does not entirely disappear as in the intervertebral regions, but, according to Gegenbaur, undergoes ultimately a direct conversion into cartilage. The contour of the sheath becomes indistinct ; the cells by the accumulation of matrix round them take on the form of cartilage-cells, so that at the time of the exclusion of the bird from the egg the limits between the altered notochord and the cartilage of protovertebral origin can only with difficulty be made out. 15. While the chief mass of a protovertebra, having given rise to a muscle-plate and a ganglion, is converted into the body and arch of a permanent vertebra with its several appurtenances, a small portion of the exterior grows down- wards as the rudiment for the formation of a rib. These costal growths are of course confined to the dorsal region. They are seen on the sixth day as cartilaginous rods, whose cells are arranged in horizontal rows. By this time they are quite separate from the bodies of the vertebre, with whose arches they are in transverse section seen to alternate. Thus in one section the vertebral arch will be distinctly seen but no trace of the rib; while in the next the rib will be visible but the arch will be absent. VI.] THE MUSCLE-PLATES. 159 16. We shall conclude our account of the protovertebree by describing the changes which take place in the muscle- lates. Jn the chick these are somewhat complicated, and have not been fully worked out. On the third day the muscle-plates end opposite the point where the mesoblast becomes split into somatopleure and splanchnopleure. On the fourth day however (Fig. 47 mp.) they extend to a certain distance into the side walls of the body beyond the point of the division into somatopleure and splanchnopleure. Into what muscles of the trunk they become converted has been somewhat disputed. There is no doubt that it is only epi- skeletal muscles, to use Professor Huxley’s term (Vertebrates, p. 46), that are derived from them, but some embryologists have stated that they only form the muscles of the back. We have, however, little doubt that all the episkeletal muscles are their products ; a view also adopted by Professors Huxley and Kélliker. According to K6lliker the muscle-plates give rise to (1) the deep dorsal muscles, such as the semispinalis multifidus &c., and (2) the visceral muscles as represented by abdominal muscles, the muscles of the breast, the superficial muscles of the neck, and the muscles of the jaws and face. The front dorso-lateral (hyposkeletal) muscles, according to Kolliker, are derived from a front (ventral) muscle-plate, which is formed from the most ventral portion of the protovertebre, but is very limited in extent in the fowl. These muscles include the longus colli, the recti antici, and quadratus. This view differs from that of Huxley, chiefly in considering only the ventral dorsal muscles as hyposkeletal, and uot also the inner visceral muscles. Huxley believes that all the episkeletal muscles are derived from the muscle- plates, but does not give an opinion as to the cells of the embryo from which the hyposkeletal muscles take their origin. His takes an entirely different view ; he believes that the muscles of the back only are derived from the muscle-plates, but that the muscles of the sides and ventral walls of the body are formed from the mesoblast of the somatopleure. There can be little doubt that the intrinsic muscles of the limbs are not cut- growths from the muscle-plates, but are formed independently in the meso- blastic tissues of which the limbs are composed. The origin of the extrinsic limb-muscles is not so certainly known. The cutaneous muscles are obviously derived from the original mesoblast of the somatopleure. It seems very probable (though the subject has not yet been worked out) that the hyposkeletal voluntary muscles underlying the vertebral column are derived from the intermediate cell-mass, which originally lies externally to the protovertebr, but into which, as we have before said, the cleavage of the mesoblast does not extend. In the first instance, as is clear from their mode of origin, the muscle-plates correspond in number with the protovertebre, and this condition is permanent 160 THE FOURTH DAY. [CHAP. in the lower vertebrates, such as fishes, where we find that the lateral muscle is divided by septa into a series of segments corresponding in number with the vertebre, 17. Of all the events of the fourth day, none perhaps are more important than those by which the rudiments of the complex urinary and generative systems are added to the simple Wolffian duct and body, which up to that time are the sole representatives of both systems. We saw that the duct arose on the second day as a solid ridge which subsequently became a tube, lying immediately underneath the epiblast above the intermediate cell-mass, close against the upper and outer angles of the proto- ‘vertebre, and reaching from about opposite to the fifth protovertebra away to the hinder end of the embryo. The exact manner in which it first appears is as yet a matter of dispute, and in our account of the second day, we gave the views of the majority of embryologists who have written on the subject. But it may be considered as quite certain that the Wolffian duct is formed out of mesoblast- cells. It is most probable that the ridge is primarily formed by simple aggregation of cells, and that it is converted into a tube by its central cells taking on a radiating arrange- ment round a central hole, which is at first small but rapidly increases in size. In whatever way it be really formed, we find before the end of the second day, in the place of the previous ridge, a duct with a distinct though small lumen. Waldeyer and some other observers have incorrectly stated that the lumen is not formed till somewhat later. At first the duct occupies a position immediately under- neath the superficial epiblast, but very soon after its forma- tion the growth of the protovertebre and the changes which take place in the intermediate cell-mass, together with the general folding in of the body, cause it to appear to change its place and travel downwards (Chap. v. § 26). While this shifting is going on, the cells lining the upper end of the pleuroperitoneal cavity (the kind of bay which, as seen in sections, is formed by the divergence of the somatopleure and splanchnopleure) become columnar, and constitute a distinct epithelium. This epithelium, which is clearly shewn in Fig. 41, g.e, and is also indicated in Fig. 44, is often called the ger- minal epithelium, because some of its cells subsequently take v1] THE WOLFFIAN BODY. 161 part in the formation of the ovary. Soon after its appearance, the intermediate cell-mass increases in size and begins to grow outwards into the pleuroperitoneal cavity, as a rounded projection which lies with its upper surface towards the somatopleure, and its lower surface towards the splanchno- pleure, but is in either case separated from these layers by a narrow chink. The Wolffian duct (Fig. 44, Wd, 47, Wd) travels down, and finally before the end of the third day is found in the upper part of this projection, near that face of it which is turned towards the somatopleure. At, or before, the fourth day, when the duct occupies this new position, the Wolffian body begins to be formed in the midst of the intermediate cell-mass. The structure of the fully developed Wolffian body is fundamentally similar to that of the permanent kidneys, and consists essentially of convoluted tubules, commencing in Malpighian bodies with vascular glomeruli, and opening into the duct. It is formed as follows. From the anterior portion of each duct and on its inner side, diverticula are given out at right angles. These gradually lengthen, and becoming twisted form the tubules, while the glomeruli of the Malpighian corpuscles seem to be derived from cells of the intermediate mass, which also gives rise to the vascular networks round the tubules. The tubules, which from their contorted course are in sections (Figs. 47, 51) seen cut at various angles, possess an epithelium which is thicker than that of the Wolffian duct. From this difference it is generally easy to distin- guish the sections of the tubules from those of the duct. The glomeruli of the Malpighian bodies are in sections of hardened embryos usually filled with blood-corpuscles. In the above statements we have followed Waldeyer (Hierstock und Fi), but it ought to be mentioned that the majority of earlier observers have believed that the tubules arise independently in the mesoblast, and only at a later period become connected with the duct. The sections which Waldeyer has drawn seem however strongly to support the view which he has brought forward; our own sections also confirm it, and we have noticed that even before the formation of the tubules, the Wolffian duct exhibits great variations in diameter, being in some cases crescent-shaped in section, in others round ; this seems clearly to indicate the giving off of diverticula. Waldeyer’s observations have moreover been since confirmed by other observers. The Wolffian body, as distinct from the duct, reaches from about the level of the fifth protovertebra to beyond the E. 11 162 THE FOURTH DAY. [CHAP. hind limbs; but the duct itself is carried on still further back. Towards the hind end of the embryo, the projection of the intermediate cell-mass spoken of above becomes smaller and smaller, and the Wolffian duct is thus brought nearer to the splanchnopleure, and in the region of the hind-gut comes to lie close to the walls of the alimentary canal. On the fourth day, the two ducts meet and open into two horns, into which the side-walls of the recently formed cloaca are at that time produced, one on either side. As we shall afterwards see, the duct of the permanent kidneys and Miiller’s duct also fall into these two horns of the cloaca. The Wolffian bodies thus constituted perform the offices of kidneys for the greater part of embryonic life. In the chick they disappear before birth; but in most of the Ichthyopsida they remain for life as the permanent kidneys. 18. Near the end of the fourth day, on the outer surface of the projection formed by the Wolffian body a furrow is formed immediately below the Wolffian duct by an involution of the germinal epithelium. This furrow, which is shewn at M.d in Fig. 47, deepens, and its walls arch over and unite. In this way a tube is formed, which separates from the germinal epithelium in the same way that the neural tube separated from the external epiblast. It is known as the Duct of Miller ; of its function we shall speak later on. This account of the origin of Miiller’s duct is due to Waldeyer (loc. cit.), whose observations have been confirmed by subsequent inquirers. An exami- nation of our own sections leads us to the same conclusions. Dr Sernoff (Centralblatt fiir Med. Wiss. 27 Jun. 1874) agrees with Bornhaupt (Untersuchung iiber die Entwickelung des Urino-genrtalsystems beim Hiihnchen) in considering that the duct of Miiller is formed by a simple invo- lution from the pleuroperitoneal cavity which grows backwards in the meso- blast between the Wolffian duct and the germinal epithelium; and thinks that Waldeyer is in error in supposing the involution to be in the form of an elon- gated furrow. This divergence of opinion is not of great importance compared with the point on which both observers are in agreement, viz. that the duct of Miiller is formed by an involution of the germinal epithelium from the pleuro- peritoneal cavity. The formation of the duct of Miiller takes place from before, backwards; but near the hind end of the embryo, where the germinal epithelium is deficient, the groove to form the duct becomes an involution which, at first solid we: THE PERMANENT KIDNEYS. 