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PubMed1988

Patterns of cholinesterase staining during neural crest cell morphogenesis in mouse and chick embryos.

Cholinesterase (ChE) activity has been reported previously in nonneuronal tissues of a variety of avian and mammalian embryos. We report here a comparison study of ChE staining in chick and mouse embryos. Transmission electron microscopy was used to study the distribution of this activity in neuroepithelial, neural crest, somite, and ectodermal cells. Our cytochemical studies show that the distribution of nonspecific ChE staining in these tissues during neurulation is similar in the two species but that acetylcholinesterase (AChE) staining, previously shown to be intense in the chick, is absent in the mouse; the only cells showing the presence of this enzyme at these stages of development in mouse are blood cells. However, AChE staining does appear later in the brain, in neural tissues derived from the neural crest and, perhaps, in some migratory neural crest cells. The differences between AChE distribution in these two species (and that reported previously in the rabbit) indicate that the timing of first appearance of AChE is unrelated to neuroepithelial morphogenesis or to neural crest cell motility. The correlation between nonspecific cholinesterases and morphogenetic movements, however, is supported by these studies.

Animals↗

PubMed1989

Fate mapping the avian neural plate with quail/chick chimeras: origin of prospective median wedge cells.

The origin of prospective M cells, which are median neuroepithelial cells that become wedge-shaped during bending of the neural plate and eventually form the midline floor of the neural tube, was determined by constructing quail/chick chimeras and using the quail nucleolar marker to identify quail donor cells in chick host blastoderms. Two possible sites of prospective M-cell origin in the epiblast were examined: a single, midline rudiment located just rostral to Hensen's node and paired rudiments flanking the cranial part of the primitive streak. Our results suggest that M cells arise exclusively from the midline, prenodal rudiment. From this rudiment, M cells extend caudally throughout the entire length of the neuroepithelium. This new information on the origin of prospective M cells will aid in the analysis of their role in neurulation.

Animals↗

PubMed1989

Cell intercalation during notochord development in Xenopus laevis.

Morphometric data from scanning electron micrographs (SEM) of cells in intact embryos and high-resolution time-lapse recordings of cell behavior in cultured explants were used to analyze the cellular events underlying the morphogenesis of the notochord during gastrulation and neurulation of Xenopus laevis. The notochord becomes longer, narrower, and thicker as it changes its shape and arrangement and as more cells are added at the posterior end. The events of notochord development fall into three phases. In the first phase, occurring in the late gastrula, the cells of the notochord become distinct from those of the somitic mesoderm on either side. Boundaries form between the two tissues, as motile activity at the boundary is replaced by stabilizing lamelliform protrusions in the plane of the boundary. In the second phase, spanning the late gastrula and early neurula, cell intercalation causes the notochord to narrow, thicken, and lengthen. Its cells elongate and align mediolaterally as they rearrange. Both protrusive activity and its effectiveness are biased: the anterioposterior (AP) margins of the cells advance and retract but produce much less translocation than the more active left and right ends. The cell surfaces composing the lateral boundaries of the notochord remain inactive. In the last phase, lasting from the mid- to late neurula stage, the increasingly flattened cells spread at all their interior margins, transforming the notochord into a cylindrical structure resembling a stack of pizza slices. The notochord is also lengthened by the addition of cells to its posterior end from the circumblastoporal ring of mesoderm. Our results show that directional cell movements underlie cell intercalation and raise specific questions about the cell polarity, contact behavior, and mechanics underlying these movements. They also demonstrate that the notochord is built by several distinct but carefully coordinated processes, each working within a well-defined geometric and mechanical environment.

Animals↗

PubMed1992

Expansion of surface epithelium provides the major extrinsic force for bending of the neural plate.

