Changes in dorsoventral but not rostrocaudal regionalization of the chick neural tube in the absence of cranial notochord, as revealed by expression of engrailed-2.

Notochord has been implicated in previous studies in both the dorsoventral and rostrocaudal patterning of the developing neural tube. This possibility has been further explored by analyzing the expression of Engrailed-2 in chick embryos developing with cranial notochord defects. Control embryos containing intact notochords expressed Engrailed-2 protein within the neural tube and in a subset of the neural crest and overlying surface ectoderm at the future mesencephalon and cranial metencephalon levels. Within the neural tube, expression was confined to cell nuclei in the roof plate and lateral walls; floor plate nuclei directly overlying the notochord typically failed to show expression. After surgical removal of Hensen's node, the source of notochord precursor cells, embryos were cultured through neurulation and assayed for expression of Engrailed-2 protein. All embryos that partially or completely lacked cranial notochord expressed Engrailed-2 in a pattern similar to that of control embryos containing intact notochords, except that when notochord and floor plate were absent, Engrailed-2 was also expressed in the most ventral part of the neural tube. These results indicate that 1) Engrailed-2 expression is suppressed in the most ventral neural tube owing to induction of the floor plate by the notochord, and 2) that the presence of an underlying notochord is not required for correct rostrocaudal expression, suggesting that multiple pathways act in the patterning of the rudiment of the central nervous system.     

Indirect immunofluorescent staining of fibronectin associated with the floor of the foregut during formation and rupture of the oral membrane in the chick embryo

The distribution of the glycoprotein, fibronectin, within the cranial region of stage 8–16 chick embryos was examined by indirect immunofluorescence using paraffin sections exposed to affinity-purified rabbit anti-human CIG and FITC-conjugated goat anti-rabbit immunoglobulins. Fluorescence was present within the matrix surrounding the cranial mesenchyme, along the basal surfaces of all epithelia, and surrounding the notochord at all stages. Fluorescence associated with the floor of the foregut was particularly intense. The fluorescent layers beneath the ectoderm and endoderm of the oral (oropharyngeal) membrane at stage 8 merged into a single, continuous, intensely fluorescent line as the extra-cellular space within the oral membrane narrowed during stages 9–12. This line of uniform fluorescence parallels the previously described histological reorganization of the extracellular compartment of the oral membrane, but the ultrastructural localization of this fluorescent material remains unknown. Fluorescence was also intense beneath the foregut endoderm in the presumptive cardiac region caudal to the oral membrane and was continuous with strands of fluorescent material extending into the matrix of the dorsal mesocardium and cardiac jelly of the developing tubular heart. These observations indicate that the extracellular matrix associated with the floor of the entire foregut contains fibronectin during stages encompassing the formation and rupture of the oral membrane. The presence of fibronectin within the oral membrane and dorsal mesocardium, as well as between Rathke's pouch and infundibulum and within the closing plates between ectodermal clefts and endodermal pouches, is consistent with the possibility that this glycoprotein may play a role in adhesion at these sites.     

低氧诱导因子-1在小鼠胚胎神经胚形成阶段的发育学表达

为研究低氧诱导因子-1(HIF-1)在小鼠胚胎神经胚形成阶段的时空表达规律,探讨HIF-1在胚胎神经系统发生和发育过程中的作用。本研究以地高辛标记的HIF-1αcRNA为探针,采用全胚胎原位杂交技术,观察HIF-1α在神经胚形成阶段的表达规律。结果显示在E8.5d,整个鼠胚神经管的头端和尾端均可观察到HIF-1αmRNA的表达,此时表达较弱。在E9.5d,随着神经管的逐渐关闭,HIF-1αmRNA在前脑泡、第一鳃弓、第二鳃弓、后脑泡处急剧增多,并在发育中的眼部有较强的表达;此外在中脑泡以及心脏原基有较弱的表达。到E10.5d,阳性信号除在E9.5d检测到的部位继续富集外,在端脑、间脑、发育中的肢芽以及尾部也出现较强的HIF-1αmRNA的表达,且在心脏原基的表达亦进一步增强。E11.5d时在连合板、喙区、第一鳃弓、第二鳃弓、后脑、延脑、发育中的肢芽以及尾部末端可检测到HIF-1αmRNA的明显表达。以上结果提示HIF-1可能参与了小鼠神经胚形成的发育过程。     