163 but subsequently hollow, bores its way through the meso- blast, and finally appears to unite on the seventh day with the Wolffian duct close to the entrance of the latter into the cloaca. Later on, this state of things becomes altered; the duct of Miiller opens directly into the cloaca without first uniting with the Wolffian duct. Its opening then lies above that of the Wolffian duct, between it and the opening into the cloaca of the true urinary canal, of which we shall speak directly. The anterior extremity of the duct of Miiller which lies about on a level with the fifth protovertebra is never closed in. Here the original furrow remains open, and forms a funnel-shaped opening into the tube from the _pleuro- peritoneal cavity. In sections of the sixth day the duct of Miiller is to be seen lying between the duct of the Wolffian body and the pleuroperitoneal cavity. Its diameter is generally smaller than that of the Wolffian duct. 19. Between the 80th and 100th hour of incubation, the permanent kidneys begin to make their appearance, and as is the case with the Wolffian bodies, the first portion of them to appear is their duct. Near its posterior extremity the Wolffian duct becomes expanded, and from the expanded portion a diverticulum is constricted off which in a trans- verse section lies above the original duct, and the blind end of which points forwards, that is, towards the head of the chick. This is the duct of the permanent kidney or ureter. At first the ureter and the Wolffian duct open by a common trunk into the cloaca, but this state of things lasts for a short time only, and by the sixth day the two ducts have independent openings. The earlier state of things was overlooked by Remak, who thus came to give an incorrect account of the origin of the duct of the kidneys. Kupffer (Untersuchung iiber die Entwickelung des Harn- und Geschlechts- systems, Archiv fiir Microscop. Anat. Vol. 11. 1866) was the first to give a correct account of the development of the duct of the permanent kidneys in the chick. His observations have since been confirmed by a number of other observers, including Waldeyer. In sections of a somewhat later period the duct of the kidneys can be seen to lie above (dorsal to) the Wolffian duct, and farther from it than the duct of Miiller. The formation of the kidneys themselves is very similar to the formation of the Wolffian bodies. 11—2 164 THE FOURTH DAY. [CHAP. From the upper end of the ureter diverticula are given off at right angles mto the intermediate cell-mass. These lengthening and becoming twisted, form the tubuli wrini- Jeri, while the mesoblast around their extremities becomes directly converted into the Malpighian bodies and the capillary network of the kidneys. Corresponding to the relative position of their ducts, the kidney lies above the Wolffian body. At its first appearance it forms an oval body, lying in the upper part of intermediate cell-mass between the Wolffian body and the vertebral column, and is placed rather nearer the median line than the Wolffian body. The formation of the kidneys takes place before the end of the seventh day, but they do not become of functional importance till considerably later. From their mode of development it clearly follows that the permanent kidneys are merely parts of the same system as the Wolffian bodies, and that their separation from these is an occurrence of a purely secondary importance. 20. Before describing the subsequent fate of the Wolffian and Miillerian ducts, it will be necessary to give an account of the formation of the true sexual glands, the ovaries and testes. At the first appearance of the projection from the inter- mediate mass, which we may now call the genital ridge, a columnar character is not only visible in the layer of cells covering the nascent ridge itself along its whole length, but may also be traced for some little distance outwards on either side of the ridge in the cells lining the most median portions of both somatopleure and splanchnopleure. Passing out- wards along these layers, the columnar cells gradually give place to a flat tesselated epithelium. As the ridge con- tinues to increase and project, the columnar character be- comes more and more restricted to cells covering the ridge itself, in which at the same time it becomes more distinct. On the outer side of the ridge, that is on the side which looks towards the somatopleure, the epithelium undergoes, as we have seen, an involution to form the duct of Miller, and for some little time retains in the immediate neighbourhood of that duct its columnar character (Fig. 51, a’), though eventually losing it, VI] THE GERMINAL EPITHELIUM. 165 The median portion of the ridge is occupied by the projection of the Wolffian body, and here the epithelium rapidly becomes flattened. On the inside of the ridge, however, that is on the side looking towards the splanchnopleure, the epithelium not only retains its columnar character, but grows several cells deep (Fig. 51, a), while at the same time the mesoblast (4) under- lying it becomes thickened. In this way, owing partly to the increasing thickness of the epithelium, and partly to the accumulation of mesoblast beneath it, a slight eminence is formed, which when viewed from above, after opening the SECTION OF THE INTERMEDIATE CELL-MAss oN THE FourtH Day. (From Waldeyer.) Magnified 160 times. m. mesentery. JZ. somatopleure. a’. portion of the germinal epithelium from which the involution to form the duct of Miiller (z) takes place. a. thickened portion of the germinal epithelium in which the primitive ova C and o are lying. #. modified mesoblast which will form the stroma of the ovary. WE. Wolffian body. y. Wolffian duct, 166 THE FOURTH DAY. [CHAP. abdominal cavity, appears in direct light as a fusiform white patch or streak, in its early stages extending along the whole length of the "Wolffian body and genital ridge, but sub- sequently restricted to its anterior portion. Ts appearance under these circumstances has been well described by Von Baer. This ‘sexual eminence’ is present in the early stages of both sexes. In both the epithelium consists of several layers of short cylindrical cells, a few of which are conspicuous on account of their size and their possessing a highly refractive oval nuckeus of considerable bulk; in both, the underlying thickened mesoblast consists—as indeed at this epoch it does generally in all parts of the body—of spindle-shaped cells. The larger conspicuous cells of the epithelium, which appear to have quite a common origin with their fellow cells, and to rise from them by direct differentiation, and which are seen at the first in male as well as female embryvs, are the primordial ova (Fig. 51, 0). Thus in quite early stages it is impossible to detect the one sex from the other. At about the 80th to the 100th hour, however, a distinction becomes apparent. In the males, the epithelium with its underlying meso- blast ceases to develope; the primordial ova neither increase nor multiply: On the contrary, they disappear, and the whole sexual eminence fades away. In females, on the other hand, the primordial ova enlarge and become more numerous, the whole epithelium growing thicker and more prominent. The spindle-shaped cells of the underlying mesoblast also increase rapidly, and thus form the stroma of the ovary. The growth of this stroma bears subsequently such a relation to that of the epithelium, that the primordial ova appear to sink into the stroma, and each ovum, as it descends, to carry with it a number of the ordinary epithelium-cells, which arrange themselves round it in a distinct layer. In this way each ovum becomes invested by a capsule of vascular connective tissue, lined internally by a layer of epithelium; the whole constituting a Graffian follicle. The large nucleus of the primordial ovum becomes the germinal vesicle, while the ovum itself remains as the true ovum; this subsequently becomes enlarged by the ad- dition of a quantity of lee derived from the epithelial lining of the follicle. Vi] THE TESTES. 167 Pfliiger (Die Eierstocke der Stiugethiere u. des Menschen, Leipzig, 1863) de- scribed the ova as arising, in mammals, out of the epithelium of tubular glands, a chain of several ova being frequently found in one tube and the tube be- coming subsequently divided by constrictions into as many follicles. According to Waldeyer however, whose account we have followed above, the primordial ova make their appearance as individual specialized epithelium-cells, without the preformation of any tubular glands, the capsule or Graffian follicle being a later product. Waldeyer’s views have been on the whole generally accepted (Leo- pold, Untersuch. ber das Epithel. des Ovariums. Inaug. Diss. Leipzig, 1870, Romiti, Max Schultze’s Archiv, 1873, Bd. x.), though opposed by Kapff (Rez- chert and Du Bois Reymond’s Archiv, 1872), and more recently by Sernoff (loc. cit.). The first traces of the testes are found in the dorsal and inner side of the intermediate cell-mass, and appear about the sixth day. From the first they differ from the rudimentary ovaries, by coming into somewhat close connection with the Wolffian bodies; but occupy about the same limits from before backwards. The mesoblast in the position we have mentioned begins to become somewhat modified, and by the eighth day is divided by septa of connective tissue into a number of groups of cells; which are the commencing tubuli seminiferi. By the sixteenth day the cells of the tubuli have become larger and acquired a distinctly epithelial character. Waldeyer is of opinion that the tubules of the Wolffian body penetrate into the tissue trom which the testes are formed, and becoming much finer than the remainder of the tubules constitute the ‘tubuli seminiferi.’ Apart from its inherent difficulties, this view has not been corroborated by any subsequent observer. It is distinctly denied by Sernoff (Joc. cit.), who further states that the testes - are entirely formed out of the mesoblast of the intermediate cell-mass, and that tneir rudiments have no connection either with the germinal epithelium or with the tubules of the Wolffian body. We have now described the origin of all the parts which form the urinary and sexual systems, both of the embryo and adult. It merely remains to speak briefly of the changes, which on the attainment of the adult condition take place in the parts described. The Wolffian body, according to Waldeyer, may be said to consist of a sexual and urinary part, which can, he states, be easily distinguished in the just-hatched chick. The sexual part becomes in the cock the after-testes or coni vasculosi, and consists of tubules which lose themselves on the one hand in the seminiferous tubules, and on the other hand, in birds, probably form the whole of what can be called the epididymis. In the hen it forms part of the parovarium of 168 THE FOURTH DAY. [CHAP. His, and is composed of well-developed tubes without pig- ment. The urinary part forms in both sexes a small rudi- ment, consisting of blindly ending tubes with yellow pigment, but is most conspicuous in the hen. The Wolffian duct remains as the vas deferens in the male. In the female it becomes atrophied and nearly dis- appears. The duct of Miller on the right side (that on the left side with the corresponding ovary generally disappearing) remains in the female as the oviduct. In the male it is almost entirely obliterated on both sides. 