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The thymosin beta 4 gene is strongly activated in neural tissues during early postimplantation mouse development.

We studied the temporal and spatial distribution of the mRNA encoding for thymosin beta 4 (T beta 4), a small acidic actin-sequestering peptide, during the early postimplantation mouse development. Analysis of total embryo RNA demonstrated a strong activation of T beta 4 gene after gastrulation and coincident with neurulation. In situ hybridization showed that T beta 4 mRNA was strongly expressed in the central nervous system and peripheral ganglia, paralleling the gradient of neuronal differentiation. An intense signal was also observed in intraventricular macrophages and blood vessels. The role of T beta 4 in mammalian neuroembryogenesis is discussed.

Animals↗

Creatine phosphokinase in Rana pipiens: expression in embryos, early larvae and adult tissues.

1. Creatine phosphokinase (CPK) isozymes found in Rana pipiens are similar to those found elsewhere; muscle-specific (MM-CPK), brain-specific (BB-CPK) and intermediate hybrid (MB-CPK) isozymes were identified. 2. Adult tissues exhibit CPK isozyme patterns similar to other vertebrates although variations are notable in nervous tissue, liver and kidney. 3. CPK isozyme patterns in ovarian eggs, ovulated eggs and selected stages of early development demonstrate a significant rise in CPK with neurulation. Most notable is a sharp increase in brain-specific BB-CPK. 4. MM-CPK purified from skeletal muscle is found to be similar, in properties studied, to the corresponding isozyme in other vertebrate species.

Animals↗

Hormonal and humoral influences on brain development.

This review discusses evidence for neurotransmitters as developmental signals in such ontogenic processes as neural tube formation (neurulation), germinal cell proliferation, and neuronal and glial differentiation during brain organogenesis, as well as evidence for other roles of these neurotransmitters in non-neuronal tissues of vertebrates and invertebrates. Evidence also is presented for hormonal regulation of brain development during postnatal neurogenesis and for interrelationships which may link neurotransmitters and hormones in a humoral milieu, providing a variety of control mechanisms for the central and peripheral nervous system during key phases of their development. Given the evidence for neurotransmitters and hormones as coordinating influence on neural ontogeny, it is possible that drugs, stress, and environmental influences may have the ability to perturb particular aspects of these developmental systems if present during those "critical periods" when such humoral influences are important for normal ontogeny.

Adrenal Cortex Hormones↗

Abnormal neural fold development in mouse trisomy 12 and trisomy 14. II. LM and TEM.

LM and TEM observations of embryonic tissue during the period of cranial neurulation are described in trisomic mouse embryos known to develop exencephaly, and are compared with tissue from normal mouse embryos. The earliest regularly observed differences were visible from the late presomite stage onwards, in the extracellular matrix of the cranial region. These were local defects of the basement membrane of the neural epithelium and enlarged areas of mesenchymal extracellular matrix, with associated abnormalities of mesenchymal cell distribution, cell number and cell contacts, and deficiency of alcian blue staining. Apical neuroepithelial microfilament bundles were observed at later somite-stages in trisomic embryos c.f. = compared with controls, and development of the concave neuroepithelial curvature was correspondingly retarded. Apposition of the neural folds at the forebrain/midbrain junction was never made, even though late neural fold fusion occurred in the hindbrain and ventral forebrain. At later stages (9-20 somites) the neuroepithelial cells showed pyknotic nuclei and dense intracellular inclusions. These are interpreted as secondary effects.

Animals↗

Ultrastructural analysis of chromosome translocation-induced neural tube defects.

Neural tube closure defects occurred in 33% of the embryos obtained from matings of male mice heterozygous for a reciprocal chromosome translocation (T(2;4)1Sn) with normal female CFLP mice. Light and electron microscopic observations of neuroepithelium and mesenchyme in affected embryos indicated two distinct types of anomalies occurred. The first consisted of neuroepithelial hypertrophy and neural tube closure defects. These defects most frequently affected the midbrain and hindbrain, but occasional defects of the lumbosacral neural tube were also observed. Unlike the highly organized, pseudostratified neuroepithelium in control embryos, neuroepithelial cells became stratified and formed cell islands with secondary lumina within the wall of the neural tube. The second condition was associated with a reduction in neuroepithelial thickness, considerable neuroepithelial and neural crest cell death, basal lamina alterations and premature invasion of the neuroepithelium by subjacent endothelial cells. In both cases, the cephalic mesenchyme cells, rather than their normal stellate appearance, were markedly elongated in shape and reduced in area. The number of cell-cell contacts between mesenchymal cells was also reduced significantly. These results are discussed in light of recent theories regarding the role of mesenchyme and extracellular matrix in neurulation.

