Early development of the chick embryo

The ultrastructure of the early chick embryo was investigated, using scanning (SEM) and transmission electron microscopy (TEM). Eggs were obtained from the shell gland by injecting hens intravenously with a synthetic prostaglandin or arginine vasopressin. Embryos were examined during late cleavage (stages IV–VI, Eyal-Giladi and Kochav, '76), formation of the area pellucida (stages VII–XI), and formation of the hypoblast (stages X–XIV). SEM highlighted the reduction in cell number at the underside of the embryo during formation of the area pellucida although it became apparent that the thickness of the embryo is not reduced to a single layer of cells at stage X. In addition, blastomeres at the perimeter of embryos (stages V–VI) project filopodial extensions onto a smooth membrane that separates the sub-embryonic cavity from the yolk. During hypoblast formation, epiblast cells generate stellate projections at their basal aspect, thus providing a meshwork for the advancing secondary hypoblast cells. By stage XII the epiblast was one cell thick and reminiscent of a columnar epithelium when viewed transversely. Cells of the deep portion of the posterior marginal zone were distinguished morphologically in the stage XII embryo by their many cell surface projections and ruffled appearance. Blastomeres at the perimeter of stage V–VI embryos projected filopodial extensions onto a smooth membrane which separates the sub-embryonic cavity from the yolk. This membrane is presumed to be confluent with the cytolemma. Evidence is presented demonstrating the presence of intracellular membrane-bound droplets which are hypothesised to contain sub-embryonic fluid. © 1993 Wiley-Liss, Inc.     

Origin of primordial germ cells in the prestreak chick embryo

The temporal and spatial pattern of segregation of the avian germline from the formation of the area pellucida to the beginning of primitive streak formation (stages VII–XIV, EG&K) was investigated using the culture of whole embryos and central and peripheral embryo fragments on vilelline membranes at stages VII–IX, immunohistological analysis of whole mount embryos and sections with monoclonal antibodies MC-480 against stage-specific embryonic antigen-1 (SSEA-1) and EMA-1, and with the culture of dispersed blastoderms at stages IX–XIV with and without an STO feeder layer. Whole embryos at intrauterine stages developed up to the formation of the primitive streak despite the absence of area pellucida expansion. Primordial germ cells (PGCs) appeared in the cultures of whole embryos and only in central fragments containing a partially formed area pellucida at stages VII–IX. When individual stage IX–XIV embryos were dispersed and cultured without a feeder layer, 25–45 PGCs/embryo were detected only with stage X–XIV, but not with stage IX blastoderms. However, the culture of dispersed cells from the area pellucida of stages IX–XIII on STO feeder layers yielded about 150 PGCs/embryo. The carbohydrate epitopes recognized by anti-SSEA-1 and EMA-1 first appeared at stage X on cells in association with polyingressing cells on the ventral surface of the epiblast and later on the dorsal surface of the hypoblast. The SSEA-1-positive hypoblast cells gave rise to chicken PGCs when cultured on a feeder layer of quail blastodermal cells. From these observations, we propose that the segregation and development of avian germline is a gradual, epigenetic process associated with the translocation of SSEA-1/EMA-1-positive cells from the ventral surface of the area pellucida at stage X to the dorsal side of the hypoblast at stages XI–XIV. © 1996 Wiley-Liss, Inc.     

Primary hypoblast development in the chick

Summary Scanning electron microscopy (SEM) indicates that the primary hypoblast forms beneath the area pellucida during the first 8 h of incubation mainly by establishment of contact among cells which move downward out of the epiblast. This movement, polyingression, begins posteriorly and continues antero-laterally during the period of primary hypoblast formation. Polyingression produces many pits and possibly a crescentic fold in the embryo upper surface with corresponding cell clusters and a ridge on the lower surface. Fixationin situ helps prevent formation of artifactual folds and wrinkles facilitating interpretation of the SEM images.Formation of intercellular adhesions which lead to development of an epithelial primary hypoblast proceeds in a posterior to anterior direction along with polyingression. This epithelialization begins with elaboration of numerous filamentous processes by cells as they arrive from the epiblast, and continues with ongoing input of cells, merging of cells and cell clusters, and cell flattening. We have also shown (Weinberger and Brick 1982) that proliferation of ingressing cells provides additional cells for hypoblast development.     

