The development of the human brain and the closure of the rostral neuropore at stage 11

Summary Twenty embryos of stage 11 (24 days) were studied in detail and graphic reconstructions of twelve of them were prepared. The characteristic feature of this stage is 13–20 pairs of somites.The notochord sensu stricto appears first during this stage, and its rostral and caudal parts differ in origin. Rostrally, the notochordal plate is being transformed into the notochord in a caudorostral direction. The caudal part, however, arises from the axial condensation in the caudal eminence in a rostrocaudal direction. The caudal eminence (or end bud) represents the former primitive streak. The somites are increasing in number at a mean rate of 6.6 h per pair.The rostral neuropore closes towards the end of stage 11. The closure is basically bidirectional, being more rapid in the roof region and producing the embryonic lamina terminalis and future commissural plate in the basal region. The caudal neuropore is constantly open. The brain comprises telencephalon medium (represented by the embryonic lamina terminalis) and a series of neuromeres: 2 for the forebrain (D1 and D2), 1 for the midbrain, and 6–7 for the hindbrain (RhA-C; Rh D is not clearly delineated). The forebrain still occupies a small proportion of the total brain, whereas the spinal part of the neural tube is lengthening rapidly. Some occlusion of the lumen of the neural tube was noted in 4 embryos, all of which had an open rostral neuropore. Hence there is at present no evidence that occlusion plays a role in expansion of the human brain. The marginal (primordial plexiform) layer is appearing, particularly in rhombomere D and in the spinal portion of the neural tube. The neural crest is still forming from both the (open) neural groove and the (closed) neural tube, and exclusively from both neural (including optic) and (mainly) otic ectoderm.The optic sulcus is now prominent, and its wall becomes transformed into the optic vesicle towards the end of stage 11. At this time also, an optic sheath derived from mesencephalic crest and optic crest is present. The mitotic figures of the optic neural crest are exceptional in being situated in the external part of the neural epithelium. The otic pit is becoming deeper, and its wall is giving rise to neural crest that is partly added to the faciovestibulocochlear ganglion and partly forms an otic sheath. The nasal plate does not yet give off neural crest.Abbreviations: Figs. 1–10. a Endoderm caudal to neurenteric canal or its site B Endoderm rostral to neurenteric canal or its site - A-H Primordium of adenohypophysis - All Allantoic primordium - Ao Aorta - C.E. Caudal eminence (caudal bud, end bud) - Caud.lim.S. Caudal limiting sulcus - Ch. Chiasmatic plate - D Diencephalon - F Foregut, pharynx - Cl. Cloacal membrane - Ggl Ganglion - H Hindgut - I Infundibulum - L.T. embryonic lamina terminalis - M Mamillary area - Mes. Mesencephalon, mesencephalic - Mit. Mitotic figure - Nas. Nasal plate - N.C. Site of neurenteric canal - N.Cr. Neural crest - Not.Pl. Notochordal plate - Not. Notochord - O-Ph Oropharyngeal membrane - Opt. Optic primordium - Opt.S. Optic sulcus - Ot. Otic pit - Ot.sh. Otic sheath - Ph.Ar. Pharyngeal arch - Pr. Mesenchyme of prechordal plate - Pros. Prosencephalon - Rec. Postoptic recess - Resp. Respiratory primordium - Rh Rhombomere - S.V. Sinus venosus - s. Somite - Tel. Telencephalon - Th. Chickening - Thyr. Thyroid primordium - Trig. Trigeminal - X Cau - al neuropore - Y Rostral neuropore Supported by research grant No. HD-16702, Institute of Child Health and Human Development, National Institutes of Health (USA)     

The development of the human brain, the closure of the caudal neuropore, and the beginning of secondary neurulation at stage 12