21. We may return to the changes which art taking place in the circulation. On the fourth day, the point at which the dorsal aorta divides into the two branches which we may now call the alzac arteries is carried much further back towards the tail. A short way beyond the point of bifurcation, each iliac gives off a branch to the newly formed allantois. It is not, however, till the second half of the fourth day, when the allantois grows rapidly, that these allantoic, or as we may now call them umbilical, arteries acquire any importance, if indeed they are present before. With the increase of the allantois they speedily acquire such a size, that the iliac trunks from which they were given off seem to be mere branches of them- selves. The omphalo-mesaraic arteries are before the end of the day given off from the undivided aortic trunk as a single but quickly bifurcating vessel, the left of the two branches into which it divides being much larger than the right. During the third day, we saw that the arterial arch running in the first visceral fold became obliterated, the obliteration being accompanied by the appearance of a new (fourth) arch running in the fourth visceral fold on either side. During the fourth day the second pair of arterial arches also becomes nearly (if not entirely) obliterated; but a new pair of arches is developed in the last (fifth) visceral fold, behind the last visceral cleft; so that there are still three pairs of arterial arches, which however now run in the third, fourth and fifth visceral folds. The last of these is as yet small, and together with the slight remains of the second vI.] THE ARTERIAL ARCHES. 169 pair of arches we may consider that there are in all four pairs of arches. When the first and second arches are obliterated, it is only the central portion of each arch on either side which absolutely disappears. The ventral portion connected with the bulbus arteriosus, and the dorsal portion which joins the dorsal aorta, both remain, and are both carried straight forward towards the head. The ventral portions of both first and second arches unite on each side to form a branch, the external carotid (Fig. 52, £, CA), which runs straight up from the bulbus arteriosus to the head. STATE OF ARTERIAL CIRCULATION ON THE FIFTH oR SrxtH Day. £. CA, external carotid. J. (A. internal carotid. AQ. dorsal aorta. wy. A. arteries to the Wolffian bodies. Ver. A. arteries given off between each of the vertebra. Of. A. omphalo-mesaraic artery. UA. umbilical artery. IA. iliac artery. In the same way the dorsal portions form a branch, the internal carotid, which takes its origin from the dorsal or far end of the third arch. 22. In the venous system important changes also occur. As the liver in the course of its formation wraps round the common trunk of the omphalo-mesaraic veins, or meatus venosus, it may be said to divide that vessel into two parts : 170 THE FOURTH DAY. [CHAP. into a part nearer the heart which is called the sinus venosus (Fig. 53, S.V.), and into a part surrounded by the liver which is called the ductus venosus. Beyond, 7.e. behind the liver, the ductus venosus is directly continuous with the omphalo-mesaraic veins, or as we may now say, vein, for the right trunk has become so small as to appear a mere branch of the left. (Fig. 53, Of.) We saw that on the third day the ductus venosus, while running through the liver, exhibited numerous bulgings Fic. 53. DIAGRAM OF THE VENOUS CIRCULATION AT THE COMMENCEMENT OF THE FIFTH Day. H. heart. D.C. ductus Cuvieri. Into the ductus Cuvieri of each side fall J. the jugular vein, or superior cardinal vein, Su. V. the superior vertebral vein, W. the vein from the wing and C. the inferior cardinal vein. S. V. sinus venosus. Of. omphalo-mesaraic vein. U. umbilical vein, which at this _ stage gives off branches to the body-walls. VI.] THE VEINS OF THE LIVER. aly indicative of branches about to be formed. These are on the fourth day actually formed, and become connected with the capillary network simultaneously developed in the hepatic substance in such a way that those branches which come off from the ductus venosus soon after its entrance between the liver-lobes, carry blood into the substance of the liver, while those which join the ductus venosus shortly before it leaves the liver, carry blood away from the hepatic substance into the ductus. The former are called vene advehentes, the latter vene revehentes. As a result of this arrangement, there is a choice of paths for the blood in passing from the omphalo- mesaraic vein to the sinus venosus; it may pass through the capillary network of the liver, going in by the vene adve- hentes, and coming back again by the venz revehentes, or it may go straight through the ductus venosus without passing at all into the substance of the liver. As the alimentary canal by its continued closing in becomes on the fourth day more and more distinct from the yolk-sac, it gradually acquires veins of its own, the mesenteric veins, which first appear as small branches of the omphalo- mesaraic vein, though eventually, owing to the change in the relative size and importance of the yolk-sac and intestine, the latter seems to be a branch of one of the former. Corresponding to the increase in the size of the head, the superior cardinal veins (Fig. 53, J.) become larger and more important and are joined by the superior vertebral (Su. V.) and wing veins (W). As before, they form the ductus Cuvieri (D.C.) by joining with the cardinal veins (C). The latter are now largely developed; they seem to take origin from the Wolffian bodies, and their size and importance is in direct proportion to the prominence of these bodies. They might be called the veins of the Wolffian bodies. As the kidneys begin to be formed, a new single median vein makes its appearance, running from them forwards, beneath the vertebral column, to fall into the sinus venosus (Fig. 53, V.C.L). This is the vena cava inferior. ‘As the lungs are being formed, the pulmonary veins also make their appearance and become connected with the left side of the auricular division of the heart. The blood carried to the allantois by the umbilical arteries is brought back by two veins which very soon after 172 THE FOURTH DAY. [CHAP. their appearance unite close to the allantois into a single trunk, the wmbilical vein, which, running along the splanch- nopleure, falls into the omphalo-mesaraic vein (Fig. 53, U). 23. Meanwhile the heart is undergoing considerable changes. ‘Though the whole organ still exhibits a marked curvature to the right, the ventricular portion becomes directed more distinctly downwards, forming a blunted cone whose apex will eventually become the apex of the adult heart. The concave (or dorsal) walls of the ventricles become much thicker, as did the convex or ventral walls on the third day. Well-marked constrictions now separate the ventricles from the bulbus arteriosus on the one hand, and from the auricles on the other. The latter constriction is very distinct, and receives the name of canalis auricularis (Fig. 54, C.A.); the former, sometimes called the fretwm Halleri, is far less conspicuous. The most important event is perhaps the formation of the ventricular septum. This, which commenced on the third day as a crescentic ridge or fold springing from the convex or ventral side of the rounded ventricular portion of the heart, now grows rapidly across the ventricular cavity towards the concave or dorsal side. It thus forms an in- complete longitudinal partition extending from the canalis auricularis to the commencement of the bulbus arteriosus, and dividing the twisted ventricular tube into two somewhat Fie. 54. HEART OF A CHICK ON THE FourtTH Day OF INCUBATION VIEWED FROM THE VENTRAL SURFACE. l.a. left auricular appendage. C. A. canalis auricularis, V. ventricle. b. bulbus arteriosus, VI] THE VENTRICULAR SEPTUM. 173 curved canals, one more to the left and above, the other to the right and below. These communicate freely with each other, above the free edge of the partition, along its whole length. Externally the ventricular portion as yet shews no division into two parts. The bulbus arteriosus (Fig. 54, b) has increased in size, and is now very conspicuous. The venous end of the heart is placed still more dorsal, and to the left of the arterial end; its walls are beginning to become thicker. The auricles are nearly if not quite as far forward as the ventricles; and the auricular appendages (Fig. 54, l.a.), which were visible even on the third day, are exceedingly prominent, giving a strongly marked external appearance of a division of the auricular portion of the heart into two chambers; but Von Baer was unable to detect at this date any internal auricular septum. 24, The chief events of the fourth day are :— (1) The increase of the cranial and body flexure. (2) The increase in the tail-fold. (3) The formation of the limbs as local thickenings of the Wolffian ridge. (4) The formation of the olfactory grooves. (5) The absorption of the. partition between the mouth and the throat. _ (6) The formation of the allantois as a diverticulum of the alimentary canal. (7) The formation of the spinal ganglia. (8) The vacuolation of the celis of the notochord. (9) The formation of the Wolffian body. (10) The involution of the germinal epithelium to form the duct of Miiller. (11) The appearance of the primitive ova in the ger- minal epithelium. (12) The development of a fifth pair of arterial arches and the obliteration of the second pair. (13) The origin from the ductus venosus of the capil- laries of the liver. (14) The development of the ‘canalis auricularis,’ the growth of the septum of the ventricles and of the auricular appendages, CHAPTER VII. THE CHANGES WHICH TAKE PLACE ON THE FIFTH DAY. 1. ON opening an egg about the middle of the fifth day, the observer's attention is not arrested by any new features ; but he notices that the progress of development, which was so rapid during the later half of the fourth day, is being con- tinued with undiminished vigour. The allantois which on the fourth day began to project from the pleuroperitoneal cavity has grown very rapidly, and now stretches away from the somatic stalk far over the right side of the embryo (which it will be remembered is lying on its left side) in the cavity between the two amniotic folds (Fig. 8, K). It is very vascular, and already serves as the chief organ of respiration. The blastoderm has spread over the whole of the yolk- sac; and the yolk is thus completely enclosed in a bag whose walls, however, are excessively delicate and easily torn. The vascular area extends over about two-thirds of the yolk. The splanchnic stalk or umbilical duct has now reached its greatest narrowness; it has become a solid cord, and will undergo no further change till near the time of hatching. The space between it and the somatic stalk is still con- siderable, though the latter is narrower than it was on the fourth day. 2. The embryo remains excessively curved, so much so indeed that the head and the tail are nearly in contact. The limbs have increased, especially in length; in each a distinction is now apparent between the more cylindrical CHAP. VII.] THE LIMBS. 