Closure of the neural tube in the cephalic region of the mouse embryo

In mouse embryos varying in age from 9 to 20 somites the first closure of the neural groove was found to occur in the cervical region. The fusion process gradually proceedéd in rhombencephalic direction until it reached a level just caudal to the otic pits. Shortly afterwards the prosencephalic walls fused together independent of the rhombencephalic closure. This prosencephalic fusion process proceeded caudally in the direction of the mesencephalon until it reached the rostral portion of the rhombencephalon. In this region the two independent fusion processes met each other. In addition the prosencephalic fusion proceeded in rostral direction toward the anterior neuropore, which was the last part of the brain vesicles to close. Hence, the closure of the brain vesicles is not a zipper-like process proceeding from the rhombencephalon to the anterior neuropore, but occurs at several places at the same time and proceeds in a rostral as well as in a caudal direction. At the cellular level considerable differences in the fusion process were found to exist between the various brain vesicles. In the rhombencephalon the first bridge between the two opposing walls was formed by surface ectoderm and neural crest cells. In the mesencephalon single squamous ectoderm and a few neuroepithelial cells established the first contact, whereas in the prosencephalon the apical ends of several neuroepithelial cells fused together to overbridge the gap between the opposing walls. The surface ectoderm cells subsequently covered the neuroepithelial bridge. In the region of the anterior neuropore the fusion was similar to that between the prosencephalic walls, the only difference being that in the anterior neuropore area many more darkly stained particles indicating cell degeneration, were present than in the prosencephalon. It is thus concluded that considerable differences exist in the fusion of the neural walls between the various brain vesicles.     

Cephalic neurulation in the mouse embryo analyzed by SEM and morphometry

A detailed account of mouse neurulation is given based mostly on SEM analysis over 20 hr of development. Many observations and measurements were made on staged living embryos and on embryos prepared for scanning and light microscopy to help deduce what mechanisms may contribute to neural tube formation. Each-lateral half of the early cephalic neural plate makes a convex bulge, opposite to the way it must fold to form a tube. Underlying mesenchyme and matrix are reported to have a role in forming these bulges. Processes that form the tube must overcome this opposed folding and the forces that produce it. Cranial flexure begins long before tube formation. The flexure commences at the rostral tip of the cephalic neural plate, then the apex of the flexure migrates caudally to the mesencephalic region. Early appearance of this flexure imposes a mechanical impediment to tube closure in forebrain and midbrain regions. Tube closure begins in the cervical region exactly where the neural plate is reflected dorsally by a bend in the embryo. This bend may mechanically assist closure in this region. Cells of the mouse neural plate are reported to contain organized microfilaments and microtubules, and the plate cells appear to change shape (reduce apical area and increase cell height) in the same manner as that suggested in embryos of some other species to contribute to neural tube formation. Measurements show that the lateral edges of the cephalic neural plate elongate craniocaudally more than the midline of the plate through each period. This elongation could contribute to the folding of the plate into a tube. The progress of cranial ventral flexure pauses while tube formation occurs, but edge elongation continues, presumably contributing to tube formation. There is considerable increase in volume of the neural plate during tube closure, and cell proliferation and enlargement of daughter cells seem sufficient to account for this growth. Mitotic spindles are positioned to place the majority of the daughter cells into the long axis of the neural plate, so ordered growth may be the main mechanism of elongation of the plate in the craniocaudal direction, which in turn may assist in tube formation. Mouse cephalic neural plates appear overlying already segmented cranial mesenchyme according to previous reports, and neuromeres develop precociously in the open plates, where their positions correlate exactly with the underlying segmented mesenchyme.     

