): Defects of the nervous system result from disruptions of the normal mechanisms that control neural development. Thus understanding the basis for inherited neurological disorders requires a knowledge of the embryonic molecular and genetic processes that control pattern formation in neural precursors. These include the tissue interactions that establish the primary axes of the neural tube, its differentiation into different neuronal types, and their interconnections to form a functional neural network. This proposal examines these issues using the genetics and embryological advantages of the zebrafish as a model. Zebrafish embryos form a simple nervous system, where homologies with regions of the human brain and spinal cord are recognizable and many of the genes that pattern these regions have been conserved between fish and humans. It is thought that a major component of anterior-posterior patterning of the nervous system is retinoic acid (RA), a signal that directs patterns of neuronal development along the body axis. Exogenous RA truncates anterior development and has been shown to cause severe neural and craniofacial defects in humans. Experiments are proposed to dissect the role of this signal using a zebrafish mutant in retinaldehyde dehydrogenase (RALDH2), an enzyme that synthesizes RA in the embryo, as well as specific antagonists or dominant negative forms of RA receptors (RARs).
Aim 1 is to characterize defects in gene expression in the RALDH2 mutant and compare them with the effects of RAR disruption, to define the extent of the requirement for RA and its possible target genes in the nervous system.
Aim 2 is to analyze the simple pattern of zebrafish primary neurons, combined with real time imaging of neural development in living embryos using a transgenic approach in which developing neurons express a fluorescent marker, both in RALDH2 mutants and in embryos with disrupted RAR signalling. This will elucidate the overall extent of the influence of RA along the body axis and its roles in the behaviors and fates of virtually every neuron in the system. The optical clarity and availability of transgenic zebrafish make it uniquely suited for this study.
Aim 3 focuses on a particular tissue interaction that requires RA, between the mesoderm and developing neural tube, by using cell transplantation in RALDH2 mutants to get to the biochemical basis for neural patterning.
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