Behaviour of dissociated hypoblast cells on the basal lamina and on extracellular fibrils in the gastrulating chicken embryo.

The spreading behaviour of dissociated hypoblast cells on and besides a band of aligned fibrils associated with the basal lamina of the epiblast was investigated by the use of scanning electron microscopy. A horse-shaped band of aligned fibrils, first demonstrated by Wakely and England (1979), is present during the gastrulation stages of chicken embryos on the ventral side of the epiblast at the cranial and lateral borders of the area pellucida. The basal lamina of the area pellucida situated inside the fibrillar band enables the spreading and probably the locomotion of dissociated cells, which appeared as polarized cells. Numerous cells were also found on the fibrillar band, and these cells lacked distinct lamellae and a polarized shape. Extensions of the cells contacted the extracellular fibrils and, at these sites of contact, the pattern of the fibrils was frequently deformed. From these observations and from previous results emerged the concept that spreading and locomotion of dissociated hypoblast cells, as well as single mesoblast cells and healing hypoblast epithelium, are inhibited by the band of extracellular fibrils, which acts as a physical barrier. The cell biological basis of the mechanism by which extracellular fibrils associated with the basal lamina arrest the migration of hypoblast and mesoblast cells, but guide the migration of primordial germ cells, is discussed.     

Nuclear beta-catenin and the development of bilateral symmetry in normal and LiCl-exposed chick embryos.

Studies in Xenopus laevis and zebrafish suggest a key role for beta-catenin in the specification of the axis of bilateral symmetry. In these organisms, nuclear beta-catenin demarcates the dorsalizing centers. We have asked whether beta-catenin plays a comparable role in the chick embryo and how it is adapted to the particular developmental constraints of chick development. The first nuclear localization of beta-catenin is observed in late intrauterine stages of development in the periphery of the blastoderm, the developing area opaca and marginal zone. Obviously, this early, radially symmetric domain does not predict the future organizing center of the embryo. During further development, cells containing nuclear beta-catenin spread under the epiblast and form the secondary hypoblast. The onset of hypoblast formation thus demarcates the first bilateral symmetry in nuclear beta-catenin distribution. Lithium chloride exposure also causes ectopic nuclear localization of beta-catenin in cells of the epiblast in the area pellucida. Embryos treated before primitive streak formation become completely radialized, as shown by the expression of molecular markers, CMIX and GSC. Lithium treatments performed during early or medium streak stages cause excessive development of the anterior primitive streak, node and notochord, and lead to a degeneration of prospective ventral and posterior structures, as shown by the expression of the molecular markers GSC, CNOT1, BMP2 and Ch-Tbx6L. In summary, we found that in spite of remarkable spatiotemporal differences, beta-catenin acts in the chick in a manner similar to that in fish and amphibia.     

From cleavage to primitive streak formation: A complementary normal table and a new look at the first stages of the development of the chick: II. Microscopic anatomy and cell population dynamics

The microscopic anatomy of uterine and freshly laid unincubated and briefly incubated chick germs is described. Special attention is paid to the difference between the three developmental periods involved: cleavage, area pellucida formation, and primary hypoblast formation. During cleavage the cytoplasm of the germinal disc divides into blastomeres, which become constantly smaller, and the subgerminal cavity is formed. The germ is accumulating extensive glycogen reserves for utilization during the next period. The most fascinating period is the formation of the area pellucida, which arises as a result of a polarized cell-shedding process. During this process all the subepithelial cells round up and fall into the subblastodermic cavity, where they assemble beneath the future anterior side of the blastoderm. The cell-shedding process is presumably energy consuming and the glycogen reserves are utilized as cell shedding progresses, starting at the posterior and terminating at the anterior side of the germ. The germ loses about one-fifth of its initial cytoplasmic mass during this process. The formation of the primary hypoblast is again polarized, posterioanteriorly. The onset of the process of polyinvagination takes place concomitantly with the shedding of the last subepithelial cells.     