Separation of neural and surface ectoderm after closure of the rostral neuropore

Summary Separation of neural and surface ectoderm after closure of the rostral neuropore in the head region has been described by investigating the integrity of the basement membranes of these epithelia in 11- to 27-somite rat embryos. The basement membranes were visualized with polyclonal antibodies against laminin. Furthermore, cell degeneration has been investigated in relation to neural crest activity, and discontinuities of the basement membrane in 9- to 30-somite mouse embryos.The separation of the basement membranes of neural and surface ectoderm in the midline is a final phase during the fusion of the neural folds, which takes place from the closure of the rostral neuropore, at the 19-somite stage, until the 27-somite stage (rat embryos), and which occurs focally with variation in the midsagittal and the transverse planes. In the prosencephalon, neural crest activity is absent during the separation phase of both epithelia, but cell degeneration may contribute to the separation of the initially connected basement membranes. A disturbance in the separation of the neural and surface ectoderm may be the pathogenetic basis of midline skull defects, and of the fronto-ethmoidal encephalocele in particular.     

Ultrastructural observations on closure of the neural tube in the mouse

Summary The fusion of the neural walls in the cephalic part of mouse embryos varying in age from 9 to 20 somites was examined with the electron microscope. In the rhombencephalic region the rim of the neural wall was formed from outside inward by ectodermal surface cells, a row of flattened cells without surface projections and neuroepithelial cells. At the junction of the surface ectoderm and the flat cells were seen large projections containing a cytoplasmic matrix without organelles and previously referred to as ruffles. The initial contact between the walls was made by the large cytoplasmic arms and numerous finger-like projections interdigitating with similar projections from the opposite wall. The projections originated from the surface ectoderm and possibly neural crest cells. During further fusion the surface ectoderm cells formed dense membrane specializations, thus establishing a firm contact.The initial contact in the mesencephalon was formed by extensions from the surface ectoderm and was followed by the formation of specialized membrane junctions, as seen between the surface ectoderm in the rhombencephalon. The neuroepithelial cells facing the gap between the neural walls with their apical ends made contact with the cells from the opposing wall by numerous finger-like projections but membrane specializations failed to develop.The closing mechanism in the prosencephalon and anterior neuropore regions differed from the previous areas in that the initial contact was established by the neuroepithelial cells. Only after this contact had been formed did the surface ectoderm cells close the gap. In contrast with the other areas many phagocytosed particles were seen in the prosencephalon and in the region of the anterior neuropore. Many particles from degenerated cells were found inside healthy surrounding cells. Some of these particles contained nuclear material and cytoplasmic organelles.     

The mechanism of cervical flexure formation in the chick

Summary Chick embryos, during stages 14 to 25, undergo an arching of the hindbrain and cervical neural tube that is termed cervical flexure. We have found that if the truncus arteriosus is severed during stage 12–13, the embryos survive for more than 24 h and do not show cervical flexure. The embryos have a beating heart, the expected number of somites, and often have discernible wing and leg buds. Light and electron micrographs reveal no histological abnormalities. The percentage of cells that become labeled with tritiated thymidine is close to normal, indicating that most of the cells are healthy. These results suggest that cervical flexure is related to normal morphogenesis of the heart. At stage 10, the heart is almost straight, with the prospective ventricle cranial to the prospective sinus venosus. The heart tube loops between stage 10 and stage 23, first to the right and then caudad, so that the ventricle becomes caudal to the sinus venosus. The heart undergoes these morphogenetic movements autonomously. The truncus arteriosus does not increase in length during caudal movement of the ventricle, so the cervical region is pulled into an arch. Bending of the cervical region into an arch can be prevented in intact embryos by injecting agar into the foregut, so that the foregut cannot bend. However, after about 24 h of further growth, if the axis cannot bend, the truncus begins to leak blood and the embryo dies. We conclude that cervical flexure is a response of the embryonic axis to the morphogenesis of the heart.     