Animals↗

Analysis of ATF2 gene expression during early Xenopus laevis development.

Activating transcription factor 2 (ATF2) is a cellular sequence-specific DNA-binding protein that mediates transcriptional activation by the adenovirus (Ad) E1A protein. In injected Xenopus laevis oocytes, E1A-induced transactivation requires an ATF2 recognition sequence within the responding promoters, thereby suggesting that ATF2 is present in oocytes and perhaps has a developmentally important function. As a first step in assessing this, an ATF2 cDNA was cloned and sequenced. The protein encoded by this cDNA contains 486 amino acids and is 92% identical to mammalian ATF2. ATF2 RNA and protein levels are very low in oocytes, but rise dramatically during blastulation. These high levels are maintained through gastrulation, but return to low levels during neurulation. In the blastula, ATF2 RNA and protein are virtually completely confined to cells of the animal pole. The temporal and spatial regulation of ATF2 suggests that it has an important function in early development.

Activating Transcription Factor 2↗

In vitro screening for anticonvulsant-induced teratogenesis in neural primary cultures and cell lines.

To establish inherent potential for the induction of neural tube defects the ability of selected anticonvulsant agents to interfere with cell division has been established in vitro using an antiproliferative assay in clonal cell lines and a cytotoxicity assay using primary cultures of cerebral cortex neurons at different stages of development. In order to evaluate the relative toxicities of these agents their in vitro effects were determined at 2-3 times the plasma therapeutic level. By these procedures valproate and the benzodiazepines, diazepam and clonazepam, exerted a potent antiproliferative action which could not be attributed to increased cytotoxicity. In contrast phenytoin was markedly cytotoxic but was without an antiproliferative action. This cytotoxicity was most pronounced during the periods of extensive fibre outgrowth. When compared to epidemiological and animal study data, agents which inhibited cell proliferation within twice therapeutic concentration were consistently associated with major neural tube malformations. However phenytoin, found to be positive in the cell cytotoxicity assay, is not associated with neural tube malformations but rather is primarily associated with mental retardation. Thus assessment of antiproliferative activity of anticonvulsant drugs may be one criterion for identification of teratogenic potential during neurulation.

Animals↗

High-resolution linkage map in the vicinity of the Lp locus.

Looptail (Lp) is a mutation that profoundly affects neurulation in mouse and is characterized by craniorachischisis, an open neural tube extending from the midbrain to the tail in embryos homozygous for the mutation. Lp maps to the distal portion of mouse chromosome 1, and as part of a positional cloning approach, we have generated a high-resolution linkage map of the Lp chromosomal region. For this, we have carried out extensive segregation analysis in a total of 706 backcross mice informative for Lp and derived from two crosses, (Lp/+ x SJL/J)F1 x SJL/J and (Lp/+ x SWR/J) F1 x SWR/J. In addition, 269 mice from a (Mus spretus x C57BL/6J)F1 x C57BL/6J interspecific backcross were also used to order marker loci and calculate intergene distances for this region. With these mice, a total of 28 DNA markers corresponding to either cloned genes or anonymous markers of the SSLP or SSCP-types were mapped within a 5-cM interval overlapping the Lp region, with the following locus order and interlocus distances (in cM): centromere--D1Mit110/Atp1 beta 1/Cd3 zeta/Cd3 eta/D1Mit145-D1Hun14/D1Mit15- D1Mit111/D1Mit112-D1Mit114-D1Mit148/D1Mit205+ ++/D1Mit36/D1Mit146/D1Mit147/D1Mit270 / D1Hun13-Fcgr2-Mpp-Apoa2/Fcer1 gamma-Lp-D1Mit149/Spna1/Fcer1 alpha-Eph1-Hlix1/D1Mit62. These studies have allowed the delineation of a maximum genetic interval for Lp of 0.5 cM, a size amenable to physical mapping techniques.

Animals↗

Effects of altered expression of the neural cell adhesion molecule, N-CAM, on early neural development in Xenopus embryos.

The neural cell adhesion molecule, N-CAM, is known to be expressed very early in the development of the vertebrate nervous system. In frog embryos, N-CAM expression increases dramatically in ectoderm coincident with the formation of the neural plate and tube, suggesting that morphogenesis of the early nervous system is controlled in part by differential expression of N-CAM. This model was tested by introducing synthetic N-CAM transcripts into Xenopus embryos so that N-CAM was indiscriminately expressed at high levels on the surface of both induced and noninduced ectodermal cells throughout gastrulation and neurulation. Analysis of these embryos shows that high levels of N-CAM misexpression do not effect neural tube formation even though ectopic expression of N-CAM in epidermis and somitic mesoderm caused specific defects in the structure of these tissues. By showing that the properly regulated expression of N-CAM is not essential for neural tube formation, these results provide compelling evidence that N-CAM on its own does not act as a regulatory molecule during early neural development.

