Inhibition of in vitro enteric neuronal development by endothelin-3: mediation by endothelin B receptors.
The terminal colon is aganglionic in mice lacking endothelin-3 or its receptor, endothelin B. To analyze the effects of endothelin-3/endothelin B on the differentiation of enteric neurons, E11-13 mouse gut was dissociated, and positive and negative immunoselection with antibodies to p75(NTR )were used to isolate neural crest- and non-crest-derived cells. mRNA encoding endothelin B was present in both the crest-and non-crest-derived cells, but that encoding preproendothelin-3 was detected only in the non-crest-derived population. The crest- and non-crest-derived cells were exposed in vitro to endothelin-3, IRL 1620 (an endothelin B agonist), and/or BQ 788 (an endothelin B antagonist). Neurons and glia developed only in cultures of crest-derived cells, and did so even when endothelin-3 was absent and BQ 788 was present. Endothelin-3 inhibited neuronal development, an effect that was mimicked by IRL 1620 and blocked by BQ 788. Endothelin-3 failed to stimulate the incorporation of [3H]thymidine or bromodeoxyuridine. Smooth muscle development in non-crest-derived cell cultures was promoted by endothelin-3 and inhibited by BQ 788. In contrast, transcription of laminin alpha1, a smooth muscle-derived promoter of neuronal development, was inhibited by endothelin-3, but promoted by BQ 788. Neurons did not develop in explants of the terminal bowel of E12 ls/ls (endothelin-3-deficient) mice, but could be induced to do so by endothelin-3 if a source of neural precursors was present. We suggest that endothelin-3/endothelin B normally prevents the premature differentiation of crest-derived precursors migrating to and within the fetal bowel, enabling the precursor population to persist long enough to finish colonizing the bowel. (+info)
A molecular pathway revealing a genetic basis for human cardiac and craniofacial defects.
Microdeletions of chromosome 22q11 are the most common genetic defects associated with cardiac and craniofacial anomalies in humans. A screen for mouse genes dependent on dHAND, a transcription factor implicated in neural crest development, identified Ufd1, which maps to human 22q11 and encodes a protein involved in degradation of ubiquitinated proteins. Mouse Ufd1 was specifically expressed in most tissues affected in patients with 22q11 deletion syndrome. The human UFD1L gene was deleted in all 182 patients studied with 22q11 deletion, and a smaller deletion of approximately 20 kilobases that removed exons 1 to 3 of UFD1L was found in one individual with features typical of 22q11 deletion syndrome. These data suggest that UFD1L haploinsufficiency contributes to the congenital heart and craniofacial defects seen in 22q11 deletion. (+info)
A subpopulation of apoptosis-prone cardiac neural crest cells targets to the venous pole: multiple functions in heart development?
A well-described population of cardiac neural crest (NC) cells migrates toward the arterial pole of the embryonic heart and differentiates into various cell types, including smooth muscle cells of the pharyngeal arch arteries (but not the coronary arteries), cardiac ganglionic cells, and mesenchymal cells of the aortopulmonary septum. Using a replication-incompetent retrovirus containing the reporter gene LacZ, administered to the migratory neural crest of chicken embryos, we demonstrated another population of cardiac neural crest cells that employs the venous pole as entrance to the heart. On the basis of our present data we cannot exclude the possibility that precursors of these cells might not only originate from the dorsal part of the posterior rhombencephalon, but also from the ventral part. These NC cells migrate to locations surrounding the prospective conduction system as well as to the atrioventricular (AV) cushions. Concerning the prospective conduction system, the tagged neural crest cells can be found in regions where the atrioventricular node area, the retroaortic root bundle, the bundle of His, the left and right bundle branches, and the right atrioventricular ring bundle are positioned. The last area connects the posteriorly located AV node area with the retroaortic root bundle, which receives its neural crest cells through the arterial pole in concert with the cells giving rise to the aortopulmonary septum. The NC cells most probably do not form the conduction system proper, as they enter an apoptotic pathway as determined by concomitant TUNEL detection. It is possible that the NC cells in the heart become anoikic and, as a consequence, fail to differentiate further and merely die. However, because of the perfect timing of the arrival of crest cells, their apoptosis, and a change in electrophysiological behavior of the heart, we postulate that neural crest cells play a role in the last phase of differentiation of the cardiac conduction system. Alternatively, the separation of the central conduction system from the surrounding working myocardium is mediated by apoptotic neural crest cells. As for the presence of NC cells in both the outflow tract and the AV cushions, followed by apoptosis, a function is assigned in the muscularization of both areas, resulting in proper septation of the outflow tract and of the AV region. Failure of normal neural crest development may not only play a role in cardiac outflow tract anomalies but also in inflow tract abnormalities, such as atrioventricular septal defects. (+info)
Development of cephalic neural crest cells in embryos of Lampetra japonica, with special reference to the evolution of the jaw.