175 stalk and the flattened terminal expansion ; and the carti- laginous precursors of the several bones have already become visible. The fore and hind limbs are still exceedingly alike, and in both the stalk is already beginning to be bent about its middle to form the elbow and knee respectively. The angles of both knee and elbow are in the first instance alike directed outwards and somewhat back- wards. By the eighth day, however, the elbow has come to look directly backwards and the knee forwards. In consequence of this change, the digits of the fore limb point directly forwards; those of the hind limb directly backwards. This state of things is altered by a subsequent rotation of the hand and foot on the arm and leg, so that by the tenth day the toes are directed straight forwards, and the digits of the wing, backwards and somewhat downwards, the elbow and knee almost touching each other. While these changes are taking place, the differences between wing and foot become more and more distinct. The cartilages of the digits appear on the fifth day as streaks in the broad flat terminal expansions, from the even curved edge of which they do not project. On the sixth or seventh day the three digits of the wing (the median being the longest), and the four (or in some fowls five) digits of the foot may be distinguished, and on the eighth or ninth day these begin to project from the edge of the expanded foot and wing, the substance of which, thin and more or less transparent, remains for some time as a kind of web between them. By the tenth day, the fore and hind extremities, save for the absence of feathers and nails, are already veritable wings and feet. At an early period of development we find the following elements in the avian manus, as separate masses of cartilage. In the carpus there are four elements. Two in the proximal row which remain distinct through life, viz. (1) the radiale, (2) the united intermedium and ulnare. In the distal row (according to some recent observations of Dr Rosenberg, Zeitschrift fiir Wiss. Zoologie, 1873, p. 139, etc.) there are also two elements. One of these is the united first and second carpal which we may call carpal'—", and the other is the united third and fourth carpal which we may call carpal’, These subsequently unite with the metacarpal bones, and form with them a united ‘carpo-metacarpus’ comparable with the tarso-metatarsus of the avian foot. + Four metacarpals are present, viz. the first, second, third, and fourth. The 176 THE FIFTH DAY. [CHAP: first, second, and third are the usually recognized elements, and to these Dr Rosenberg’s investigations (loc. cit.) have added a fourth. The first, second and third persist in the adult, though they become anchylosed in all recent birds. They also fuse, as we have said above, with the distal row of the carpals. Phalanges belonging to the first, second, and third metacarpals are present. There thus seem in the avian manus to be no representatives of the centrale, the fifth carpal, the fifth metacarpal and the phalanges of the fourth and fifth digits. “Of the elements we have spoken of in the avian hand, the only ones which require further notice are the carpal'—", carpal™'—, and the fourth meta- carpal. The united first and second carpal first appears as a small mass of cartilage close to the proximal end of the second metacarpal. In this condition it persists for some time but commences finally to fuse with the first metacarpal ; and at a slightly subsequent period with the second metacarpal. These rela- tions with the first and second metacarpals shew without doubt that this little mass of cartilage is the representative of the first and second bones of the distal row of the carpus. In a still later stage carpal! fuses also with carpal!4V, Tts distinct nature as a separate element in the bird’s manus is again shewn during ossification, when there appears for it a separate centre of ossification. Carpal” appears about the same time as carpal’ but is at its first appear- ance united with metacarpals three and four; it soon becomes separated from metacarpal three, and afterwards also from metacarpal four. It subsequently undergoes considerable changes of shape, and rather later fuses with carpal}. Its true nature is again, as with carpal’ shewn during ossification by the appearance, of a separate centre of ossification for it. The fourth metacarpal is, as we have described, at first united with carpal, but subsequently the neck connecting the two becomes constricted, and finally they become completely separated from each other. The small independent mass of cartilage thus formed represents the fourth metacarpal; it applies itself closely to the side of the third metacarpal, though without becoming united with it. It ossifies very late—some time after the hatching of the chick, and after ossification fuses with the third metacarpal—and then in most cases disappears completely. The pes of a fowl in its early embryonic condition consists of (t) a mass of cartilage close to the distal end of the tibia. It represents (Gegenbaur) the proximal row of tarsal bones, viz. the ‘tibiale,’ the ‘inter- medium,’ the ‘fibulare,’ and the ‘centrale.’ This cartilage fuses in the adult with the distal end of the tibia. (2) a mass of cartilage representing the five bones of the distal row of the tarsus. In the adult this unites with the metatarsus, forming a tarso- metatarsus. (3) the metatarsus. There are usually stated to be four metatarsal bones present in the metatarsus of a fowl, which are anchylosed in the adult, but aré represented by separate rods of cartilage in the embryo. They are the distal extremity of a first metatarsal, and complete second, third and fourth metatarsal bones. In addition to these Dr Rosenberg (loc. cit.) has found a small oval mass of cartilage representing a fifth metatarsal. Soon after its appearance this becomes fused with the end of the tarsal mass of cartilage representing the fifth tarsal, but later entirely atrophies. (4) There are four phalanges present both in the embryo and the adult, a number which is never exceeded in birds (except amongst some abnormal breeds of fowls, e.g, the Dorking fowls) ; though one or more of the four are frequently deficient. VIL] THE INVESTING MASS, Va 3. As we mentioned in the last chapter, the formation of the primitive cranium commenced upon the fourth day. This in its earliest stage, inasmuch as it is composed of con- densed but otherwise only slightly differentiated mesoblast, may be spoken of as the membranous cranium. On the sixth day, true hyaline cartilage makes its ap- pearance; and the primitive membranous cranium gives place to the primitive cartilaginous cranium. The cartilage which is the first to appear, forms a thick plate called the investing mass of Rathke (Fig. 55, zv.), sur- rounding the whole of that portion of the notochord which projects in front of the foremost protovertebra. The hinder Fic. 55. VIEW FROM ABOVE OF THE INVESTING MASS AND OF THE TRABECULZ ON THE FourTH Day oF IncuBation. (From Parker.) In order to shew this, the whole of the upper portion of the head has been sliced away. ‘The cartilaginous portions of the skull are marked with the dark horizontal shading. cv. I. cerebral vesicles (sliced off). e¢. eye. me. notochord. iw. investing mass. g. foramen for the exit of the ninth nerve. cl. cochlea. hsc. horizontal semicircular canal. g. quadraie. 5. notch for the passage of the fifth nerve. lg. expanded anterior end of the investing mass. pts. pituitary space. tr. trabecule. The reference line ¢r. has been accidentally made to end a little short of the cartilage, E, 12 178 THE FIFTH DAY. [CHAP. portion of this investing mass sends upwards along the sides of the brain two lateral projections or wings, which enclose the rudiments of the internal ear. In the chick the portions which thus inclose the auditory sacs seem never to be at any time separate from the remainder of the investing mass. At the front end of the notochord the cartilaginous investing mass divides into two horizontal branches in the form of two cartilaginous rods called the trabecule (Fig. 55, tr.), which passing forward (in a somewhat different plane from the investing mass), meet again in front, and so enclose a space called the pituitary space pts, into which the infundibulum extends downward. In front of this junction, the trabeculz expand into a somewhat broad plate (subsequently developed into the ethmoid and nasal cartilages), which ends in two horns in the interior of the fronto-nasal process. The front end of the notochord probably defines the anterior boundary of the basi-occipital. At first it extends quite up to the pituitary space and the starting-point of the trabecule. Subsequently, however, there takes place be- tween it and the pituitary space a growth of cartilage in which the ossification for the basi-sphenoid takes place. The lateral projections at the hinder end of the investing mass grow up behind, and completely enclose that part of the neural canal from which the medulla oblongata is de- veloped, and in it ossifications arise to form the occipital bones and the bones which invest the auditory labyrinth. It is important to notice that the only segment of the skull, which primarily forms a cartilaginous roof to any part of the brain, is the occipital segment. The roof of the re- mainder of the skull is formed by membrane-bones. For the histological differences observable in the develop- ment of cartilage and membrane bones, we must refer the reader to treatises on histology; for our purpose it is sufficient to say that a membrane-bone is one which is not preformed in cartilage, while a cartilage-bone is one in which the ossification takes place in a bed of cartilage, which fills the place subsequently occupied by the bone. The trabecule together with the cartilage between the pituitary space and the end of the notochord give rise to the sphenoid bone, while in the cartilage in front of the trabecul the ethmoid and nasal bones are formed. vi1.] THE VISCERAL ARCHES. 