Animals↗

Neuronal determination without cell division in Xenopus embryos.

Cell division in the Xenopus CNS was blocked by incubating embryos in a mixture of the DNA synthesis inhibitors hydroxyurea and aphidicolin. Surprisingly, embryos treated at the beginning of gastrulation proceeded normally through neurulation, neural tube closure, and CNS subdivision. Thus, cell division is not critical for neural induction or early morphogenetic events in the CNS. Neuroblasts in treated embryos differentiated into neurons of many classes, indicating that cellular determination in the CNS can be dissociated from lineage and birth date. Axonal tracts and embryonic reflexes also developed. The remarkable amount of normal CNS development that occurs in these animals may be explained by a series of sequential inductions that are largely independent of cell proliferation.

Animals↗

Delayed formation of the floor plate after ablation of the avian notochord.

We have examined the long-term effects of notochord ablation at chick stages 9-10 on formation of the floor plate and motor neurons. Although missing or reduced 2 days postablation, the floor plate and motor neurons were morphologically normal by 4 postoperative days. When isolated whole or ventral, but not lateral, neural plate fragments from stage 9 embryos were cultured for 4 days in collagen gels, floor plate and neural markers were observed. Our results suggest that floor plate and motor neurons can form in a delayed fashion in vivo after notochord ablation and in vitro from isolated neural plates. This suggests that either there is an early induction of floor plate by the chordamesoderm of Hensen's node, or only limited interactions between the neural plate and notochord immediately after neurulation are required for floor plate determination.

Animals↗

Neural expression of a sodium channel gene requires cell-specific interactions.

In the protochordate Halocynthia roretzi, voltage-activated sodium current undergoes a change in kinetics within 48 hr of fertilization. Molecular cloning and microinjection of antisense DNA into single cells suggest that the kinetic changes are due to the increased expression of a putative neural-specific sodium channel gene, TuNa I. TuNa I gene transcription is first induced in late stage gastrulae, preceding the appearance of the rapidly inactivating sodium current unique to neural cells. In cleavage-arrested and intact embryos, cell interactions between specific animal and vegetal blastomeres are required for induction of TuNa I gene expression. Our results implicate cell contact, prior to neurulation, as a mechanism for selectively activating the TuNa I gene expressed in cells of the neural lineage.

Animals↗

Congenital anomalies in relatives of schizophrenic probands may indicate a retinoid pathology.

Retinoic acid, the morphogenic derivative of vitamin A, has been shown to alter patterns of neurulation and to regulate the expression of many genes involved in central nervous system development. Retinoid toxicity can result in craniofacial, limb, digit, heart and urogenital abnormalities. Hydrocephalus, due to increased ventricular size and/or decreased size of the hind- or forebrain, occurs frequently. Comparison of the frequency and type of congenital anomalies in extended pedigrees of 12 Ashkenazi probands with schizophrenia and seven normal Ashkenazi control probands indicates that relatives of the schizophrenic probands present a gamut of both minor and major congenital anomalies similar to, but less severe than, those caused by retinoid excess or deficiency, and at a frequency significantly greater than in control pedigrees. Within schizophrenic pedigrees, those diagnosed with schizophrenia spectrum illnesses are more likely to present such anomalies than are non-spectrum members. Retinoic acid receptors are present in all parts of the cranial region and delivery of retinoids is exquisitely controlled throughout embryonic and fetal development. Alterations in the functioning of the retinoid cascade may have profound implications for neurodevelopmental disorders like schizophrenia.

Abnormalities, Multiple↗

Activin A induced expression of a fork head related gene in posterior chordamesoderm (notochord) of Xenopus laevis embryos.

A gene family encoding the fork head/HNF-3 domain has been identified in the South African clawed frog, Xenopus laevis. Screening of genomic DNA and gastrula stage derived cDNA libraries with a DNA fragment encoding the Drosophila fork head domain led to the isolation of a number of different clones encoding this motif. While one of the Xenopus fork head sequences, XFD-3, represents the Xenopus counterpart to rat HNF-3 beta, all other sequences encode novel types of fork head related proteins. Here we report on XFD-1, a DNA binding protein which can bind to the HNF-3 alpha target sequence. Analysis of temporal and spatial expression revealed that the gene is activated at blastula stage and that transcripts are localized in a rather thin stripe of cells invaginating the dorsal blastopore lip (organizer) during gastrulation. XFD-1 mRNA is localized within the notochord and, by the end of neurulation, is no longer detectable. In the animal cap assay the gene is activated by incubation with the vegetalizing factor (activin A) but not with bFGF.