Neurulation, formation of the neural tube, requires both intrinsic forces (i.e., those generated within the neural plate) and extrinsic forces (i.e., those generated outside the neural plate in adjacent tissues), but the precise origin of these forces is unclear. In this study, we addressed the question of which tissue produces the major extrinsic force driving bending of the neural plate. We have previously shown that 1) extrinsic forces are required for bending and 2) such forces are generated lateral to the neural plate. Three tissues flank the neural plate prior to its bending: surface epithelium, mesoderm, and endoderm. In the present study, we removed two of these layers, namely, the endoderm and mesoderm, underlying and lateral to the neural plate; bending still occurred, often with complete formation of a neural tube, although the latter usually rotated toward the side of tissue depletion. These results suggest that the surface epithelium, the only tissue remaining after microsurgery, provides the major extrinsic force for bending of the neural plate and that the mesoderm (and perhaps endoderm) stabilizes the neuraxis, maintaining its proper orientation and position on the midline.

Animals↗

PubMed1993

Developmental anomalies of Xenopus embryos following microinjection of SPARC antibodies.

The function of SPARC (Secreted Protein, Acidic, Rich in Cysteine) in early embryonic development was assayed by microinjecting affinity-purified antibodies directed against SPARC into the blastocoel cavity of Xenopus embryos. Microinjection of SPARC antibodies did not appear to interfere with development until late neurulation. By hatching, a broad spectrum of external developmental anomalies were observable, including bent embryonic axes, accentuated ventral masses, shortened embryonic axes, and lack of visible eye pigment. Histological sections of injected embryos demonstrated that lack of visible eye pigmentation was often associated with deformities in eye development. Bending and shortening of the embryonic axis was associated with highly disorganized myotome patterns and loss of segmental boundaries. The results indicate a requirement for SPARC in the early morphological development of several tissues in Xenopus.

Animals↗

PubMed1995

Comparative study of Msx-1 expression in early normal and vitamin A-deficient avian embryos.

Homeobox-containing genes may play an important role in establishing embryonic patterns during development of vertebrates. Retinoic acid is able to induce expression of Hox genes in cells in culture and to alter expression patterns in the developing vertebrate embryos. Using wholemount in situ hybridization, we have examined and compared the expression patterns of a homeobox-containing gene, Msx-1, in early normal and vitamin A-deficient quail embryos. At gastrulation stage, Msx-1 is primarily expressed in the posterior half of both normal and vitamin A-deficient embryos. However, the gene is expressed wider and stronger in the vitamin A-deficient embryos. At neurulation stages, Msx-1 is continuously expressed in the posterior region up to Hensen's node and in the edge of the neural fold in both normal and vitamin A-deficient embryos. Notably, in the vitamin A-deficient embryos, Msx-1 is expressed more strongly and is also expressed ectopically in the anterior and precardiac regions. These results provide evidence that endogenous retinoids are involved in the normal expression of Msx-1 in avian embryo and that the expression of Msx-1 is downregulated by endogenous and physiological retinoids in vivo during early avian embryogenesis.

Animals↗

PubMed2003

Positional information by Rauber's sickle and a new look at the mechanisms of primitive streak initiation in avian blastoderms.

The present experimental in vitro study suggests that a primitive streak (PS) in avian blastoderms is induced by diffusion of morphogenetic substances emanating from Rauber's sickle. Indeed, even without direct contact between a quail Rauber's sickle and the reacting upper layer (by interposition of a vitelline membrane), a PS can be induced in the isolated area centralis or antisickle region of unincubated chicken blastoderms. The so-formed PSs are localized below the vitelline membrane in the immediate neighborhood of the apposed Rauber's sickle material. This seems to indicate that Rauber's sickle organizes the formation of the avian PS according to the basic concept of "positional information." The morphogenetic substances seem to have an effect only on the formation of a PS. Each part of Rauber's sickle seems to have, point by point, the same thickening and PS-inducing effect on each corresponding part of the underlying upper layer (UL). By a mechanism of sliding over the basement membrane and fusion, this finally results in the formation of one single median PS. Our study shows that a PS can be induced in the total absence of hypoblast (sickle endoblast) or caudal marginal zone, by only the presence of Rauber's sickle material. In contrast, the differentiation of mesoblast into blood islands under the influence of Rauber's sickle and neural tissue development are impaired by the interposition of a vitelline membrane. The latter could be due to the absence of a normal interaction of Rauber's sickle-derived sickle endoblast with endophyll and/or upper layer and the absence of cranial migration of the mesoblast. Thus, earlier studies and the present study indicate the existence of a temporospatially bound cascade of gastrulation and neurulation phenomena and blood island formation in the avian blastoderm, starting from Rauber's sickle, the primary major organizer with inducing, inhibiting, and dominating potencies. The latter not only plays a role by secretion of signaling molecules, but also influences development by its cell lineages (junctional endoblast and sickle endoblast).