179 From the study of the development of the skull, especially in some of the lower vertebrates, Mr Parker and Professor Huxley have shewn, that the trabecule are developed independently of the investing mass, and that their subsequent connection with it is due to a secondary process. Professor Huxley is of opinion that they are to be regarded as the remains of a pair of visceral arches, corresponding with the other five pairs of arches which we find developed in the chick.. The stage in which they exist as simple visceral arches with a core of undifferentiated mesoblast is not seen in the chick. They first attract notice when they become cartilaginous rods. The ordinary visceral arches are, as we have seen, suffici- ently obvious, while as yet their mesoblast is quite undiffer- entiated ; but in them, as in the trabeculz, rods of cartilage are subsequently developed and begin to make their appear- ance about the fifth day. The first arch, it will be remembered, budded off a process called the superior maxillary process. The whole arch, therefore, comes to consist of two parts, viz. a superior and an inferior maxillary process; in each of these, carti- laginous rods are developed. In the superior maxillary process, the rod does not appear till the fifth day. It is called from its subsequent fate, the pterygo-palatine rod, and consists of a pterygoid and of a palatine part. In the inferior maxillary process two developments of cartilage take place; one which forms the guadrate in the upper or prox- imal portion close to the origin of the superior maxillary process, a second in the lower or distal portion, which goes by the name of Meckel’s cartilage. Cartilaginous rods are also formed in the second and third arches. These, which give rise to the hyoids and branchials respectively, quickly come to lie within the first arch, but do not form a conspicuous portion of the skeleton of the face. 4. Closely connected with the development of the skull is the formation of the parts of the face. After the appearance of the nasal grooves, on the fourth day the mouth (Fig. 56 M.) appears as a deep depression inclosed by five processes. Its lower border is entirely formed by the two inferior maxillary processes (Fig. 56, F.1), at its sides lie the two superior maxillary processes 8. I, while above it is bounded by the fronto-nasal process nf. After a while the cuter angles of the fronto-nasal process, enclosing the termination of the ethmovomerine plate, pro- ject somewhat outwards on each side, giving the end of the 12—2 180 THE FIFTH DAY. [CHAP. Fic. 56. A. HEAD oF AN Empryo CHICK OF THE FourTH DAY VIEWED FROM BELOW AS AN OPAQUE OBJECT. (Chromic acid preparation.) OH. cerebral hemispheres. FB. vesicle of the third ventricle. Op. eyeball. nf. naso-frontal process. M. cavity of mouth. S. M. superior maxillary process of /. 1, the first visceral fold (inferior maxillary process). F. 2, F. 3, second and third visceral folds. NV. nasal pit. In order to gain the view here given the neck was cut across between the third and fourth visceral folds. In the section e thus made, are seen the alimentary canal al with its collapsed walls, the neural canal m.c., the noto- chord ch., the dorsal aorta A0O., and the vertebral veins V. The incision has been carried just below the upper limit of the pleuroperi- toneal cavity, consequently a portion of the somatopleure appears at the angle hetween the two third visceral folds. Almost embraced by the piece of somato- pleure is seen the end of the bulbus arteriosus Ao. In the drawing the nasal groove has been rather exaggerated in its upper part. On the other hand the lower part of the groove, where it runs between the superior maxillary process S. M. and the broad naso-frontal process, was in this particular embryo extremely shallow and indeed hardly visible. Hence the end of the superior maxillary process seems to join the inner and not, as described in the text, the outer margin of the nasal groove. A few hours later the separation of the two would have been very visible. B. The same seen sideways, to shew the visceral folds. Letters as before. process a rather bilobed appearance. These projecting portions of the fronto-nasal process form on each side the inner margins of the rapidly deepening nasal grooves, and are sometimes spoken of as the znner nasal processes. The outer margin of each nasal groove is raised up into a projection frequently spoken of as the outer nasal process which runs downwards to join the superior maxillary process, frem which, bse vul.| THE NASAL LABYRINTH. 181 however, it is separated by a shallow depression. This de- pression, which runs nearly horizontally outwards towards the eyeball, is, according to Coste and Kolliker, subsequently con- verted into the lachrymal duct. On the fifth day, the inner nasal processes or lower and outer corners of the fronto-nasal process arching over, unite on each side with the superior maxillary processes. (Com- pare Fig. 57, which, however, is a view of the head of a chick of the sixth day.) In this way each nasal groove is converted into a canal, which leads from the nasal pit above, into the cavity of the mouth below, and places the two in direct communication. This canal, orliase lining consists of epiblast, is the rudiment of the nasal labyrinth. HEAD oF A CHICK AT THE SrxtH Day FROM BELOW. (Copied from Huxley’s Elements of Comparative Anatomy.) Ia. cerebral vesicles. a. eye, in which the remains of the choroid slit can still be seen. g. nasal pits. &. fronto-nasal process. J. superior maxillary process. I. inferior maxillary process or first visceral arch. 2. second visceral arch. «x. first visceral cleft between the first and second visceral arches. The cavity of the mouth is seen enclosed by the fronto-nasal process, the superior maxillary processes and the first pair of visceral arches. At the back of it is seen the opening leading into the throat. The nasal grooves leading from the nasal pits to the mouth are already closed over and converted into canals. By the seventh day (Fig. 58), not only is the union of the superior maxillary and fronto-nasal processes completed, and the upper boundary of the mouth thus definitely con- 182 - THE FIFTH DAY. [CHAP. stituted, but these parts begin to grow rapidly forward, thus deepening the mouth, and giving rise to the appearance of a nose or beak (Fig. 58), which, though yet blunt, is still distinct. The whole of the lower boundary of the buccal cavity is formed by the inferior maxillary processes. bea \ HEAD oF A CHICK OF THE SEVENTH Day FROM BELOW. (Copied from Huxley’s Elements of Comparative Anatomy.) I a. cerebral vesicles. a. eye. g. nasal pits. /. fronto-nasal process. J. superior maxillary process, 1. first visceral arch. 2. second visceral arch. «. first visceral cleft. The external opening of the mouth has become much constricted, but it is still enclosed by the fronto-nasal process and superior maxillary processes above, and by the inferior maxillary process (first pair of visceral arclies) below. The superior maxillary processes have united with the fronto-nasal process, along the whole length of the latter, with the exception of a small space in front, where a narrow angular opening is left between the two. As we have before mentioned, the ethmovomerine car- tilage is developed in the fronto-nasal process, the pterygo- palatine bar in the superior maxillary process, Meckel’s cartilage and the quadrate in the inferior maxillary process ; the other bones which form the boundaries of the mouth in the adult are developed later after all external trace of these parts as separate processes has disappeared. For an account of their formation, however, we must refer the reader to the chapter upon the development of the skull. At first the mouth is a simple cavity into which the nasal canals open directly. When however the various VII. | THE MOUTH. 185 processes unite together to form the upper boundary of the mouth, each superior maxillary process sends inwards a lateral bud. These buds become flattened and form horizontal plates which stretch more and more inward towards the middle line. There they finally meet, and by their union, which is effected first in front, and thence extends backwards, they con- stitute a horizontal plate, stretching right across the mouth, and dividing it into two cavities—an upper and a lower one. In the front of the mouth their union is quite complete, so that here there is no communication between the two cavities. Behind, however, the partition is not a complete one; so that the two divisions of the buccal cavity com- municate at the back of the mouth. The external opening of the mouth passes into the lower of these two cavities, which may therefore be called the mouth proper. Into the upper chamber the nasal ducts open; it may be called the respiratory chamber and forms the commencement of the chamber of the nose. In birds generally the upper nasal cavity becomes subsequently divided by a median partition into two chambers, which communicate with the back of the mouth by separate apertures. The original openings of the nasal pits remain as the nostrils. 5. One important occurrence of the fifth day is the appearance of the anus, which is formed very much in the same way as the mouth. Beneath the tail an involution of the epiblast takes place towards the cloaca. At this point the wall of the cloaca, which has here taken no share in the cleavage of the meso- blast, becomes thinner, and is finally perforated. An orifice thus places the cloaca in communication with the exterior, and constitutes the anus. 6. On this day also important changes take place in the spinal cord; and a brief history of the development of this organ may fitly be introduced here. At the beginning of the third day, the cavity of the neural canal (Fig. 41) is still of considerable width, and when examined in vertical section its sides may be seen to be nearly parallel, though perhaps approximating to each other more below than above. We say below and above, because a vertical section is naturally examined with its dorsal side uppermost. In the ordinary terminology of the spinal cord, 184 THE FIFTH DAY. [CHAP. above would be posterior and below anterior. These latter terms it will be henceforward most convenient to adopt. The exact shape, however, varies according to the region of the body from which the section is taken. The epiblast walls are at this time composed of radiately arranged columnar cells. The cells are much elongated, but somewhat irregular; and it is very difficult in sections to make out their individual boundaries. They contain granu- lar oval nuclei in which a nucleolus can almost always be seen. The walls of the canal are both anteriorly and pos- teriorly considerably thinner than in the middle. Towards the end of the third day, changes take place in the shape of the cavity. In the lumbar region its vertical section becomes more elongated, and at the same time very narrow in the middle while expanded at each end into a some- what bulbous enlargement, producing an hour-glass appear- ance (Fig. 44). Its walls however still preserve the same histological characters as before, : On the fourth day (Fig. 47) coincidently with the appear- ance of the spinal nerves, important changes may be observed in the hitherto undifferentiated epiblastic walls. In the anterior region of the cord, the external portions of the epiblast become modified into grey matter, forming an anterior grey column, which in turn is covered superficially by a mass of white matter forming an anterior white column. The internal portions of the epiblast remain as the epithe- lium