Neural crest cells contribute extensively to vertebrate head morphogenesis and their origin is an important question to address in understanding the evolution of the craniate head. The distribution pattern of cephalic crest cells was examined in embryos of one of the living agnathan vertebrates, Lampetra japonica. The initial appearance of putative crest cells was observed on the dorsal aspect of the neural rod at stage 20.5 and ventral expansion of these cells was first seen at the level of rostral somites. As in gnathostomes, cephalic crest cells migrate beneath the surface ectoderm and form three major cell populations, each being separated at the levels of rhombomeres (r) 3 and r5. The neural crest seems initially to be produced at all neuraxial levels except for the rostral-most area, and cephalic crest cells are secondarily excluded from levels r3 and r5. Such a pattern of crest cell distribution prefigures the morphology of the cranial nerve anlage. The second or middle crest cell population passes medial to the otocyst, implying that the otocyst does not serve as a barrier to separate the crest cell populations. The three cephalic crest cell populations fill the pharyngeal arch ventrally, covering the pharyngeal mesoderm laterally with the rostral-most population covering the premandibular region and mandibular arch. The third cell population is equivalent to the circumpharyngeal crest cells in the chick, and its influx into the pharyngeal region precedes the formation of postotic pharyngeal arches. Focal injection of DiI revealed the existence of an anteroposterior organization in the neural crest at the neurular stage, destined for each pharyngeal region. The crest cells derived from the posterior midbrain that express the LjOtxA gene, the Otx2 cognate, were shown to migrate into the mandibular arch, a pattern which is identical to gnathostome embryos. It was concluded that the head region of the lamprey embryo shares a common set of morphological characters with gnathostome embryos and that the morphological deviation of the mandibular arch between the gnathostomes and the lamprey is not based on the early embryonic patterning. (+info)
Regulation of Hoxa2 in cranial neural crest cells involves members of the AP-2 family.
Hoxa2 is expressed in cranial neural crest cells that migrate into the second branchial arch and is essential for proper patterning of neural-crest-derived structures in this region. We have used transgenic analysis to begin to address the regulatory mechanisms which underlie neural-crest-specific expression of Hoxa2. By performing a deletion analysis on an enhancer from the Hoxa2 gene that is capable of mediating expression in neural crest cells in a manner similar to the endogenous gene, we demonstrated that multiple cis-acting elements are required for neural-crest-specific activity. One of these elements consists of a sequence that binds to the three transcription factor AP-2 family members. Mutation or deletion of this site in the Hoxa2 enhancer abrogates reporter expression in cranial neural crest cells but not in the hindbrain. In both cell culture co-transfection assays and transgenic embryos AP-2 family members are able to trans-activate reporter expression, showing that this enhancer functions as an AP-2-responsive element in vivo. Reporter expression is not abolished in an AP-2(alpha) null mutant embryos, suggesting redundancy with other AP-2 family members for activation of the Hoxa2 enhancer. Other cis-elements identified in this study critical for neural-crest-specific expression include an element that influences levels of expression and a conserved sequence, which when multimerized directs expression in a broad subset of neural crest cells. These elements work together to co-ordinate and restrict neural crest expression to the second branchial arch and more posterior regions. Our findings have identified the cis-components that allow Hoxa2 to be regulated independently in rhombomeres and cranial neural crest cells. (+info)
Early specification of sensory neuron fate revealed by expression and function of neurogenins in the chick embryo.