The role of extracardiac factors in normal and abnormal development of the chick embryo heart: cranial flexure and ventral thoracic wall

The present study was designed to investigate whether the formation of the cranial flexure is involved in the normal positional changes of the embryonic heart tube that occur during its transformation from the c- to s-shaped loop. For this purpose, the formation of the cranial flexure was locally suppressed in chick embryos by introducing a straight hair into the neural canal. In the experimental embryos, prevention of cranial flexure did not suppress the normal positional changes of the heart tube. However, other anomalies in the looping of the heart tube were frequently observed. These anomalies were caused by alterations in the formation of the ventral thoracic wall, which in turn seemed to be related not to the prevention of the cranial flexure but rather to accidental injuries during the implantation of the hair. In the embryos with abnormal looping of the heart tube, the incidence of delayed/defective septation of the heart was significantly higher than in embryos with normal looping. These results show that in the chick embryo: (1) cranial flexure is not involved in normal positional changes of the heart loop; (2) manipulations at the head region of the embryo can unintentionally result in developmental disorders of the ventral thoracic wall; (3) such disorders can result in congenital heart defects through mechanical interference with normal looping of the embryonic heart. The possible significance of these findings for the evaluation of experimental studies of chick embryos is discussed in the context of anomalies observed after surgical ablation of the premigratory cranial neural crest.     

Contributions of placodal and neural crest cells to avian cranial peripheral ganglia

The method of embryonic tissue transplantation was used to confirm the dual origin of avian cranial sensory ganglia, to map precise locations of the anlagen of these sensory neurons, and to identify placodal and neural crest-derived neurons within ganglia. Segments of neural crest or strips of presumptive placodal ectoderm were excised from chick embryos and replaced with homologous tissues from quail embryos, whose cells contain a heterochromatin marker. Placode-derived neurons associated with cranial nerves V, VII, IX, and X are located distal to crest-derived neurons. The generally larger, embryonic placodal neurons are found in the distal portions of both lobes of the trigeminal ganglion, and in the geniculate, petrosal and nodose ganglia. Crest-derived neurons are found in the proximal trigeminal ganglion and in the combined proximal ganglion of cranial nerves IX and X. Neurons in the vestibular and acoustic ganglia of cranial nerve VIII derive from placodal ectoderm with the exception of a few neural crest-derived neurons localized to regions within the vestibular ganglion. Schwann sheath cells and satellite cells associated with all these ganglia originate from neural crest. The ganglionic anlagen are arranged in cranial to caudal sequence from the level of the mesencephalon through the third somite. Presumptive placodal ectoderm for the VIIIth, the Vth, and the VIIth, IXth, and Xth ganglia are located in a medial to lateral fashion during early stages of development reflecting, respectively, the dorsolateral, intermediate, and epibranchial positions of these neurogenic placodes.     

Neural crest and placodal contributions in the development of the glossopharyngeal-vagal complex in the chick

By using the method of quail-to-chick transplantation of neural crest in one series (VNG) and placodal ectoderm in a second series (VPG) we were able to determine the relative contribution of cranial neural crest and placodal ectoderm to the formation of the Glossopharyngeal-vagal complex. In chimeric embryos, quail cells originating from cranial neural crest grafts of postotic levels end up in the root ganglia, while quail cells originating from placodal ectoderm of postotic levels end up in the trunk ganglia. The results clearly indicate that the caudal levels of the medulla and rostral cervical segments represent the site, and the neural crest the source, for the neurons of the root ganglia. The neurons form a homogenous population of the small-cell type. This clearly rules out any contribution to the root ganglia from placodal ectoderm. On the basis of our experiments, it is also concluded that the neurons of the trunk ganglia are purely placodal in origin and are composed of a population of cells of the large-cell type. Our experiments also provide convincing evidence for a neural crest origin for Schwann cell and ganglionic Satellite cells.     