Summary Twenty-four embryos of stage 12 (26 days) were studied in detail and graphic reconstructions of five of them were prepared. The characteristic features of this stage are 21–29 pairs of somites, incipient or complete closure of the caudal neuropore, and the appearance of upper limb buds. The caudal neuropore closes during stage 12, generally when 25 somititc pairs are present. The site of final closure is at the level of future somite 31, which corresponds to the second sacral vertebral level. Non-closure of the neuropore may be important in the genesis of spina bifida aperta at low levels. The primitive streak probably persists until the caudal neuropore closes, when it is replaced by the caudal eminence or end-bud (Endwulst oder Rumpfknospe). The caudal eminence, which appears at stage 9, gives rise inter alia to hindgut, notochord, caudal somites, and the neural cord. The material for somites 30–34 (which appear in stage 13) is laid down during stage 12, and its absence would be expected to result in sacral agenesis. Aplasia of the caudal eminence results in cloacal deficiency and various degrees of symmelia.The junction of primary and secondary development (primäre und sekundäre Körperentwicklung) is probably at the site of final closure of the caudal neuropore. Secondary neurulation begins during stage 12. The cavity of the already formed spinal cord extends into the neural cord, and isolated spaces are not found within the neural cord. Primary and secondary neurulation are probably coextensive with primary and secondary development of the body, respectively. The telencephalon medium has enlarged two mesencephalic segments (M1 and M2) are distinguishable, and rhombomere 4 is reduced. The sulcus limitans is detectable in the spinal cord and hindbrain (RhD), and in the mesencephalon and diencephalon, where it extends as far rostrally as the optic sulcus in D1. A marginal layer is appearing in the rhombencephalon and mesencephalon. The first nerve fibres are differentiating, chiefly within the hindbrain (from the nucleus of the lateral longitudinal tract). Optic neural crest is at its maximum, and the otic vesicle is giving crest cells to ganglion 7/8. Neural crest continues to develop in the brain and contributes to cranial ganglia 5, 7/8, and 10/11. The spinal crest extends as far caudally as somites 18–19 but shows no subdivision into ganglia yet. Placodal contribution to the trigeminal ganglion is not certain at stage 12. Such a contribution to ganglion 7/8 is not unlikely. Involvement of neural crest in the formation of the derivatives of pharyngeal arches 1 and 2 is possible but has not yet been confirmed in the human embryo.Supported by research grant No. HD-16702, Institute of Child Health and Human Development, National Institutes of Health (USA)     

Deceleration and acceleration in the rate of posterior neuropore closure during neurulation in the curly tail (ct) mouse embryo

Summary Curly tail (ct) is a mouse mutant producing spinal neural tube defects as a result of delayed closure of the posterior neuropore (PNP). The purpose of the present study was to determine in ct/ct embryos the time of onset of the delay in PNP closure, and the pattern of this closure, as well as to study the possibility that reopening of the neural tube occurs. Normal spinal neurulation was studied in non-mutant Swiss (Sw) embryos. In the latter, the average PNP length diminished steadily between the 7- and 25-somite stages, and then decreased more rapidly, indicating an acceleration of closure rate, until the 30- to 32-somite stage, when all PNPs closed. PNP width decreased steadily between the stages of 7 and 30 somites. In ct/ct embryos the average PNP length showed a slight increase between the stage of 23 to 28 somites, indicating a temporary deceleration of closure rate, and the range of PNP sizes increased markedly. This was followed by a decrease in PNP length until the 37-somite stage, indicating an acceleration of closure rate. From the stage of 32 somites onwards, the proportion of embryos with closed PNPs gradually increased to 90%. The population of ct/ct embryos was subdivided. Embryos with large PNPs showed a marked deceleration of closure rate during a period of 11 somite stages, followed by a brief but very high acceleration of closure rate. This resulted in closure of the PNP in a proportion of these embryos, while in the remainder of the embryos the deceleration phase had been too enhanced to allow complete catch up of closure during the acceleration phase; these embryos would develop spina bifida. Embryos with relative small PNPs also showed a deceleration of closure rate, but only during a period of four somite stages. This was followed by an acceleration, resulting in closure of all PNPs at the stage of 32 to 33 somites. The enlargement of the PNP in ct/ct embryos was not due to re-opening of a closed neural tube, but resulted from a sharp decline in the rate of PNP closure combined with a normal rate of caudal elongation of the embryo. It is concluded that the ct strain forms a homogeneous population, with a large variation of its specific phenotype: deceleration of PNP closure during a restricted period. The disturbance of spinal neurulation in ct/ct embryos takes the form of a deceleration/acceleration pattern, resulting in a net delay of closure. It is suggested that, due to the ct mutation, forces are generated in the embryonic axis which oppose a normal neurulation process at a specific stage of development.     