The effect of ambient temperature on turkey embryonic development at oviposition

Stage of embryonic development at oviposition was measured in turkey breeder hens maintained in relatively warm and cool environments. The premise was that variations in embryonic development at oviposition might account for the decreased hatchability associated with warm summer temperatures. No treatment effect was found, as judged by somite counts after 52 h of incubation. Variation in embryonic development was as great within a hen as between hens, indicating that causative factor(s) other than stage of development at oviposition is the reason for reduced hatch of fertile eggs during periods of relatively high environmental temperature.     

Characterization of stage-specific embryonic antigen-1 (SSEA-1) expression during early development of the turkey embryo.

SSEA-1 is a carbohydrate epitope associated with cell adhesion, migration and differentiation. In the present study, SSEA-1 expression was characterized during turkey embryogenesis with an emphasis on its role in primordial germ cell development. During hypoblast formation, SSEA-1 positive cells were identified in the blastocoel and hypoblast and later in the germinal crescent. Based on location and morphology, these cells were identified, as PGCs. Germ cells circulating through embryonic blood vessels were also SSEA-1 positive. During the active phase of migration, PGCs in the dorsal mesentery and gonad could no longer be identified using the SSEA-1 antibody. The presence of PGCs at corresponding stages was verified using periodic acid Schiff stain. Pretreatment of PGCs with trypsin, alpha-galactosidase and neuraminidase did not restore immunoreactivity to SSEA-1. In general, expression was not limited to the germ cell lineage. SSEA-1 was also detected on the ectoderm, yolk sac endoderm, gut and mesonephric tubules. During neural tube closure, SSEA-1 was expressed by the neural epithelium of the fusing neural folds. Later SSEA-1 was detected in regions of the developing spinal cord. Enzyme pretreatment unmasked the epitope on some neural crest cells and cells in the sympathetic ganglion. The temporal and spatial distribution of SSEA-1 in the turkey embryo suggests a role in early germ cell and neural cell development. The absence of SSEA-1 on turkey gonadal germ cells was different from that observed for the chick. Therefore, while features of avian germ cell development appear to be conserved, expression of SSEA-1 can vary with the species.     

Isolation of chicken vasa homolog gene and tracing the origin of primordial germ cells

To obtain a reliable molecular probe to trace the origin of germ cell lineages in birds, we isolated a chicken homolog (Cvh) to vasa gene (vas), which plays an essential role in germline formation in Drosophila. We demonstrate the germline-specific expression of CVH protein throughout all stages of development. Immunohistochemical analyses using specific antibody raised against CVH protein indicated that CVH protein was localized in cytoplasm of germ cells ranging from presumptive primordial germ cells (PGCs) in uterine-stage embryos to spermatids and oocytes in adult gonads. During the early cleavages, CVH protein was restrictively localized in the basal portion of the cleavage furrow. About 30 CVH-expressing cells were scattered in the central zone of the area pellucida at stage X, later 45-60 cells were found in the hypoblast layer and subsequently 200-250 positive cells were found anteriorly in the germinal crescent due to morphogenetic movement. Furthermore, in the oocytes, CVH protein was predominantly localized in granulofibrillar structures surrounding the mitochondrial cloud and spectrin protein-enriched structure, indicating that the CVH-containing cytoplasmic structure is the precursory germ plasm in the chicken. These results strongly suggest that the chicken germline is determined by maternally inherited factors in the germ plasm.     

Transfer of extracellular matrix components between germ layers in chimaeric chicken-quail blastoderms

