The retina is the most amenable part of the central nervous system (CNS) for studies of development. The order of appearance of different cell types and the formation of their connections have been well described using classical techniques. For example, we know that there is an order of birth of different retinal cell types that is generally conserved among all vertebrate species. We now wish to study the mechanisms that guide development of the retina. As a first step in this pursuit, we propose to define molecules expressed by the progenitor cells of the developing retina. We then wish to use these markers to assess whether retinal progenitor cells change during development. This information will help to resolve whether progenitors are totipotent throughout development and different cell types are generated at different times due to environmental influences alone, or whether progenitors change over time, perhaps becoming restricted as development proceeds. These possibilities have been suggested and discussed for many types of CNS progenitors, but we have little definitive evidence regarding this issue. Several parallel strategies employing immunological approaches, nucleic acid-based approaches, and combinations of these 2 approaches will be used. In one strategy, we will search for temporally regulated expression of classes of surface receptors known to play a role in development. In another, we will use the immune system to make monoclonal and polyclonal antisera to define markers expressed on the surface of progenitors. Expression of these markers in progenitors will be studied to determine if they exhibit temporal regulation in order to see if they reveal changes in retinal progenitors. These markers will be useful for other applications as well. They provide a tool for manipulation of retinal progenitors (e.g. killing of subpopulations of cells using complement-mediated lysis, selection of cells bearing surface receptors using fluorescence activated cell sorting) which will be useful for in vitro studies of progenitors aimed towards an understanding of cell fate determination mechanisms. The classes of genes chosen also lend themselves to studies of function through the creation of dominant negative alleles. Such alleles will allow studies of function via gene transfer into the retina using retrovirus vectors. These studies also can contribute to our understanding of diseases and potentially inform our choices regarding therapy. For example, in diseases where certain cell types die, such as rods and/or cones, replacement of the dying cells with healthy progenitors that can differentiate and replace the dying cells has been proposed. if certain types of progenitors are restricted to making rode or cones, these progenitors need to be identified and methods for purifying them must be devised. Our studies will provide this information.
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