Odor enrichment influences neurogenesis in the rostral migratory stream of young rats

The olfactory bulb is one of a few brain structures characterized by high plasticity due to the fact that new neurons are continually integrated into the olfactory bulb circuit throughout life. The new cells originate from the subventricular zone of the forebrain and migrate through the rostral migratory stream (RMS) to the olfactory bulb that also represents the first synaptic relay of the olfactory system. Data accumulating in recent years have confirmed that sensory inputs can influence the level of postnatal neurogenesis in the olfactory bulb. In this study, we studied neurogenesis in the rostral migratory stream of Wistar albino rat pups after exposure to an odor-enriched environment. The rats were olfactory stimulated twice daily with different odorants from the day of their birth up to 1, 2 or 3 weeks, respectively. Using bromodeoxyuridine, a marker of cell proliferation, we found an increased number of proliferating cells in the rostral migratory stream of rat pups submitted to olfactory stimulation. Conversely, the number of dying cells, labeled with the fluorescent dye Fluoro Jade-C, was down-regulated in groups of rats exposed to an odor-enriched environment.     

喙端迁移流的发生及其细胞凋亡

目的 研究生后不同日龄小鼠喙端迁移流（RMS）的发育,神经干细胞增殖和凋亡的规律。方法 利用Caspase-8免疫荧光标记法和5’-溴脱氧尿嘧啶核苷（BrdU）法,对小鼠RMS内的神经干细胞增殖和凋亡进行研究（n =92）。结果 生后早期小鼠脑内,尤其是室管下区（SVZ）和RMS,存在大量的增殖细胞。随着小鼠年龄的增加,脑内干细胞逐渐减少,到成年,大脑皮质几乎见不到增殖的神经干细胞,但在SVZ和RMS仍可以看到许多增殖的神经干细胞。在RMS,神经干细胞增殖的同时伴随着细胞凋亡,干细胞的增殖与凋亡存在着正相关关系。结论 RMS的神经干细胞增殖与凋亡有重要的生理意义,通过细胞凋亡,RMS可以调节神经干细胞向嗅球迁移的数量,也可以调节干细胞向颗粒细胞分化。     

A segmented pattern of cell death during development of the chick embryo

Summary During the early development of the chick embryo, specific groups of cells die in characteristic patterns. In this study, Nile Blue sulphate staining was used to reveal a novel pattern of segmentally repeated cell death in the paraxial mesoderm of the chick prior to stage 23. This pattern varies according to the developmental stage of the embryo and shifts rostrocaudally, corresponding to progressing somite differentiation. Initially, during early somite differentiation, cell death is restricted to the rostral half of the somite (the rostral pattern of cell death). After the somite has differentiated into dermomyotome and sclerotome, dead cells appear in superficial tissues in a pyramidal pattern which lies in register (rostrocaudally) with the central part of the sclerotome. Finally, small bands of dying cells are seen between the neural tube and the expanding sclerotome. This third pattern (the ventral path) lies in register with the rostral part of the caudal half of the sclerotome. We show by fluorescent labelling of the migrating neural crest that these patterns of cell death correspond to the routes of neural crest migration. In addition, serial sectioning of stage 23 chick embryos confirms that the position of dying cells correlates with the known routes of neural crest migration and with the sites of development of certain neural crest-derived tissues.     

Wnts and the neural crest

The neural crest is a multipotent tissue that originates between the neural epithelium and non-neural ectoderm, which can develop into numerous cell types, including neurons, glia, pigment cells, smooth muscle, cartilage and bone. Work in a variety of animal models has shown that a number of signalling factors are necessary for the induction, delamination and differentiation of neural crest cells. However one family of proteins, the Wnts, shows an overriding influence on this tissue. Here we review recent studies that pinpoint specific roles that Wnts play in the development of the neural crest.     