Summary A chemical basis for the transmission of signals during gastrulation has been investigated by using chimaeric embryos resulting from the combination of ³H-glucosamine-labelled and unlabelled hypoblast with epiblast taken from chicken and quail embryos at stage 3 of Vakaet (1970). The ability to distinguish chicken from quail cells on the basis of their different nuclear distribution of heterochromatin after Feulgen staining made it possible to determine the origin of the cells in the chimaerae. Tritiated quail hypoblast (after incubation of the embryo in the presence of ³H-glucosamine) was transplanted onto unlabelled chicken blastoderm deprived of its hypoblast. After culture of the chimaera for 5 h, the autoradiographic pattern shows silver grains not only over the graft, but also at the ventral surface of the epiblast of the host. Transfer of label may occur to mesoblast cells, but not between chicken and quail hypoblast cells. Chase experiments exclude the possibility that unprocessed, tritiated glucosamine is transferred. Chemical fixation of the host before transplantation of a labelled quail hypoblast also allows visualization of a transfer of macromolecules from hypoblast to the basement membrane of the epiblast, suggesting that an intervention of the epiblast cells in this process is not necessary. The morphology of the chimaeric embryos, as studied by scanning electron microscopy, suggests a direct deposition of these macromolecules by filopodia of the dorsal surface of the hypoblast. The possibility of diffusion of free macromolecules has been considered and can reasonably be discarded on the basis of several observations. The reverse experiment, in which unlabelled quail hypoblast and possibly some mesoblast have been combined with a tritiated host deprived of its hypoblast, also shows the transfer of label from the host to the cellular surface of the graft. A two-way exchange of glucosamine-containing molecules thus occurs in the blastoderm. It is hypothesized that: (1) low molecular weight compounds, macromolecular material, and/or catabolic products, are exchanged between the different germ layers during gastrulation; (2) the components of the extracellular matrix turn over and are continuously changing; (3) this transfer is a possible mechanism of transmission for developmental or inductive signals during embryonic development. The present results also demonstrate the participation of underlying tissue in the biosynthesis of basement membrane components of an epithelium.     

Nucleolar ontogenesis in the uterine chick germ correlated with morphogenetic events

Nucleolar development in the cleaving chick germ up to the formation of the primary hypoblast was followed through a series of well-defined uterine and early incubated stages both by light and electron microscopy. Well-established criteria of nucleolar morphology were used for determining the developmental stage of onset of rRNA synthesis. By these criteria rRNA synthesis was first observed at midcleavage in uterine stage VII [1] germs. This could be correlated with the first morphogenetic event—the posterio-anteriorly orientated formation of the area pellucida which results in a bilaterally symmetrical blastoderm.     

The hypoblast of the chick embryo positions the primitive streak by antagonizing nodal signaling

The hypoblast (equivalent to the mouse anterior visceral endoderm) of the chick embryo plays a role in regulating embryonic polarity. Surprisingly, hypoblast removal causes multiple embryonic axes to form, suggesting that it emits an inhibitor of axis formation. We show that Cerberus (a multifunctional antagonist of Nodal, Wnt, and BMP signaling) is produced by the hypoblast and inhibits primitive streak formation. This activity is mimicked by Cerberus-Short (CerS), which only inhibits Nodal. Nodal misexpression can initiate an ectopic primitive streak, but only when the hypoblast is removed. We propose that, during normal development, the primitive streak forms only when the hypoblast is displaced away from the posterior margin by the endoblast, which lacks Cerberus.     

Hypoblast controls mesoderm generation and axial patterning in the gastrulating rabbit embryo

Gastrulation in higher vertebrate species classically commences with the generation of mesoderm cells in the primitive streak by epithelio-mesenchymal transformation of epiblast cells. However, the primitive streak also marks, with its longitudinal orientation in the posterior part of the conceptus, the anterior-posterior (or head-tail) axis of the embryo. Results obtained in chick and mouse suggest that signals secreted by the hypoblast (or visceral endoderm), the extraembryonic tissue covering the epiblast ventrally, antagonise the mesoderm induction cascade in the anterior part of the epiblast and thereby restrict streak development to the posterior pole (and possibly initiate head development anteriorly). In this paper we took advantage of the disc-shape morphology of the rabbit gastrula for defining the expression compartments of the signalling molecules Cerberus and Dickkopf at pre-gastrulation and early gastrulation stages in a mammal other than the mouse. The two molecules are expressed in novel expression compartments in a complementary fashion both in the hypoblast and in the emerging primitive streak. In loss-of-function experiments, carried out in a New-type culturing system, hypoblast was removed prior to culture at defined stages before and at the beginning of gastrulation. The epiblast shows a stage-dependent and topographically restricted susceptibility to express Brachyury, a T-box gene pivotal for mesoderm formation, and to transform into (histologically proven) mesoderm. These results confirm for the mammalian embryo that the anterior-posterior axis of the conceptus is formed first as a molecular prepattern in the hypoblast and then irrevocably fixed, under the control of signals secreted from the hypoblast, by epithelio-mesenchymal transformation (primitive streak formation) in the epiblast.Edited by D. Tautz     