Localization of cells of the prospective neural plate, heart and somites within the primitive streak and epiblast of avian embryos at intermediate primitive-streak stages

By constructing avian transplantation chimeras using fluorescently-labeled grafts and antibodies specific for grafted cells, we have generated a prospective fate map of the primitive streak and epiblast of the avian blastoderm at intermediate primitive-streak stages (stages 3a/3b). This high-resolution map confirms our previous study on the origin of the cardiovascular system from the primitive streak at these stages and provides new information on the epiblast origin of the neural plate, heart and somites. In addition, the origin of the rostral endoderm is now documented in more detail. The map shows that the prospective neural plate arises from the epiblast in close association with the rostral end of the primitive streak and lies within an area extending 250 microm rostral to the streak, 250 microm lateral to the streak and 125 microm caudal to the rostral border of the streak. The future floor plate of the neural tube arises within the midline just rostral to the streak, confirming our earlier study, but unlike at the late-primitive streak stages when both Hensen's node and the midline area rostral to Hensen's node contribute to the floor plate, only the area rostral to the primitive streak contributes to the floor plate at intermediate primitive-streak stages. Instead of contributing to the floor plate of the neural tube, the rostral end of the primitive streak at intermediate primitive-streak stages forms the notochord as well as the rostromedial endoderm, which lies beneath the prechordal plate mesoderm and extends caudolaterally on each side toward the cardiogenic areas. The epiblast lateral to the primitive streak and caudal to the neural plate contributes to the heart and it does so in rostrocaudal sequence (i.e., rostral grafts contribute to rostral levels of the straight heart tube, whereas progressively more caudal grafts contribute to progressively more caudal levels of the straight heart tube), and individual epiblast grafts contribute cells to both the myocardium and endocardium. The prospective somites (i.e., paraxial mesoderm) lie within the epiblast just lateral to the prospective heart mesoderm. Comparing this map with that constructed at late primitive-streak stages reveals that by the late primitive-streak stages, prospective heart mesoderm has moved from the epiblast through the primitive streak and into the mesodermal mantle, and that some of the prospective somitic mesoderm has entered the primitive streak and is undergoing ingression.     

Attachment of neural crest cells to endogenous extracellular matrices

Newly emerging neural crest (NC) cells will enter either the lateral pathway under the surface ectoderm or the vental pathway along the neural tube depending on the axial level (Pratt et al.: Dev. Biol., 44:298-305, 1975; Thiery et al.: Dev. Biol., 93:324-343, 1982; Newgreen et al.: Cell Tissue Res., 221:521-549, 1982; LeDouarin et al.: In: The Role of Extracellular Matrix in Development. Alan R. Liss, Inc., New York, pp. 373-398, 1984; Brauer et al.: Anat. Rec., 211:57-68, 1985). A number of studies have shown a correlation between the type of extracellular matrix (ECM) associated with adjacent tissues (e.g., ectoderm, neural tube, and mesoderm) and the initial pathway taken by NC cells. Our working hypothesis is that the direction of NC cell migration (ventral vs. lateral pathway) depends on the composition of the ECM associated with the surface ectoderm and its ability to support NC cell attachment. In this study, we tested this hypothesis by isolating endogenous ECM associated with the ectoderm of each region and examining the ability of each endogenous ECM to support cranial and trunk NC cell attachment in vitro. Results indicated that both cranial and trunk NC cells preferentially attached to cranial ectodermal ECM as compared to trunk ectodermal ECM. The differences in NC cell attachment were not due to a preferential adsorption of cranial ectodermal ECM onto the ECM-conditioned plastic substrate over trunk ectodermal since approximately equal amounts of ECM bound to the plastic. These results supported the hypothesis and provide evidence that endogenous ectodermal ECM may be one factor potentially responsible for directing the NC cells along a ventral or a lateral pathway.     