Multiphoton microscope imaging: The behavior of neural progenitor cells in the rostral migratory stream

Neural progenitor cells (NPCs) in the subventricular zone (SVZ) travel a long distance along the rostral migratory stream (RMS) to give rise to interneurons in the olfactory bulb (OB). Using the multiphoton microscope and time-lapse recording techniques we here report the behavior of NPCs in the RMS under both intact and ischemic conditions in living brain slices. The NPCs were visualized in 3-week-old transgenic mice that carry the reporter gene, green fluorescent protein (GFP), driven by the nestin promoter. Cortical brain ischemia was induced by permanent occlusion of the right common carotid artery and the middle cerebral artery. We observed that the RMS contained two populations of NPCs: nonmigrating cells (bridge cells) and migrating cells. Bridge cells enabled migrating cells to travel and also produced new cells in the RMS. The direction of NPC migration in the RMS was bidirectional in both intact and ischemic conditions. Cortical ischemia impeded NPC travel in the RMS next to the lesion area during the early period of ischemia. Cell–cell contact was a prominent feature affecting NPC translocation and migratory direction. These data suggest that behavior and function of nestin-positive NPCs in the RMS are variable. Cell–cell contacts and microenvironmental changes influence NPC behavior in the RMS. This study may provide insights to help in understanding NPC biology.     

Analysis of hindbrain neural crest migration in the long-tailed monkey (Macaca fascicularis)

Neural crest cells make a substantial contribution to normal craniofacial development. Despite advances made in identifying migrating neural crest cells in avian embryos and, more recently, rodent embryos, knowledge of crest cell migration in primates has been limited to what was obtained by conventional morphological techniques. In order to determine the degree to which the nonhuman primate fits the mammalian pattern, we studied the features of putative neural crest cell migration in the hindbrain of the long-tailed monkey (Macaca fascicularis) embryo. Cranial crest cells were identified on the basis of reported distributional and morphological criteria as well as by immunocytochemical detection of the neural cell adhesion molecule (N-CAM) that labels a subpopulation of these cells. The persistent labeling of a sufficient number of crest cells with antibodies to N-CAM following their exit from the rostral, preotic and post-otic regions of the hindbrain facilitated tracking them along subectodermal pathways to their respective destinations in the first, second and third pharyngeal arches. Peroxidase immunocytochemistry was also employed to localize laminin and collagen-IV in neuroepithelial basement membranes. At stage 10 (8–11 somites), crest emigration occurred in areas of unfused neural folds through focal disruptions in the neuroepithelial basement membrane in both the rostral and pre-otic regions, although there was little evidence of crest migration in the post-otic hindbrain. By stage 11 (16–17 somites), the neural folds were fused (pre- and post-otic hindbrain) or in the process of fusing (rostral hindbrain), yet crest cell emigration was apparent in all three areas through discontinuities in the basement membrane. Emigration was essentially complete at stage 12 (21 somites) as indicated by nearly continuous cranial neural tube basement membranes. At this stage the pre-ganglia (trigeminal, facioacoustic and glossopharyngeal) were consistently stained with N-CAM. The current study has provided new information on mammalian neural crest in a well-established experimental model for normal and abnormal human development, including its use as a model for the retinoic acid syndrome. In this regard, the current results provide the basis for probing the mechanisms of retinoid embryopathy which may involve perturbation of hindbrain neural crest development.     

Loss and restoration of preoptic thermoreactiveness after lesions of the rostral raphe nuclei

Summary Preoptic neurons, extracellularly recorded in the rat's brain, were tested for their responses to thermal stimulation of the scrotal and abdominal skin before and after electrolytic lesions of about 1 mm³ in the area of the rostral raphe nuclei, nucleus raphe dorsalis and centralis (NRD/NRC). All analyzed neurons were of the switching type, i.e. they changed their firing rates to a higher or lower level when a threshold of the peripheral stimulation temperature was exceeded. When major parts of NRD or NRC were destroyed, the preoptic neurons no longer changed their firing rates after thermal stimulation, whereas transmission of noxious information in most cases was not impeded. Smaller lesions in NRD or NRC did not abolish the responses, but brought about essentially modified responses compared to those before the lesions. Lesions lateral to NRD or NRC had no effect. If the lesions were effective and the neurons could be observed for a longer period after the lesions, the response was restored in many cases. As the noxious response had often not been abolished and the lateral lesions were without any effect, it might be that the lesion effects and the restoration of responses involve short-term plasticity. However, temporary block of input to the neurons by unspecific effects cannot be excluded.     