The Tridimensional Morphology of Rice Embryo Development with Special Reference to the Scutellum and the Coleoptile

Using scanning electron microscopy and semi-thin plastic sections, the pattern of development of the rice ( Oryza sativa L. ) embryo from 2 days after pollination (DAP) to maturity was followed. ( 1 ) At 2 DAP, the young embryo was observed to consist of an embryo proper, a hypoblast and a suspensor. The trum-pet-shaped hypoblast was a transitional region situated between the suspensor and the embryo proper. To label the hypoblast as suspensor is incorrect. During this time, dorsiventrality was established, but a radicle was not yet differentiated. Therefore it is still referred to as a proembryo. (2) 3 ~ 5 DAP, the embryo underwent definite morphological and anatomical changes. In the young embryo at 3 DAP the scutellum and colcoptile appeared simultaneously directly from the proembryo. The coleoptile did not originate from the scutellmn. During these foremost 3 days, the coleoptile primordium underwent a special kind of morphological change and formed a young coleeptile having the shape of an inverted hollow cone. This process revealed the true mechanism of c61eeptile formation. Anatomical observation indicated that the embryo at 3 DAP began to differentiate procambium, ground meristem and root cap. At 4 DAP a dome-like growth cone and protoderm of radicle appeared. Then the shoot-root axis became established. At 5 DAP the plumule, hypocotyl and radicle were formed. (3) It was shown that the embryo of rice actually has two cotyledons: the scutellum (a part of the embryonic envelope) and the coleeptile (The scutellum being the lateral cotyledon, a part of outside cotyledon, and the coleoptile the apical cotyledon--the coleoptile may be considered to be a modified form of a cotyledon). This kind of structural arrangemem can be referred to as dimorphic cotyledon.     

Activation of epiblast gene expression by the hypoblast layer in the prestreak chick embryo

Axis formation is a highly regulated process in vertebrate embryos. In mammals, inductive interactions between an extra-embryonic layer, the visceral endoderm, and the embryonic layer before gastrulation are critical both for anterior neural patterning and normal primitive streak formation. The role(s) of the equivalent extra-embryonic endodermal layer in the chick, the hypoblast, is still less clear, and dramatic effects of hypoblast on embryonic gene expression have yet to be demonstrated. We present evidence that two genes later associated with the gastrula organizer (Gnot-1 and Gnot-2) are induced by hypoblast signals in prestreak embryos. The significance of this induction by hypoblast is discussed in terms of possible hypoblast functions and the regulation of axis formation in the early embryo. Several factors known to be expressed in hypoblast, and retinoic acid, synergistically induce Gnot-1 and Gnot-2 expression in blastoderm cell culture. The presence of retinoic acid in prestreak embryos has not yet been directly demonstrated, but exogenous retinoic acid appears to mimic the effects of hypoblast rotation on primitive streak extension, raising the possibility that retinoid signaling plays some role in the pregastrula embryo.     

The determination of myogenic and cartilage cells in the early chick embryo and the modifying effect of retinoic acid

Summary The time of determination of cartilage and skeletal muscle was studied by making chimeric grafts or explants of small tissue pieces from several stages of early chick or quail embryos. Chondrogenesis was assessed by histology or with antibodies directed against type II collagen or cartilage proteoglycan, while myogenesis was detected immunohistochemically with antibodies directed against 3 different muscle markers, including muscle myosin. Grafts from Hensen's node, primitive streak and segmental plate of donor embryos of Stage 3–5 (Hamburger and Hamilton) were transplanted under the ectoderm in the extraembryonic area of Stage 12 host embryos. In addition, explants and mesodermal cells were cultured on glass in DMEM+F12 medium supplemented with 10% FCS. The results showed that determined myogenic cells could first be detected in Hensen's node and the primitive streak at Stage 3⁺–4 and that they developed from mesodermal cells located between the epiblast and hypoblast. Myogenic cells also appeared in grafted and explanted segmental plate with or without notochord from Stage 5 embryos. On the other hand, cartilage cells only formed in grafted and explanted segmental plate that also contained notochord. RA (1 ng/ml) could induce the formation of cartilage cells in the explanted primitive streak without Hensen's node or notochord taken from Stage 3–5 embryos and could also promote the differentiation of myogenic cells in primitive streak from Stage 3 embryo. Thus RA can substitute for Hensen's node or the notochord in the induction of cartilage cells and has some stimulatory effects on the differentiation of myogenic cells. Additional evidence indicates that the hypoblast might play an inductive role in the formation of the notochord which may subsequently promote the differentiation of cartilage cells. Offprint requests to: M. Solursh     