Animals↗

PubMed2003

Developmental transformations in a normal series of embryos of the sea lamprey Petromyzon marinus (Linnaeus).

Lamprey development is of interest to evolutionary biologists because it can inform our understanding of primitive vertebrate developmental patterns. In this study, we describe and illustrate some of the principle landmarks of organogenesis in the embryonic sea lamprey Petromyzon marinus L. at different chronological ages. We examined 63 fixed embryos spanning Piavis developmental stages 11-18+ (5-70 days postfertilization) by gross observation and histology. This period begins at late neurulation stages and ends with the formation of the larva (ammocoete). A significant difference with some previous accounts is that the anus develops not from a persistent blastopore, but by secondary canalization and proctodeum formation at the former site of the blastopore. Further, we show that the ciliated bands of the pharyngeal roof originate in the esophagus, distinguishing it from the intestine. We clarify the epithelialization of the gut, showing that the secondary gut cavity is progressively epithelialized from each end. We identify possible germ cells in the coelomic and cloacal walls. Balfour's "subnotochordal rod" is lacking in our specimens; we suggest that he may have misinterpreted the corpus adiposum. Our study is of potential value to the growing number of biologists interested in lamprey development and provides a character set that will be used : 1) in a phylogenetic study of vertebrate development, and 2) to prepare a staging series for the lamprey based on parsimony analysis.

Animals↗

PubMed2006

Early steps in neural development.

We studied early neurulation events in vitro by transplanting quail Hensen's node, central prenodal regions (before the nodus as such develops), or upper layer parts of it on the not yet definitively committed upper layer of chicken anti-sickle regions (of unincubated blastoderms), eventually associated with central blastoderm fragments. We could demonstrate by this quail-chicken chimera technique that after the appearance of a pronounced thickening of the chicken upper layer by the early inductive effect of neighboring endophyll, a floor plate forms by insertion of Hensen's node-derived quail cells into the median part of the groove. This favors, at an early stage, the floor plate "allocation" model that postulates a common origin for notochord and median floor plate cells from the vertebrate's secondary major organizer (Hensen's node in this case). A comparison is made with results obtained after transplantation of similar Hensen's nodes in isolated chicken endophyll walls or with previously obtained results after the use of the grafting procedure in the endophyll walls of whole chicken blastoderms.

Animals↗

PubMed1991

Distribution and expression of two interactive extracellular matrix proteins, cytotactin and cytotactin-binding proteoglycan, during development of Xenopus laevis. I. Embryonic development.

An immunohistochemical study of the localization of cytotactin and cytotactin-binding (CTB) proteoglycan throughout embryonic development of the anuran Xenopus laevis reveals that both appear in a restricted pattern related to specific morphogenetic events. CTB proteoglycan expression is first detected during gastrulation at the blastopore lip. Later, it is seen in the archenteron roof around groups of cells forming the notochord, somites and neural plate. Cytotactin first appears after neurulation, and is restricted to the intersomitic regions. Both molecules appear along the migratory pathways of neural crest cells in the trunk and tail. Later, cytotactin is present at sites where neural crest cells differentiate, around the aorta and in the smooth muscle coat of the gut; CTB proteoglycan is absent from these sites. In the head, cytotactin is initially restricted to the regions between cranial somites, while CTB proteoglycan is distributed throughout the cranial mesenchyme. The expression of both molecules is later associated with key events in chondrogenesis during the development of the skull. After chondrogenesis, CTB proteoglycan is distributed throughout the cartilage matrix, while cytotactin is restricted to a thin perichondrial deposit. Both molecules are expressed in developing brain. These findings are compared to studies of the chick embryo and although distinct anatomical differences exist between frog and chick, the expression of these molecules is associated with similar developmental processes in both species. These include mesoderm segmentation, neural crest cell migration and differentiation, cartilage development, and central nervous system histogenesis.