On the differentiation and migration of some non-neuronal neural crest derived cell types

Summary Neural tubes containing premigratory neural crest cells from head and trunk levels as well as somites containing neural crest cells that have migrated away from the neural crest were grafted orthotopically and heterotopically from quail embryos to chicken embryos. Schwann cells and melanocytes of donor origin developed after all grafting procedures. Cartilage developed only from neural crest cells of head levels. No skeletal muscle was ever observed to develop from the neural crest. The development of these different cell types from heterotopically grafted premigratory neural crest cells indicates that the neural crest is not a population of pluripotent undeterminated cells, but that at least some determinated cells are present within it before the onset of emigration of neural crest cells from the neural crest. Different neural-crest-derived cell populations exhibit different migratory behaviour: After heterotopically grafting quail neural crest cells to the wing buds of chicken embryos. Schwann cells and non-epidermal melanocytes were found to have migrated proximally and distally away from the grafts. Epidermal melanocytes of donor origin were found to have migrated in a distal direction essentially.This work was supported by the Österreichischer Fonds zur Förderung der wissenschaftlichen Forschung (P 4680)     

The ultrastructure of early cephalic neural crest cell migration in the mouse

Summary A study of the ultrastructural changes associated with the detachment of the presumptive neural crest cells from the neuroepithelium in the midbrain region in mouse embryos at 9 and 91/2 days of gestation was carried out. The first sign of neural crest cell formation occurred in this region before fusion of the neuroepithelium had occurred. Neural crest cells arose from both the neural plate and the adjoining surface ectoderm. Initially, the cells of the neural plate and the surface ectoderm were attached to each other by zonula occludens and zonula adherans at their apical surfaces however, these junctions disappeared just prior to the beginning of the migration of the crest cells. The first sign of migration of the crest cells was the disappearance of the basal lamina in the region of the presumptive crest cells. Once the basal lamina was lost, cell junctions were formed between the epithelial cells and the underlying mesenchymal cells. Once the crest cells had migrated into the underlying mesenchyme, they tended to form clumps of closely related, irregularly shaped cells. Phagosomes and accumulations of glycogen particles were found within some crest cells when they were still within 50 to 100 microns of the epithelium.     

Role of the extracellular matrix in neural crest cell migration

Development of the neural crest involves a remarkable feat of coordinated cell migration in which cells detach from the neural tube, take varying routes of migration through the embryonic tissues and then differentiate at the end of their journey to participate in the formation of a number of organ systems. In general, neural crest cells appear to migrate without the guidance of long-range physical or chemical cues, but rather they respond to heterogeneity in the extracellular matrix that forms their migration substrate. Molecules such as fibronectin and laminin act as permissive substrate components, encouraging neural crest cell attachment and spreading, whereas chondroitin sulphate proteoglycans are nonpermissive for migration. A balance between permissive and nonpermissive substrate components seems to be necessary to ensure successful migration, as indicated by a number of studies in mouse mutant systems where nonpermissive molecules are over-expressed, leading to inhibition of neural crest migration. The neural crest expresses cell surface receptors that permit interaction with the extracellular matrix and may also modify the matrix by secretion of proteases. Thus the principles that govern the complex migration of neural crest cells are beginning to emerge.     

The first appearance of the neural tube and optic primordium in the human embryo at stage 10