玉米胚胎发育、萌发与胚的结构及子叶二型性

运用扫描电镜与半薄切片技术,观察了玉米(Zea mays L.)的胚发育过程,得到以下认识:第一、关于原胚.玉米合子细胞分裂形成的原胚分为胚柄、胚基与胚体三部分.胚柄短小,寿命短暂.胚基具有生长带,纵向伸长长度大,胚基的上部参与形成胚根鞘,其余部分干缩后附在胚根鞘末端.第二、玉米胚的背腹极性及二型子叶.原胚初期胚体出现背腹极性,腹面的细胞小,细胞质稠密,液泡较少;背面的细胞较大,细胞质稠密度略低,液泡较多.原胚后期胚体分化为腹部与背部,腹部从腹面的中央突起,背部在腹部的周围(从左至右侧)及整个胚体背面.进入幼胚时期,腹部分化为胚芽鞘、生长锥、胚轴、胚根和胚根鞘(大部分).期间,胚芽鞘原基和根原始细胞的分化都从胚体的中轴部位开始,然后向两侧和四周扩展,表现出胚体腹面形态的两侧对称性.原胚的背部形成盾片原基,盾片原基经历向左、右、上、下的迅速扩展和加厚的生长,将整个腹部所分化形成的构件藏于盾片的纵沟之中,最后盾片从纵沟的边缘长出的左、右侧鳞均向胚体的中轴线生长,完整显示出玉米胚腹面的两侧对称.玉米胚由腹部顶端形成胚芽鞘和生长锥的情况与水稻胚的胚芽鞘(顶生子叶)和生长锥的形成相同,玉米的胚芽鞘也是顶生子叶,盾片则是侧生子叶.玉米异型子叶的由来在于胚体的背腹极性.第三、玉米胚的真实形态结构及胚胎发育时期的划分.玉米胚是一个胚轴,其顶部是具胚芽鞘的胚芽,中部是具侧生子叶(盾片)的胚轴,下部是具胚根鞘的胚根.盾片从背面到腹面包住整个胚轴系统,在胚的腹面上可见从盾片边缘衍生的左、右侧鳞的边缘相迭,只在接缝线的上、下端留下蝌蚪状的小孔,使胚芽鞘和胚根鞘的末端稍露出.胚胎发育分为4个时期: 1.原胚期--从合子细胞分裂开始至分化背部与腹部为止;2.腹部迅速分化期;3.盾片快速生长期;4.侧鳞生长、胚套形成期.第四、获取垂直于胚腹面正中央纵切面是正确认识玉米胚形态的关键.     