lining the spinal canal. Both columns are formed at the point of entrance of the anterior nerve-roots; and these may easily be traced through the white into the grey matter. The grey column is composed of numerous small nuclei, each of which appears to be surrounded by a definite mass of protoplasm, though the boundaries of the protoplasm belong- ing to each nucleus can only occasionally be made out. The nuclei le in the meshes of a network of fibres continu- ous with the fibres of the nerve-root, and passing through the mass of grey matter in two directions :—(1) round the anterior end of the spinal canal, immediately outside its epithelium and so to the grey matter on the opposite side, forming in this way an anterior commissure through which a decussation of the fibres from the opposite sides is effected : (2) upwards along the outside of the lateral walls of the canal, VII. THE SPINAL CORD. 185 The posterior roots of the spinal nerves enter the cord near its posterior surface, and at this point the posterior grey columns are formed in a Similar way to the anterior. In some cases also the rudiment of a posterior white column may be seen at the junction of the nerve with the epiblast of the canal. The fibres of the posterior root cannot be traced so far into the cord as those of the anterior root. The grey matter of the cord seems undoubtedly to be formed by a meta- morphosis of the external cells of the epibiast of the neural tube, and is directly continuous with the epithelium; there being no strong line of demarcation between them. Whether the fibres which traverse it, and which seem to be partly nervous and partly connective tissue in their nature, are derived from mesoblast or epiblast our observations have not enabled us to determine. The white matter which caps the grey mass, and which forms the com- mencement of the anterior white column, is a peculiar tissue. It consists of a network of fibres somewhat resembling the connective tissue network of the white matter of the adult cord, to which it has a further likeness in not being easily stained by carmine. The fibres of which it is composed have a general tendency to be disposed in radiating septa, a peculiarity which is especially noticeable with low powers. Along the fibres and more especially in the septa, numerous highly refracting granules are embedded, and in the meshes pale spherical nuclei with nucleoli are to be seen. The boundary between the white and grey matter is very sharply defined, and we have always failed to trace the fibres of which we are speaking into the fibres present in the grey matter, though Lockhart Clarke (Phil. Trans. 1862) asserts that they are continuous. Nor can the fibres of the nerve-roots be seen to come into connection with these same fibres. It has generally been assumed that the white matter like the grey is derived from the epiblast: this does not however appear ever to have been clearly proved, while the peculiarities of the tissue, and the fact that it first appears at the origin of the spinal nerves, might seem to indicate that it is directly derived from the mesoblast surrounding the cord ; a view which we are inclined to accept. On the fourth day there is no trace of either an anterior or a posterior fissure, and in the lumbar region the shape of the spinal canal is not very different from what it was on the third day. It appears in sections as a narrow slit dilated somewhat at either end (Fig. 47). The epithelium surround- ing the slit is still very thin, especially above and below, but at the anterior end forms a somewhat arched projection with the convex surface turned downwards. On the fifth and sixth days important changes take place. By the great increase of the grey matter, which now comes to form the chief mass of the cord, the epithelium is reduced to a thin layer of cells immediately surrounding the canal, In the dorsal region, the side walls of the laterally com- 186 © THE FIFTH DAY. [CHAP. pressed canal come into absolute contact in the middle. So that sections no longer shew an hour-glass cavity, but two more or less elliptical cavities, représenting the former term- inal enlargements, one anterior and one posterior, separated by a neck in which the epithelium of the one side is closely applied to that of the other. In other words, the original single canal has been divided longitudinally into an anterior and posterior canal. Of these the anterior will alone remain as the permanent central canal of the spinal cord. In the lumbar region this division has as yet not taken place. The anterior white columns have very much increased in quantity ; the posterior white columns have also become distinct, and the two form together a thick covering for the grey matter. The two columus of each side are continuous with each other, but their line of junction is clearly marked ; and on the sixth day there may be seen at this spot a small mass of white matter, differing somewhat from the rest in appearance, which perhaps may be looked upon as the first commencement of the lateral column. The columns of the one side are not continuous with those of the other either posteriorly or anteriorly. In other words, there are as yet no white commissures. The anterior ends of the cord on each side of the middle line have commenced to grow downwards. These outgrowths, in which both the white and the grey matter take part, have an important function. They enclose between them a somewhat linear space: the commencement of the anterior fissure. This, which is at first not very deep and rather wide, may be noticed already on the fifth day (L. Clarke) and on the sixth day is very clearly marked. Corresponding with these grosser changes, certain histo- logical features make their appearance. Between the an- terior and posterior parts into which the grey matter is divided on each side, or, as we may now call them, the anterior and posterior cornua, there is found a rather lighter hand of grey matter in which the nuclei are somewhat more seattered. ‘he anterior cornu exhibits a further division into an outer and upper part, and a lower and inner part, in both of which the nuclei are more numerous than in the intervening mass. The posterior cornu is of considerably darker colour than the anterior, the difference being due to evil.| THE, POSTERIOR FISSURE. 187 the greater number of nuclei present in the former. The outlines of the cells are more clearly marked and somewhat more angular in shape than they were on the fourth day. The distinctions between the several parts of the grey matter are chiefly brought about by variations in the number of nuclei in a given area. Throughout the cord fibres of the grey matter seem to be continuous with the epithelium of the neural canal, but this is much more strongly marked in the posterior than in the anterior region. In the posterior region also, it is still much more difficult to trace the routs of the nerves than in the anterior, Of the three columns into which the white matter on each side is divided, the anterior column differs from the posterior in being thicker and also in having wider meshes and fewer granules. The lateral column is the most granular of all and very conspicuous. The minute structure of the white matter remains about the same as on the fourth day. Meanwhile an alteration is taking place in the external outline of the cord. From being, as on the fourth and fifth days, oval in section, it becomes, chiefly through the increase of the white matter, much more nearly circular. On the seventh day the most important event is the formation of the posterior fissure. This is brought about by the absorption of the roof of the posterior of the two parts into which the neural canal has become divided. Between the posterior horns of the cord, the epithelium forming the roof of the, so to speak, posterior canal is along the middle line covered neither by grey nor by white matter, and on the seventh day is partially absorbed, thus transforming the canal into a wedge-shaped fissure, whose mouth however is seen in section to be partially closed by a triangular clump of elongated cells (Fig. 59 c.). Below this mass of cells the fissure is open. It is separated from the ‘true spinal canal’ by a very narrow space along which the side walls have coalesced. In the lumbar and sacral regions the two still communicate. We thus find, as was first pointed out by Lockhart Clarke, that the anterior and posterior fissures of the spinal cord are, morphologically speaking, entirely different. ‘Vhe ante- rior fissure is merely the space left between two lateral downward growths of the cord, while the posterior fissure is part of the original neural canal separated from the rest of 188 THE FIFTH DAY. [CHAP. Fig. 59. acw SECTION THROUGH THE SPINAL CorD oF A SEVEN Days’ CHIcK. p.c.w. posterior white column, J.c.w. lateral white column. a. c.w. anterior a. f. a. 9. white column. p c. posterior cornu, of grey matter, consisting of very small cells. a.c. anterior cornu of grey matter, with a peculiar mass of very large cells. ep. epithelium lining the original medullary canal. p. f. pos- terior fissure. The posterior fissure is chiefly formed by the upper portion of the original medullary canal which becomes open above. The upper portion of it is now filled with tissue (c) which is probably derived from the epithelium of the medullary canal. The lower portion of the medullary canal becomes the spinal canal (sp.c.) and is eventually entirely shut off from posterior fissure. The communication between the spinal canal and the posterior fissure is already narrowed, and if the section had been made further forwards, the two would have been entirely separated from each other. anterior fissure. This is formed in an entirely different manner from the posterior fissure. It is produced by the anterior column of white, and the anterior cornu of grey matter, growing downwards and leaving between them a fissure. It is at this time filled up with connective tissue. ¢. anterior grey commissure. cc. tissue filling up the end of the posterior fissure. sp.c. spinal canal. Only the right half of the cord is represente:l in the figure. The section passes through the cord between the entrance of two spinal nerves. The angular form of the cells of the cord has not been done justice to by the engraver. VII.] THE WHITE COLUMNS. 