Chordate evolution and the origin of craniates: an old brain in a new head

The earliest craniates achieved a unique condition among bilaterally symmetrical animals: they possessed enlarged, elaborated brains with paired sense organs and unique derivatives of neural crest and placodal tissues, including peripheral sensory ganglia, visceral arches, and head skeleton. The craniate sister taxon, cephalochordates, has rostral portions of the neuraxis that are homologous to some of the major divisions of craniate brains. Moreover, recent data indicate that many genes involved in patterning the nervous system are common to all bilaterally symmetrical animals and have been inherited from a common ancestor. Craniates, thus, have an "old" brain in a new head, due to re-expression of these anciently acquired genes. The transition to the craniate brain from a cephalochordate-like ancestral form may have involved a mediolateral shift in expression of the genes that specify nervous system development from various parts of the ectoderm. It is suggested here that the transition was sequential. The first step involved the presence of paired, lateral eyes, elaboration of the alar plate, and enhancement of the descending visual pathway to brainstem motor centers. Subsequently, this central visual pathway served as a template for the additional sensory systems that were elaborated and/or augmented with the "bloom" of migratory neural crest and placodes. This model accounts for the marked uniformity of pattern across central sensory pathways and for the lack of any neural crest-placode cranial nerve for either the diencephalon or mesencephalon. Anat Rec (New Anat) 261:111-125, 2000.     

Evidence against involvement of Bmp receptor 1b signaling in fate specification of the chick mesencephalic alar plate at HH16

Studies in the developing spinal cord have established that morphogenes secreted from the roof- and floor plate influence pattern formation along the dorsal-ventral axis of the neural tube. Bone morphogenetic proteins (Bmps), secreted from the roof plate, act on the more laterally located alar plates to induce position dependent gene expression and cell fate changes. The dorsalizing activity of Bmps is counteracted by Sonic hedgehog (Shh), which is secreted from the floor plate and underlying notochord. Bmps are also expressed in the roof plate of the mesencephalic vesicle, yet it is unclear at present if they also provide patterning information to the mesencephalic alar plates. We have experimentally tested the hypothesis that Bmp signaling is required for fate specification of the mesencephalic alar plate by manipulating Bmp receptor signaling in the early chick embryo through ectopic expression of mutated forms of Bmp receptor 1b (BmpR1b), which render the receptor constitutively active or dominant negative, respectively. In contrast to published data on the embryonic spinal cord, neither activation nor blockage of BmpR1b signaling in stage 16 embryos altered expression of markers of the mesencephalic alar plates including Pax3, Pax7, Meis2 and efnb1. Moreover, simultaneous activation of BmpR1b signaling and blockage of Shh signaling was not sufficient to induce Meis2 expression in the ventral mesencephalon. Therefore, whereas the importance of Bmp signaling for dorsal differentiation in the spinal cord is well established, it appears to play a less prominent role in the dorsal specification of the developing mesencephalon during the same developmental stages.     

Avian photogenic epilepsy and embryonic brain chimeras: neuronal activity of the adult prosencephalon and mesencephalon

Photogenic genetic epilepsy was studied in an avian model, using either the Fayoumi epileptic chicken (Fepi) or neural chimeras obtained by replacement of em bryonic brain vesicles in normal chickens with those of Fepi embryos. In these two kinds of animals motor seizures accompanied by electroencephalographic (EEG) desynchronization and flattening (DF) were evoked by intermittent light stimulation (ILS). In chimeras with on ly the prosencephalon grafted, motor seizures were less severe but DF remained. ILS-induced DF persisted un der paralysis by gallamine triethiodide (Flaxedil). Extra cellular recordings were made in the prosencephalon (wulst) and in the mesencephalon (optic tectum) of paralysed animals. Units recorded in the prosencephalon of Fepi and chimeras showed abnormal interictal burst ing activity, distinctly different from the non-epileptic Fayoumi heterozygotes (Fhtz) and normal chickens. The mesencephalic units of Fepi and chimeras having both prosencephalon and mesencephalon grafted showed two types of abnormal activities during ILS-induced DF, which were distinct from the non-epileptic chickens: type I neurons displaying early, high sensitivity to ILS fol lowed by a prolonged suppression of activity; type II neurons displaying an early and prolonged suppression of activity. The results are discussed with respect to the brain structures generating ictal and interictal EEG ac tivities and motor seizures.     