玉米胚胎发育、萌发与胚的结构及子叶二型性

运用扫描电镜与半薄切片技术,观察了玉米(Zea mays L.)的胚发育过程,得到以下认识：第一、关于原胚。玉米合子细胞分裂形成的原胚分为胚柄、胚基与胚体三部分。胚柄短小,寿命短暂。胚基具有生长带,纵向伸长长度大,胚基的上部参与形成胚根鞘,其余部分干缩后附在胚根鞘末端。第二、玉米胚的背腹极性及二型子叶。原胚初期胚体出现背腹极性,腹面的细胞小,细胞质稠密,液泡较少;背面的细胞较大,细胞质稠密度略低,液泡较多。原胚后期胚体分化为腹部与背部,腹部从腹面的中央突起,背部在腹部的周围(从左至右侧)及整个胚体背面。进入幼胚时期,腹部分化为胚芽鞘、生长锥、胚轴、胚根和胚根鞘(大部分)。期间,胚芽鞘原基和根原始细胞的分化都从胚体的中轴部位开始,然后向两侧和四周扩展,表现出胚体腹面形态的两侧对称性。原胚的背部形成盾片原基,盾片原基经历向左、右、上、下的迅速扩展和加厚的生长,将整个腹部所分化形成的构件藏于盾片的纵沟之中,最后盾片从纵沟的边缘长出的左、右侧鳞均向胚体的中轴线生长,完整显示出玉米胚腹面的两侧对称。玉米胚由腹部顶端形成胚芽鞘和生长锥的情况与水稻胚的胚芽鞘(顶生子叶)和生长锥的形成相同,玉米的胚芽鞘也是顶生子叶,盾片则是侧生子叶。玉米异型子叶的由来在于胚体的背腹极性。第三、玉米胚的真实形态结构及胚胎发育时期的划分。玉米胚是一个胚轴,其顶部是具胚芽鞘的胚芽,中部是具侧生子叶(盾片)的胚轴,下部是具胚根鞘的胚根。盾片从背面到腹面包住整个胚轴系统,在胚的腹面上可见从盾片边缘衍生的左、右侧鳞的边缘相迭,只在接缝线的上、下端留下蝌蚪状的小孔,使胚芽鞘和胚根鞘的末端稍露出。胚胎发育分为4个时期：1．原胚期——从合子细胞分裂开始至分化背部与腹部为止;2．腹部迅速分化期;3．盾片快速生长期;4．侧鳞生长、胚套形成期。第四、获取垂盲于胚腹面正中央纵切面是正确认识玉米胚形态的关键。     

Epiblast and primitive-streak origins of the endoderm in the gastrulating chick embryo

Gastrulation is characterized by the extensive movements of cells. Fate mapping is used to follow such cell movements as they occur over time, and prospective fate maps have been constructed for several stages of the model organisms used in modern studies in developmental biology. In chick embryos, detailed fate maps have been constructed for both prospective mesodermal and ectodermal cells. However, the origin and displacement of the prospective endodermal cells during crucial periods in gastrulation remain unclear. This study had three aims. First, we determined the primitive-streak origin of the endoderm using supravital fluorescent markers, and followed the movement of the prospective endodermal cells as they dispersed to generate the definitive endodermal layer. We show that between stages 3a/b and 4, the intraembryonic definitive endoderm receives contributions mainly from the rostral half of the primitive streak, and that endodermal movements parallel those of ingressing adjacent mesodermal subdivisions. Second, the question of the epiblast origin of the endodermal layer was addressed by precisely labeling epiblast cells in a region known to give rise to prospective somitic cells, and following their movement as they underwent ingression through the primitive streak. We show that the epiblast clearly contributes prospective endodermal cells to the primitive streak, and subsequently to definitive endoderm of the area pellucida. Finally, the relationship between the hypoblast and the definitive endoderm was defined by following labeled rostral primitive-streak cells over a short period of time as they contributed to the definitive endoderm, and combining this with in situ hybridization with a riboprobe for Crescent, a marker of the hypoblast. We show that as the definitive endodermal layer is laid down, there is cell-cell intercalation at its interface with the displaced hypoblast cells. These data were used to construct detailed prospective fate maps of the endoderm in the chick embryo, delineating the origins and migrations of endodermal cells in various rostrocaudal levels of the primitive streak during key periods in early development.     

The marginal zone and its contribution to the hypoblast and primitive streak of the chick embryo

The marginal zone of the chick embryo has been shown to play an important role in the formation of the hypoblast and of the primitive streak. In this study, time-lapse filming, fate mapping, ablation and transplantation experiments were combined to study its contribution to these structures. It was found that the deep (endodermal) portion of the posterior marginal zone contributes to the hypoblast and to the junctional endoblast, while the epiblast portion of the same region contributes to the epiblast of the primitive streak and to the definitive (gut) endoderm derived from it. Within the deep part of the posterior marginal zone, a subpopulation of HNK-1-positive cells contributes to the hypoblast. Removal of the deep part of the marginal zone prevents regeneration of the hypoblast but not the formation of a primitive streak. Removal of both layers of the marginal zone leads to a primitive streak of abnormal morphology but mesendodermal cells nevertheless differentiate. These results show that the two main properties of the posterior marginal zone (contributing to the hypoblast and controlling the site of primitive streak formation) are separable, and reside in different germ layers. This conclusion does not support the idea that the influence of the posterior marginal zone on the development of axial structures is due to it being the source of secondary hypoblast cells.