Summary Thirteen embryos of stage 10 (22 days) were studied in detail and graphic reconstructions of most of them were prepared. The characteristic feature of this stage is 4–12 pairs of somites. Constantly present are the prechordal and notochordal plates (the notochord sensu stricto is not yet apparent), the neurenteric canal or at least its site, the thyroid primordium, probably the mesencephalic and rhombencephalic neural crest and the adenohypophysial primordium. During this stage, the following features appear: terminal notch, optic sulcus, initial formation of neural tube, oropharyngeal membrane, pulmonary primordium, cardiac loop, aortic arches 1–3, intersegmental arteries, and laryngotracheal groove. The primitive streak is still an important feature.Graphic reconstructions have permitted the detection of the telencephalic portion of the forebrain, for the first time at such an early stage. It is proposed that the remainder of the forebrain comprises two subdivisions: D1, which becomes largely the optic primordium during stage 10, and D2, which is the future thalamic region. The optic sulcus is found in D1 but does not extent into D2, as has been claimed in the literature. An indication of invagionation of the otic disc appears towards the end of the stage. As compared with the previous stage, the prosencephalon has increased in length, the mesencephalon has remained the same, the rhombencephalon has decreased, and the spinal part of the neural plate has increased fivefold in length. The site of the initial closure of the neural groove is rhombencephalic, upper cervical, or both. The neural plate extends caudally beyond the site of the neurenteric canal. Cytoplasmic inclusions believed to indicate locations of great activity were always detected in the forebrain (especially in the optic primordium), and also in the rhombencephalon, spmal part, and mesencephalon.Supported by research grant No. HD-16702, Institute of Child Health and Human Development, National Institutes of Health (USA)     

Morphological and quantitative studies in the otic region of the neural tube in chick embryos suggest a neuroectodermal origin for the otic placode

Careful histological observation of the development of the anlage of the inner ear in chicken embryos led us to question the traditional view of otic placode (OP) formation. First, morphological studies in the cephalic region carried out on stages preceding the appearance of the placodal epithelium revealed that the medial placodal cells are continuous temporally and spatially with cells belonging to the neural fold (NF). Second, both the formation of the basal lamina between the dorsal region of the neural tube (NT) and ectoderm and the pattern of formation of the neural crest present distinctive characteristics between otic levels and regions located anteriorly and posteriorly. Third, numerical comparisons of parameters for the NT and the OP between different levels of the rhombencephalon allowed us to assign a differential behaviour in the growth pattern of the otic region. These results indicated that the medial part of the OP is not derived from already independent ectoderm that increases in thickness under the influence of the NT (as previously accepted) but that it develops directly from the NFs. Although we do not exclude other possibilities, we propose that at least a proportion of the OP cells originate directly from cells committed to be neural crest. After this incorporation, basal laminal formation would delimit the NT from the OP without transition of the otic cells to ectoderm. This hypothesis would imply that part of the otic cells originate directly from neuroepithelial cells having a neuroectodermal (rather than the previously established ectodermal) origin.     

成纤维细胞生长因子信号调控早期鸡胚胎神经嵴细胞的迁移

目的 利用Sprouty2基因阻断成纤维细胞生长因子（FGF）信号,探讨FGF在早期鸡胚胎发育过程中对神经嵴细胞迁移的影响及其机制。方法 通过体内培养的方法孵育鸡胚至HH9期,通过显微注射的方法将Sprouty2-绿色荧光蛋白（GFP）质粒注射入神经管腔内。实验侧使用电穿孔转染的方法转染胚胎半侧神经管,另一侧正常神经管设为对照侧。采用神经嵴细胞特异标记物HNK1免疫荧光的方法检测Sprouty2基因阻断FGF信号后是否影响胚胎头部和躯干部神经嵴细胞的迁移过程。随后,进一步通过检测神经细胞钙黏分子N-Cadherin的表达来观察细胞之间黏附作用的改变。结果 HNK1免疫荧光检测结果显示,Sprouty2转染侧即阻断FGF信号通路后,HNK1在早期鸡胚胎的头部和躯干部的表达量均比对照侧的表达量增多;而神经细胞钙黏分子N-Cadherin检测结果表明,Sprouty2转染侧和正常对照侧N Cadherin在头部和躯干部神经管上表达量的差异均无显著性。结论 Sprouty2基因阻断FGF信号后,促进了早期鸡胚胎神经嵴细胞的迁移,但是FGF信号对此过程的影响可能不是由神经钙黏分子N-Cadherin介导的。     

Interdigital chondrogenesis and extra digit formation in the duck leg bud subjected to local ectoderm removal