189 the cavity (which goes to form the true spinal canal) by a median coalescence of the side walls. The lateral white columns have on the seventh day increased in size and become less granular, and the lines of junction between them and the anterior columns have now to be arbitrarily selected. The posterior white columns are still much thinner and more granular than the anterior. The nuclei of the white matter are more numerous than before. Some of the septa of the white matter can now be traced in the one direction into the grey matter, and in the other direction into the connective tissue around the cord. Whether these are nerve-fibres which have separated trom the remainder of the fibres, to enter the cord at a different point, or are merely trabecule of connective tissue, cannot be absolutely determined. The latter view however seems most probable. In the grey matter, the anterior and posterior divisions are better distinguished than at an earlier date. In particular the nuclei of the cells of the posterior division are both smaller and more numerous than those of the anterior. Some ot the fibres from the posterior root, after entering the grey matter, quickly pass out again into the posterior column of the white matter. In the anterior division of the grey matter, near the entrance of the anterior roots, there is a peculiar and well-marked mass of somewhat triangular cells, with large and distinct nuclei, more deeply stained with carmine than the remainder of the grey matter. This mass of cells is present in the lumbar and sacral regions, but is deficient or very inconspicuous in the dorsal portion of the cord. The nuclei of the whole anterior region of the grey matter have increased in size, and the cells to which they belong (when clearly visible) are usually jound to be angular. Around the true spinal canal, the line of separation between the epithelium and the grey matter is sharply defined, but elsewhere is very indistinct. By the end of the seventh day, the following important parts of the cord have been definitely established : (1) The anterior and posterior fissures. (2) The anterior and posterior horns of grey matter. (3) The anterior, posterior and lateral columns of white matter. (4) The spinal canal. As yet, however, the grey mass of the two sides of the cord only communicate by the anterior grey commissure, and the white columns of opposite sides do not communicate at all. The grey matter, moreover, still far preponderates over the white matter in quantity. By the ninth day the posterior fissure is fully formed, and the posterior grey commissure has also appeared. In the centre of the sacral enlargement this commissure is absent, and the posterior columns at a later period separate 190 . THE FIFTH DAY. [CHAP. widely and form the ‘sinus rhomboidalis,’ which is not, as has been sometimes stated, the remains of the primitive ‘sinus rhomboidalis’ visible during the second day. The anterior white columns have much increased on this day, and now form the sides of the already deep anterior fissure. The anterior white commissure does not however appear till somewhat later. 7. The fifth day may perhaps be taken as marking a most important epoch in the history of the heart. The changes which take place on that and on the sixth day, added to those previously undergone, transform the simple tube of the early days of incubation into an almost com- pletely formed heart. The venous end of the heart, though still lying somewhat to the left and above, is now placed as far forwards as the arterial end, the whole organ appearing to be drawn together. The ventricular septum is complete. The apex of the ventricles becomes more and more pointed. In the auricular portion a small longitudinal fold appears as the rudiment of the auricular septum, while in the canalis auricularis, which is now at its greatest length, there is also to be seen a commencing transverse partition tending to separate the cavity of the auricles from those of the ventricles. About the 106th hour, a septum begins to make its appearance in the bulbus arteriosus in the form of a longitu- dinal fold, which according to Dr Tonge (Proc. of Royal Soc. 1868) starts, not (as Von Baer thought) at the end of the bulbus nearest to, but at that furthest removed from, the heart. It takes origin from the wall of the bulbus between the fifth and fourth pairs of arches and grows downwards in such a manner as to divide the bulbus into two channels, one of which leads from the heart to the fourth and third pair of arches and the other to the fifth pair. The free edge of the septum is somewhat V-shaped, so that its two legs as it were project downwards towards the heart, further than its central portion; and this shape of the free edge is maintained during the whole pericd of its growth. Its course downwards is not straight but spiral, and thus the two channels into which it divides the bulbus arteriosus, wind spirally the one over the other. The existence of the septum can only be Wit] ’ THE SEMILUNAR VALVES. 197 ascertained at this stage by dissection or by sections, there being as yet no external signs of the division. At the time when the septum is first formed, the opening of the bulbus arteriosus into the ventricles is narrow or slit- like, apparently in order to prevent the flow of the blood back into the heart. Soon after the appearance of the septum, however, semilunar valves (Tonge, loc. cit.) are developed from the wall of that portion of the bulbus which lies between the free edge of the septum and the cavity of the ventricles. These arise as six solid outgrowths of the wall arranged in pairs, an anterior, an inner, and an outer pair, one valve of each pair belonging to the one and the other to the other of the two main divisions of the bulbus which are now being established. The anterior and the inner pairs of valves are the first to _ appear: the former as two small solid prominences separated from each other by a narrow groove; the latter as a single shallow ridge, in the centre of which is a prominence indi- eating the point where the ridge will subsequently become divided into two. The outer pair of valves appear opposite each other, at a considerably later period, between the ends of the other pair of valves on each side. As the septum grows downwards towards the heart, it finally reaches the position of these valves. One of its legs then passes between the two anterior pair of valves, and the other unites with the prominence on the inner valve-ridge. At the same time the growth of all the parts causes the valves to appear to approach the heart and thus to be placed quite at the top of the ventricular cavities. The free edge of the septum of the bulbus now fuses with the ventricular septum, and thus the division of the bulbus into two separate channels, each provided with three valves, and each com- municating with a separate side of the heart, is complete, the position of the valves not being very different from what it is in the adult heart. That division of the bulbus which opens into the fifth pair of arches is the one which communicates with the right ventricle, while that which opens into the third and fourth pairs communicates with the left ventricle. The former becomes the pulmonary artery, the latter the commencement of the systemic aorta. 192 THE FIFTH DAY. [CHAP. The external constriction actually dividing the bulbus into two vessels, does not begin to appear till the septum has extended some way back towards the heart. The semilunar valves become pocketed at a period con- siderably later than their first formation (from the 147th to the 165th hour) in the order of their appearance. ' 8. Towards the end of the fifth and in the course of the sixth day further important changes take place in the heart. The venous end with its two very conspicuous auricular appendages, comes to be situated more above (dorsal to) the arterial end, though it still turns rather towards the left. The venous portion of the heart undergoes on the sixth day or even near to the end of the fifth, ach a development of the muscular fibres of its walls, that the canalis auricularis becomes almost entirely concealed. ‘he point of the heart is now Fic. 60. Two VIEWS OF THE Hrart oF A CHICK UPON THE FirtH Day OF INCUBATION, A from the ventral, B from the dorsal side. i.a. left auricular appendage, r.a. right auricular appendage, 7.v. right ventricle, 1.v. left ventricle. &, bulbus arteriosus, Vil. ] THE BULBUS ARTERIOSUS. 193 directed nearly backwards (7.e. towards the tail), but also a little downwards. An alteration takes place during the sixth day in the relative position of the parts of the ventricular division of the heart. The right ventricle is now turned towards the abdominal surface, and also winds to a certain extent round the left ventricle. It will be remembered that on the fourth day the right ventricle was placed above (dorsal to) the left. The right ventricle is now also the smaller of the two, and the constriction which divides it from the left ventricle does not extend to the apex of the heart (Fig. 60). It has, however, a very marked bulge towards the right. At first the bulbus arteriosus appeared to come off chiefly from the left ventricle; during the fifth day, and still more on the sixth, it appears to come from the right chamber. This is caused by the canal from the right ven- tricle into the bulbus arteriosus passing towards the left, and on the ventral side, so as entirely to conceal the origin of the canal from the left chamber of the heart. On the seventh day the bulbus arteriosus appears to come less markedly from the right side of the heart. All these changes, however, of position of the bulbus arteriosus only affect it externally; during the whole time the two chambers of the heart open respectively into the two divisions of the bulbus arteriosus, The swelling of the Fic. 61. Heart of a CHIcK UPON THE SixtH Day oF INCUBATION, FROM THE VENTRAL SURFACE. 1. a. left auricular appendage, .a. right auricular appendage, 7,v.right ventricle, 1. v. left ventricle, b, bulbus arteriosus, ~ E, 13 194 THE FIFTH DAY. [CHAP. bulbus is much less marked on the seventh day than it was before. At the end of the sixth day, and even on the fifth day (Figs. 60, 61), the appearance of the heart itself, without reference to the vessels which come from it, is not very dissimilar from that which it presents when adult. The original curvature to the right now forms the apex of the ventricles, and the two auricular appendages are placed at the anterior extremity of the heart. The most noticeable difference (in the ventral view) is the still externally undivided condition of the bulbus arte- riosus. About the sixth or, perhaps, even on the fifth day, the pericardium, according to Von Baer, makes its appearance. | Its mode of formation is not exactly known, but it probably takes origin from folds of the lining of the thoracic cavity which meet and coalesce. 9. The subsequent changes which the heart undergoes are concerned more with its internal structure than with its external shape. Indeed, during the next three days, viz. the eighth, ninth, and tenth, the external form of the heart re- mains nearly unaltered. In the auricular portion however, the septum which com- menced on the fifth day becomes now more conspicuous. It is placed vertically, and arises from the ventral wall; com- mencing at the canalis auricularis and proceeding backwards, it does not as yet reach the opening into the sinus venosus. The blood from the sinus, or, as we may call it, the inferior vena cava, enters the heart obliquely from the right, so that it has a tendency to flow towards the left auricle of the heart, which is at this time the larger of the two. The valves between the ventricles and auricles are now well developed, and it is about this time that the division of the bulbus arteriosus into the aorta and pulmonary artery becomes visible on the exterior. By the eleventh or thirteenth day the right auricle has become as large as the left, and the auricular septum much more complete, though there is still a small opening, the foramen ovale, by which the two cavities communicate . with each other. Through this foramen the greater part of the blood of the vena cava inferior, which is now joined just vu. ] THE EUSTACHIAN VALVE. 