The generation of sensory and autonomic neurons from the neural crest requires the functions of two classes of basic helix-loop-helix (bHLH) transcription factors, the Neurogenins (NGNs) and MASH-1, respectively (Fode, C., Gradwohl, G., Morin, X., Dierich, A., LeMeur, M., Goridis, C. and Guillemot, F. (1998) Neuron 20, 483-494; Guillemot, F., Lo, L.-C., Johnson, J. E., Auerbach, A., Anderson, D. J. and Joyner, A. L. (1993) Cell 75, 463-476; Ma, Q., Chen, Z. F., Barrantes, I. B., de la Pompa, J. L. and Anderson, D. J. (1998 Neuron 20, 469-482). We have cloned two chick NGNs and found that they are expressed in a subset of neural crest cells early in their migration. Ectopic expression of the NGNs in vivo biases migrating neural crest cells to localize in the sensory ganglia, and induces the expression of sensory neuron-appropriate markers in non-sensory crest derivatives. Surprisingly, the NGNs can also induce the expression of multiple pan-neuronal and sensory-specific markers in the dermomyotome, a mesodermal derivative. Taken together, these data suggest that a subset of neural crest cells may already be specified for a sensory neuron fate early in migration, as a consequence of NGN expression. (+info)
Early embryonic lethality in Bmp5;Bmp7 double mutant mice suggests functional redundancy within the 60A subgroup.
Members of the BMP family of signaling molecules display a high conservation of structure and function, and multiple BMPs are often coexpressed in a variety of tissues during development. Moreover, distinct BMP ligands are capable of activating common pathways. Here we describe the coexpression of two members of the 60A subfamily of BMPs, Bmp5 and Bmp7, at a number of different sites in the embryo from gastrulation onwards. Previous studies demonstrate that loss of either Bmp5 or Bmp7 has negligible effects on development, suggesting these molecules functionally compensate for each other at early stages of embryonic development. Here we show this is indeed the case. Thus we find that Bmp5;Bmp7 double mutants die at 10.5 dpc and display striking defects primarily affecting the tissues where these factors are coexpressed. The present analysis also uncovers novel roles for BMP signaling during the development of the allantois, heart, branchial arches, somites and forebrain. Bmp5 and Bmp7 do not appear to be involved in establishing pattern in these tissues, but are instead necessary for the proliferation and maintenance of specific cell populations. These findings are discussed with respect to potential mechanisms underlying cooperative signaling by multiple members of the TGF-beta superfamily. (+info)
Prospective identification, isolation by flow cytometry, and in vivo self-renewal of multipotent mammalian neural crest stem cells.
Multipotent and self-renewing neural stem cells have been isolated in culture, but equivalent cells have not yet been prospectively identified in neural tissue. Using cell surface markers and flow cytometry, we have isolated neural crest stem cells (NCSCs) from mammalian fetal peripheral nerve. These cells are phenotypically and functionally indistinguishable from NCSCs previously isolated by culturing embryonic neural tube explants. Moreover, in vivo BrdU labeling indicates that these stem cells self-renew in vivo. NCSCs freshly isolated from nerve tissue can be directly transplanted in vivo, where they generate both neurons and glia. These data indicate that neural stem cells persist in peripheral nerve into late gestation by undergoing self-renewal. Such persistence may explain the origins of some PNS tumors in humans. (+info)