Activins↗

Dynamic expression of the murine Achaete-Scute homologue Mash-1 in the developing nervous system.

The Drosophila Achaete-Scute Complex genes encode transcriptional regulators belonging to the basic-helix-loop-helix family which control early steps of development of the central and peripheral nervous systems. We have isolated two mouse homologues of Achaete-Scute Complex genes, Mash-1 and Mash-2, by using the conservation of the basic-helix-loop-helix domain in this family. In this article, we analyse the expression of Mash-1 from its onset during neurulation to adult stages by RNA in situ hybridization on whole mounts and sections. As was observed for the rat Mash-1 protein, mouse Mash-1 RNA expression is restricted to cells of the developing central and peripheral nervous systems. We have observed three successive phases in the distribution of Mash-1 transcripts in the developing central nervous system. Initially, between embryonic day 8.5 and 10.5, Mash-1 transcripts are found in restricted domains in the neuroepithelium of the midbrain and ventral forebrain, as well as in the spinal cord. Between embryonic day 10.5 and 12.5, Mash-1 expression pattern changes from a restricted to a widespread one. Mash-1 transcripts are then found at variable levels in the ventricular zone in all regions of the brain. From embryonic day 12.5 to post-natal stages, Mash-1 is also expressed in cells outside of the ventricular zone throughout the brain. In addition, Mash-1 is expressed during development of the olfactory epithelium and neural retina. Overall, its expression pattern suggest that Mash-1 plays a role at early stages of development of specific neural lineages in most regions of the central nervous system and of several lineages in the peripheral nervous system. We have also compared the expression of Mash-1 and mouse Notch because their Drosophila homologues have been shown to interact genetically. The two genes show very similar expression patterns, both spatially and temporally, in the early developing brain and in the retina, suggesting that both genes may participate in the development of the same neural lineages.

Amino Acid Sequence↗

The c-ets-1 proto-oncogenes in Xenopus laevis: expression during oogenesis and embryogenesis.

We previously reported the cloning and sequencing of two cDNAs derived from the Xenopus laevis ets-1 gene (Stiegler et al., 1990). The Xl-ets-1a cDNA encodes a polypeptide highly homologous to known ets-1 proteins. The 3'-UTR contains two AATAAA polyadenylation signals together with three copies of the TTTTTAT sequence thought to confer a maturation-specific polyadenylation and implicated in the deadenylation of dormant mRNAs. Several transcripts with maternal characteristics were detected in oogenesis and early embryogenesis. A marked augmentation of the major transcript in the poly(A)+ fraction was detected at fertilization. Ets-1 transcripts were observed at constant levels during the cleavage stages but decreased abruptly at gastrulation, to reappear from neurulation to late embryogenesis. The possible contribution of 3'-UTR sequence elements to this behavior is discussed.

Adenosine↗

Stimulation of circus movement by activin, bFGF and TGF-beta 2 in isolated animal cap cells of Xenopus laevis.

Lobopodium is a hyaline cytoplasmic protrusion which rotates circumferencially around a cell. This movement is called circus movement, which is seen in dissociated cells of amphibian embryos. Relative abundance of the lobopodia-forming cells changes temporally and spatially within Xenopus embryos, reflecting stage-dependent difference of morphogenetic movements. The lobopodia-forming activity of dissociated animal cap cells was stimulated strongly by activin and bFGF, and weakly by TGF-beta 2. In addition, activin A was found to stimulate cellular attachment to the substratum when the cultivation lasted long. Thus, mesoderm-inducing growth factors stimulate lobopodia formation and cellular movements which may be necessary for gastrulation and neurulation in Xenopus early embryos.

Activins↗

Integrin alpha 5 during early development of Xenopus laevis.

The full length sequence of the Xenopus integrin alpha 5 subunit is reported. Analysis of cloned cDNA fragments reveals that alternative polyadenylation of alpha 5 mRNA occurs in the embryo. Furthermore, a variant form of the alpha 5 mRNA is expressed which encodes an integrin alpha 5 subunit with a truncated cytoplasmic domain. Integrin alpha 5 mRNA and protein are expressed in oocytes, eggs and throughout development. Spatial expression of alpha 5 mRNAs is first detected by whole mount in situ hybridization in presumptive neural crest cells and in the somitic mesoderm from the midgastrula stage onwards. In contrast, the alpha 5 protein is present on newly formed plasma membranes beginning at first cleavage. During neurulation, the integrin alpha 5 subunit disappears from the outer layer of the ectoderm, the notochord and the neural tube and accumulates in the sensorial layer of the ectoderm, the somites and the neural crest cells. These results provide evidence for the position specific regulation of alpha subunit expression in early vertebrate embryos.

Amino Acid Sequence↗