We propose to address four crucial questions about retinal development: (1) what is the embryonic origin of rod photoreceptor progenitor cells?; (2) What is the cellular lineage of rod progenitor cells?; (3) What controls the rate of proliferation of rod specific progenitor cells?; (4) What is the time course of differentiation of newly generated rod photoreceptors? Since retinal histogenesis follows a similar pattern throughout vertebrate pylogeny, we propose to continue our experiments using a teleost fish because of the unique experimental advantages it offers. The fish eye grows throughout life from a germinal zone at the margin of the eye, providing a spatial recapitulation of embryonic retinal neurogenesis. In addition, we can study embryos directly where necessary since we can produce animals of known developmental stage with in vitro fertilization. We answer these questions by using a variety of cellular and molecular techniques. We will use incorporation of 3H-thymidine to label dividing cells in the experiments focussed on the origin of rod photoreceptor progenitor cells. We will use the intracellular injection of fluorescent dye to trace the lineage of individual cells. We will use in situ hybridization of opsin mRNA to trace the time course of differentiation following cell division. We know that the teleost retina has two kinds of stem cells which differ greatly in their developmental strategies. The marginal zone consists of cells in close physical proximity, derived from the cells which give rise to the original retina and they produce progeny arranged in an orderly array. In contrast, the rod progenitor cells are isolated in the retina, arise from an unknown origin and have a single type of progeny. Understanding the basis for having these two strategies for the assembly of the retina will contribute to our understanding of fundamental principles of retinal development.
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