Animals↗

PubMed1991

Expression of ras-like proteins in embryonic and adult cells of Xenopus laevis.

A polyclonal antibody raised against v-Ha-ras p21 was purified and its specificity was checked on Ha-ras transformed cell lines. It was used to immunoprecipitate p21 from different Xenopus laevis cell types: brain cells, blood cells, and embryonic material. By one-dimensional Western blot analysis, we show that ras p21 is synthesized very early in oogenesis and accumulates throughout vitellogenesis. The ras p21 content, estimated to be 1.1 ng in the full-grown oocyte, remains constant during oocyte maturation and egg cleavage. Increase in the amount of ras p21 occurs at the beginning of neurulation. Two-dimensional Western blot patterns reveal the presence of multiple molecular forms of p21 in all Xenopus cell types studied. The numerous resolved polypeptides were ascribed to the expression of at least two different ras genes. Furthermore, specific charge modifications of the ras polypeptides are observed in brain, blood, and embryonic cells. During oogenesis and early embryonic development, differences in two-dimensional patterns mainly concern variations in the relative amounts of the different polypeptides. The results are discussed in relation to the well documented synthesis activities of the growing oocyte and of the early developing embryo.

Animals↗

PubMed1984

Delayed fertilization of anuran amphibian (Xenopus) eggs leads to reduced numbers of primordial germ cells.

Several media were tested for the extent to which they promoted high fertilization efficiencies in ovulated, stripped Xenopus eggs. One medium was selected for maintaining eggs in a 'delayed fertilization' (DelF) condition. DelF eggs displayed several unusual characteristics, including shift of the center of gravity, prominent sperm entrance site, and occasional polyspermy. The frequency of normal pattern formation varied according to the length of time eggs were maintained in the DelF condition. Various developmental abnormalities were observed during gastrulation, neurulation, and organogenesis. Most abnormalities appeared, however, to be related to morphogenesis of the endoderm. Primordial germ cell (PGC) development was examined in DelF eggs which displayed normal external morphological features at the swimming tadpole stage. PGC counts were usually normal in short-duration (eg, 5 hr) DelF eggs, but frequently substantially reduced or completely diminished in longer-duration (eg, 25h) tadpoles. Six spawnings were compared and shown to exhibit considerable variability in fertility, morphogenesis, and PGC development. Yolk platelet shifts and developmental parameters were examined in two additional spawnings. The subcortical cytoplasm in which the germ plasm is normally localized appeared to be disrupted in longer duration DelF eggs. That observation may account for low PGC counts in DelF tadpoles.

Animals↗

PubMed2006

Retinoids and spinal cord development.

The role that RA plays in the development and patterning of the spinal cord is discussed. The morphogenetic process of neurulation is described in which RA plays a role because in the absence of RA signaling spina bifida results. Following neural induction, RA is involved in several patterning events in the spinal cord. It is one of the posteriorizing factors along with FGFs and Wnts and as such patterns the cervical spinal cord acting via the Hoxc transcription factors. It is involved in the induction of neural differentiation via genes such as NeuroM. It plays a part in patterning the dorsoventral axis of the anterior spinal cord where it interacts with FGF, Shh, and BMPs and induces an interneuronal population of neurons called V0 and V1 and a subset of motor neurons known as LMCs. To perform these actions RA is synthesized in the adjacent paraxial mesoderm by the enzyme RALDH2 and acts in a paracrine fashion on the developing neural tube. In the final action of RA, it begins to be synthesized within the neural tube at brachial and lumbar levels in the LMCs. Later-born neurons migrate through this RALDH2-expressing region and induce differentiation in these migrating neurons, which become a subset of LMC neurons known as LMCls. Thus RA acts several times over in the development of the spinal cord and not on the cells in which it is synthesized, but in adjacent cells in a paracrine manner.