Differential expression of beta3 integrin gene in chick and mouse cranial neural crest cells.

RNA in situ hybridization on early chicken embryos revealed that the beta3 integrin gene started to be expressed after Hamburger and Hamilton (HH) stage 6 in the presumptive epidermis adjacent to the neural plate, before closure of the neural tube. The beta3 integrin gene was also strongly expressed in cephalic neural crest cells at the same stage in which they begin their migration but disappeared progressively in these cells along the route they take to the branchial arches. The gene was weakly expressed in the differentiating cranial neural crest cells. The alphaVbeta3 integrin protein complex was also mainly detected in the migratory cephalic neural crest cells. However, during early mouse embryogenesis and in contrast to the chick, the beta3 integrin gene was expressed in the foregut diverticulum and in the heart and not in the cephalic neural crest cells. Therefore, the difference in the beta3 integrin expression suggests that mouse and chicken cranial neural crest cells may have distinct integrin requirements during their ontogenesis.     

Immunocytological study of the differentiation of chick and quail adenohypophysis epithelial rudiments,grafted or cultivated in vitro: Evidence for polypeptidic hormones

Epithelial rudiments of adenohypohysis were removed from chick and quail embryos between days 3 and 5 of development. Chick rudiments were grafted for 11–13 days onto the chorioallantoic membrane of decapitated chick embryo hosts. Quail rudiments were cultivated in vitro for 6 days. Both grafted and cultivated Rathke's pouches differentiated into adenohypophyseal tissue. The adenohypophyseal tissue cultured on chorio-allantoic membrane exhibited cells reacting with the following immune sera: anti-β-(1–24)ACTH, anti-α-(17–39)-ACTH, anti-α-endorphin, anti-β-endorphin and anti-β-LPH, which also gave a positive reaction when applied to adenohypophysis of corresponding age which had differentiated in situ. In situ, corticotrophs were located exclusively in the cephalic lobe of adenohypophysis. Therefore, the differentiation of corticotrophs in the whole graft, i.e., from both cephalic and caudal lobes of Rathke's pouch, showed that the cells of the caudal lobe, or at least some of them, were uncommitted when the rudiment was removed. In vitro, tissue derived from Rathke's pouch contained cells reacting with antibodies to β-(1–24)-ACTH, α-(17–39)-ACTH, and β-LPH, as did adenohypophysis from quail embryos of corresponding age (9–10 days), differentiated in situ. The differentiation of quail Rathke's pouch in vitro corroborates that differentiation can occur without influence from hypothalamus and, moreover, shows that at least some kinds of cells can differentiate without influence exerted by any other encephalic factors, and in the absence of mesenchyme. The question arises whether fibroblastic cells derived from Rathke's pouch cells act as feeder-cells and/or secrete some factors promoting differentiation.     

Antagonists of Wnt and BMP signaling promote the formation of vertebrate head muscle

Recent studies have postulated that distinct regulatory cascades control myogenic differentiation in the head and the trunk. However, although the tissues and signaling molecules that induce skeletal myogenesis in the trunk have been identified, the source of the signals that trigger skeletal muscle formation in the head remain obscure. Here we show that although myogenesis in the trunk paraxial mesoderm is induced by Wnt signals from the dorsal neural tube, myogenesis in the cranial paraxial mesoderm is blocked by these same signals. In addition, BMP family members that are expressed in both the dorsal neural tube and surface ectoderm are also potent inhibitors of myogenesis in the cranial paraxial mesoderm. We provide evidence suggesting that skeletal myogenesis in the head is induced by the BMP inhibitors, Noggin and Gremlin, and the Wnt inhibitor, Frzb. These molecules are secreted by both cranial neural crest cells and by other tissues surrounding the cranial muscle anlagen. Our findings demonstrate that head muscle formation is locally repressed by Wnt and BMP signals and induced by antagonists of these signaling pathways secreted by adjacent tissues.