195 at its entrance into the heart by the right vena cava superior, is directed into the left auricle. The left vena cava superior enters the right auricle independently ; between it and the inferior vena cava is a small valve which directs its blood entirely into the right auricle. On the sixteenth day the right vena cava superior, when viewed from the exterior, still appears to join the inferior vena cava before entering the heart; from the interior how- ever the two can now be seen to be separated by a valve. This valve, called the ‘Eustachian valve,’ extends to the opening of the left vena cava superior, and into it the valve which in the earlier stage separated the left superior and inferior venze cavee has apparently become merged. There is also on the left side of the opening of the inferior cava a membrane, stretching over the foramen ovale, and serving as a valve for that orifice. The blood from the inferior cava still passes chiefly into the left auricle through the foramen ovale; while the blood from the other two venz cave now falls into the right auricle, being prevented from entering the left chamber by the Eustachian valve. Hence, since at this period also the blood from the left ventricle passes to a great extent to the anterior portion of the body, there is a species of double-circulation going on. The greater part of the blood from the allantois entering the left auricle from the inferior vena cava passes into the left ventricle and is thence sent chiefly to the head and anterior extremities; from these it is brought back through the right auricle to the right ventricle, from whence it is returned along the aorta to the allantois. From the seventeenth to the nineteenth day, the right auricle becomes larger than the left. The large Eustachian valve still prevents the blood from the superior cave from entering the left auricle, while it conducts the blood from the inferior vena cava into that chamber through the foramen ovale. The entrance of the inferior vena cava is however further removed than it was from the foramen ovale, and the increased flow of blood from the lungs prevents all the blood of the inferior cava from entering into the left auricle. At the same time the valve of the foramen ovale prevents the blood in the left auricle from entering the right auricle. During the period from the seventh day onwards, the 13 —2 196 THE FIFTH DAY. [CHAP. apex of the heart becomes more marked; the arterial roots are more entirely separated and the various septa completed, so that when the foramen ovale is closed and the blood of the inferior vena cava thereby entirely confined to the right auricle, the heart has practically acquired its adult con- dition. 10, The fifth day may also be taken as marking the epoch at which histological differentiation first becomes distinctly established. It is of course true that long before this date, even from the earliest hours, the cells in each of the three fundamental layers have ceased to be everywhere alike. Nevertheless the changesundergone by the several cells have been few and slight. The cells of epiblastic origin, both those going to form the epidermis and those included in the neural involution, are up to this time simple more or less columnar cells; they may be seen here elongated, there oval, and in another spot spheroidal ; here closely packed with scanty protoplasm, there scattered with each nucleus well surrounded by cell-substance ; but wherever they are found they may still be recognized as cells of a distinctly epithelial character. So also with the cells of hypoblastic origin, whether simply lining the alimentary canal or taking part in the formation of the compound glands. Even in the mesoblast, which undergoes far more changes than either of the other layers, not only increasing more rapidly in bulk but also serving as the mother tissue for a far greater number of organs, the alterations im the individual cells are, till near upon the fifth day, insignificant. Up to this time, the mesoblast may be spoken of as consisting of little more than indifferent tissue :—of nuclei imbedded in a, protoplasmic cell-substance. In one spot the nuclei are closely. packed together, and the cell-substance scanty and compact; at another the nuclei are scattered about with spindle-shaped. masses of protoplasm attached to each, and there is a large development either of intercellular spaces or of intracellular vacuoles, filled with clear fluid. The proto- plasm differs in various places, chiefly in being more or less granular, and less or more transparent, having as yet under- gone but slight chemical transformation. Up to this epoch (with the exception of the early differentiated blood), there are no distinct ¢éssues, and the rudiments of the various VIL] THE EPIBLAST, 197 organs are simply marked out by greater or less condensation of the simple mesoblastic substance. From the fifth day onwards, however, histological differ- entiation takes place rapidly ; and it soon becomes possible to speak of this or that part as being composed of muscular, or cartilaginous, or connective &c. tissue. It is not within the scope of the present work to treat in detail of these histo- genetic changes, for information concerning which we would refer the reader to histological treatises. We have already had occasion to refer incidentally to many of the earliest histological events; and shall content ourselves by giving a brief summary of the derivation of the tissues of the adult animal from the three primary layers of the blastoderm. The epiblast or upper layer of many embryologists forms primarily two very important parts of the body, viz. the central nervous system and the epidermis. It is from the involuted epiblast of the neural tube, that the whole of the grey matter of the brain and spinal cord appears to be developed, the simple columnar cells of the epiblast being apparently directly transformed into the characteristic caudate nerve-cells. There is, however, some doubt whether mesoblast cells may not possibly enter into its formation, and it is very probable that the white matter of the brain and spinal cord is derived from the mesoblast alone. The epithelium (ciliated in the young animal) lining the canalis centralis of the spinal cord, together with that lining the ventricles of the brain, all which cavities and canals are, as we have seen, derivatives of the primary neural canal, is the undifferentiated remnant of the primitive epiblast. The epiblast as we have said also forms the epidermis; not however the dermis, which is of mesoblastic origin. The line of junction between the epiblast and the meso- blast coincides with that between the epidermis and the dermis. From the epiblast are formed all such tegumentary organs or parts of organs as are epidermic in nature. In addition to these, the epiblast plays an important part in the formation of the organs of special sense. According to their mode of formation, these organs may be arranged into two divisions. In the first come the eases where the sensory expansion of the organ of special 198 THE FIFTH DAY. [CHAP. sense is derived from the involuted epiblast of the medullary canal. To this class belongs the Retina, including the epi- thelial pigment of the choroid, whichis formed from the engimal optic vesicle budded out from the fore- brain. To the second class belong the epithelial expansions a: the membranous labyrinth of the ear, and the cavity of the nose, which are formed by involution from the superficial epiblast covering the external surface of the embryo. These accordingly have no primary connection with the brain. We may also fairly suppose that the ‘taste bulbs’ and the nervous cells which have lately been described as present in the epidermis are also structures formed from the epiblast. In addition to these we have the crystalline lens formed of involuted epiblast and the cavity of the mouth lined by it. These are the most important parts which are derived from the epiblast. From the hypoblast are derived the epithelium of the digestive canal, the epithelium of the trachea, bronchial tubes and air cells, the cylindrical epithelium of the ducts of the liver, pancreas and other glands of the alimentary canal, as well as the hepatic cells constituting the parenchyma of the liver, developed as we have seen from the solid hypoblast cylinders given off around the primary hepatic diverticula. Homologous probably with the hepatic cells, and equally of hypoblastic origin, are the spheroidal ‘secreting cells’ of pancreas and other glands. The epithelium of the salivary glands, though these so exactly resemble the pancreas, is of epiblastic origin, inasmuch as the cavity of the mouth (Chap. vi. § 8) is entirely lined by epiblast. The hypoblast also lines the allantois. From the mesoblast are formed all the remaining parts of the body. The muscles, the bones, the connective tissue and the vessels, both arteries, veins, capillaries and lymphatics with their appropriate epithelium, are entirely formed from the mesoblast. All the nerves of the body, both the cranial nerves (the so-called optic and olfactory nerves alone excepted), the spinal nerves,’and the sympathetic system, are also formed from the mesoblast. The nerve-cells of the sympathetic ganglia as well as those of the ganglia on the posterior roots of the spinal nerves are of mesoblastic origin, and thus appaz- VIL] THE MESOBLAST. 199 ently are in striking contrast with the nerve-cells in the brain and cord. The fibres constituting the white matter of both brain and spinal cord are also probably derived from mesoblast. The generative and urinary organs are entirely derived from the mesoblast. It is worthy of notice that their epithe- lium, though resembling so closely the hypoblastic epithelium of the alimentary canal, is distinctly mesoblastic. From the mesoblast lastly are derived all the muscular, connective, and nervous and vascular elements, as well of the alimentary canal and its appendages as of the skin and the tegumentary organs. Just as it is only the epidermic moiety of the latter which is derived from the epiblast, so it is only the epithelium of the former which comes from the hypoblast. In the present state of our knowledge we cannot in all cases with certainty say which parts of “the mesoblast. enter into the formation of particular organs ; the more important facts in this part of our subject will however already have been gathered, from the earlier part of this work. 11. The important events then which characterize the fifth day are :— 1. The growth of the allantois. 2. The appearance of the knee and elbow, and of the cartilages which precede the bones of the digits and limbs. 3. The formation of the primitive cartilaginous cranium, more especially of the investing mass, the trabeculee, and the ethmo-vomerine plate ; and the appearance of rods of cartilage in the visceral arches. 4. The developments of the parts of the face: the closing in of the nasal passages by the nasal processes. 5. The formation of the anus. 6.