Aldehyde Oxidoreductases↗

PubMed1987

Benomyl-induced craniocerebral anomalies in fetuses of adequately nourished and protein-deprived rats.

Benomyl, a benzimidazole fungicide, produced craniocerebral and systemic malformations in fetal rats when administered by gavage in doses of 31.2, 62.5, and 125 mg/kg of maternal body weight on days 7-21 of gestation. Malformations increased in incidence and severity with increasing benomyl dosage and nearly doubled when coupled with a protein-deficient diet. Protein deficiency alone produced only decreased fetal weight. High benomyl doses produced higher percentages of fetal resorptions and late fetal deaths, and these percentages also increased with protein deficiency. A benomyl dose of 62.5 mg/kg in protein-deficiency dams, the optimal combination for a high incidence of anomalies and low fetal wastage, produced hydrocephalus in 69.4% of fetuses, meningocele in 8.2%, encephalocele in 14.3%, exencephaly in 44.9%, anencephaly in 14.3%, corpus callosum agenesis in 26.5%, periventricular necrosis in 26.5%, and periventricular cellular "overgrowth" in 55.1%. The most common combination of anomalies was hydrocephalus, exencephaly, and periventricular "overgrowth." Common systemic malformations included cleft palate, micromelia, hydroureter, and misshapen tails. No fetus was entirely normal at the highest benomyl dose. Benomyl has been shown by others to bind tubulin and inhibit the formation of microtubules that are important in neurulation, mitosis, and cell migration during early brain development. Thus, it is suggested that benomyl, coupled with a protein-deficient diet, offers a teratogenic model with a spectrum of abnormalities similar to hypervitaminosis A but with a higher yield of specific craniocerebral anomalies.

Animals↗

PubMed2002

Effects of folate supplementation on the risk of spontaneous and induced neural tube defects in Splotch mice.

BACKGROUND: Neural tube defects (NTDs) are among the most common human congenital malformations. Although clinical investigations have reported that periconceptional folic acid supplementation can reduce the occurrence of these defects, its mechanism remains unknown. Therefore, the murine mutant Splotch, which has a high incidence of spontaneous NTDs, along with the inbred strains SWV and LM/Bc, were used to investigate the relationship between folate and NTDs. METHODS: To investigate whether folates could reduce spontaneous NTDs, heterozygous Splotch dams (+/Sp) were treated with either folate or folinic acid throughout neurulation, gestational day (GD) 6.5 to 10.5. On GD 18.5 the dams were sacrificed and the fetuses examined for any neural tube defects. Subsequently, Sp/+ dams were treated with arsenic while receiving either a folate or folinic acid supplementation. Similar experiments were performed in the LM/Bc and SWV strains. RESULTS: Neither folate nor folinic acid supplements reduced the frequency of spontaneous NTDs in the embryos from Splotch heterozygote crosses. Arsenic increased the frequency of NTDs and embryonic death in the Splotch, LM/Bc and SWV litters and folinic acid failed to ameliorate the teratogenic effect of this metal. A folate supplement given to arsenic-treated dams proved to be maternally lethal in all three strains. CONCLUSIONS: Splotch embryos were not protected from either spontaneous or arsenic-induced NTDs by folinic or folic acid supplementation. Furthermore, folinic acid supplements did not reduce the incidence of arsenic-induced NTDs in either the LM/Bc or SWV litters.

Animals↗

PubMed2002

Prevention of fumonisin B1-induced neural tube defects by folic acid.