Summary In the chick embryo the interdigital tissue in the stages previous to cell death exhibits in vitro a high chondrogenic potential, and forms extra digits when subjected in vivo to local ectodermal removal. In the present work we have analyzed the chondrogenic potential both in vivo and in vitro of the interdigital mesenchyme of the duck leg bud. As distinct from the chick, the interdigital mesenchyme of the duck leg bud exhibits a low degree of degeneration, resulting in the formation of webbed digits. Our results show that duck interdigital mesenchyme exhibits also a high chondrogenic potential in vitro until the stages in which cell death starts. Once cell death is finished chondrogenesis becomes negative and the interdigital mesenchyme forms a fibroblastic tissue. In vivo the interdigital mesenchyme of the duck leg bud subjected to ectoderm removal forms ectopic foci of chondrogenesis with a range of incidence similar to that in the chick. Unlike those of the chick the ectopic cartilages of the duck are rounded and smaller, and appear to be located at the distal margin of the interdigital mesenchyme. Formation of extra digits in the duck occurs with a lower incidence than in the chick. It is concluded that ectopic chondrogenesis and formation of extra digits is related to the intensity of interdigital cell death. The non-degenerating interdigital mesenchymal cells destined to form the interdigital webs of the duck appear to contribute very little to the formation of interdigital cartilages.     

Association of cephalic neural crest cells with cardiovascular development, particularly that of the semilunar valves

Summary The quail-chick chimera method was used to examine whether neural crest cells were associated with the formation of semilunar valves. From the metencephalon to somite 5, or from the otocyst to somite 3, left, right, or bilateral neural folds, including the neural crest, were transplanted. Among embryos used for the experiment, three into which left neural crest cells were transplanted, two into which right neural crest cells were transplanted, and two into which bilateral neural crest cells were transplanted had a morphologically normal heart. In these embryos, neural crest cells were found in all cusps of the aortic and pulmonary semilunar valves.Although neural crest cells have been thought to have no association with the formation of the semilunar valves, our experiment indicates that such association indeed occurs.     

Distribution of the neural crest cells in the heart of birds: a three dimensional analysis

Summary The distribution of neural crest derived cells (NC) in the heart of quail-chick chimeric embryos was analyzed three-dimensionally after computer reconstruction. During the division of the truncus arteriosus into the aorta and the pulmonary trunk, ventral and dorsal columns of NC-derived cells were found in the truncal swellings. These columns were elongations from the aorticopulmonary (AP) septum. The dorsal column extended more proximally than did the ventral column. Around hatching, NC-derived cells located between the proximal aorta and the pulmonary trunk, differentiated into cartilage and connective tissue. They formed a part of the cardiac skeleton. A small number of NC-derived cells were scattered in the cusps of the arterial valves. Cells derived from the right NC were located around the aorta and the right arch arteries but not around the distal pulmonary trunk and the left arch arteries. At the proximal level, cells derived from the rigth NC were located in both the dorsal and ventral columns. These results suggest that the AP septum is mainly formed by NC-derived cells, right and left NC cells migrating into assigned areas in the heart. Location of two columns of NC-derived cells may support a translocation hypothesis for the AP septum during truncal division.     

Development of cranial nerves in the chick embryo with special reference to the alterations of cardiac branches after ablation of the cardiac neural crest

Summary Development of cranial nerve branches in the cardiac region was observed in whole-mount specimens which were stained with a monoclonal antibody, E/C8, after the ablation of the cardiac neural crest. In early embryos, nerve trunks of IX and X were lacking or only poorly developed, while the early development of pharyngeal branch primordia was normal. In day 5 embryos, the nerve trunks of IX–X were present in all the embryos, however; extensive communication was observed between X and XII. On day 6 and later, the spiral pattern of superior cardiac branches was disturbed, as were the blood vessels. Furthermore, the distal branches of XII passed within the superficial layer of cardiac outflow mesenchyme. Vagal branches passed within the deeper layer. There was no apparent change in the development of the sinal branch. Using quail — chick chimeras, it was found that the cardiac neural crest cells formed the Schwann cells of XII, and that they were also associated with the hypobranchial muscle primordium, suggesting that the absence of the cardiac neural crest not only disturbs the development of the cardiac outflow septation, but also affects the normal morphogenesis of the hypobranchial musculature and its innervation. Embryologically, the tongue is located close to the cardiac outflow tract, which is the migration pathway of the cardiac neural crest-derived cells.