BACKGROUND: The mycotoxin fumonisin B1 (FB1) inhibits sphingolipid synthesis, blocks folate transport, and has been associated with increased incidences of cancer and neural tube defects. Results from reproductive studies in animal models in vivo and in vitro have demonstrated toxicity in some cases, but no specific terata after fumonisin exposure. No information is available about folic acid's potential to protect against this toxicity. METHODS: Neurulating mouse embryos were exposed to fumonisin or folinic acid in whole embryo culture and assessed for effects on growth and development. RESULTS: Fumonisin exposure inhibited sphingolipid synthesis, reduced growth, and caused cranial neural tube defects in a dose dependent manner. Supplemental folinic acid ameliorated the effects on growth and development, but not inhibition of sphingolipid synthesis. CONCLUSION: Fumonisin has the potential to inhibit embryonic sphingolipid synthesis and to produce embryotoxicity and neural tube defects. Folic acid can reverse some of these effects, supporting results showing that fumonisin disrupts folate receptor function.

Animals↗

PubMed2002

Effects of VLA-4 antagonists in rat whole embryo culture.

BACKGROUND: Pharmacological antagonism of VLA-4 (Very Late Antigen 4, alpha(4)beta(1) integrin) has become an attractive target for the treatment of predominantly eosinophil mediated disease states such as asthma, allergic rhinitis, multiple sclerosis, rheumatoid arthritis, diabetes, and inflammatory bowel disease. Gene knockouts of the alpha(4)-integrin subunit of VLA-4 or its cell surface ligand, VCAM-1, however, have been shown to result in embryo-lethality in homozygous null mice due to defects in chorio-allantoic or epi-myocardial fusion. Although gene knockout phenotypes are not always manifested by pharmacological antagonism, those studies suggested that VLA-4 antagonists might cause embryo-lethality or drug-induced malformations. METHODS: To test these concepts, early neurulating rat embryos were cultured by the methods of New ('78) after intra-coelomic microinjection of a VLA-4 blocking antibody or in the presence of small molecule VLA-4 antagonists. RESULTS: Defects in chorio-allantoic fusion were induced after microinjection of VLA4 blocking antibody and after continuous exposure to small molecule antagonists. In a minority of affected embryos chorio-allantoic fusion was completely blocked whereas the majority of affected embryos had only superficial chorio-allantoic fusion and the allantois was enlarged and edematous. Although the allantoic mesoderm covered the trophoblasts of the chorionic plate and contained blood vessels there was only minimal invasion of the trophoblasts by the allantoic mesoderm. The lowest observed effect level generally correlated with the IC(approximately 95), as determined in 90% plasma. DISCUSSION: Based on these data, VLA-4 antagonism might represent a significant risk to the developing embryo/fetus. In vitro exposure, however, is "constant" and does not take into account the elimination phase of these xenobiotics in vivo. Given the high concentrations required to elicit an effect, therapeutic blood levels in vivo may be several fold lower than those that affect the conceptus, depending on the tissue penetration of the compound and the route of administration. VLA-4 also exists in a range of conformations and activation states in vivo and the gene KOs and present studies do not define whether these developmental processes are dependent upon a particular activation state of VLA-4. Therefore, state-selective antagonists may have an improved embryonic safety profile. Additional studies will be required to determine potential effects of VLA-4 antagonists on embryo/fetal development in vivo.

Allantois↗

PubMed1978

Teratogenic potential of blighted potato concentrate in rabbits, hamsters, and miniature swine.

Freeze-dried preparations of potatoes infected with phytophthora infestans or Alternaria solani were fed to pregnant hamsters, rabbits, and miniature swine. No evidence was observed that these diets or the extracts of respective blights were capable of producing teratologic effects in hamsters. Lesions of neurulation and anencephaly were noticed in rabbits and swine, respectively. The need for further pharmacological and toxicological evaluation in sensitive species is emphasized.

Alternaria↗