OLIGODENDROCYTE ONTOGENY AND DIFFERENTIATION

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In the central nervous system myelin is produced by the oligodendrocytes. The extent of myelination ultimately depends on the allocation and proliferation of oligodendrocyte progenitor cells and on the morphological and biochemical differentiation of mature oligodendrocytes. We propose to study these two phenomena in primary cultureof newborn mouse brain and to analyze the effect thereon of several neurological mutations of the mouse - jp, msd, qk, shi, mld - that are characterized by hypomyelination. The population dynamics of oligodendrocyte ontogeny will be analyzed using: limiting dilution analysis of progenitor cells, 3H-thymidine labeling of proliferative cells and immunofluorescent counting of mature cells. The biochemical parameters of oligodendrocytes differentiation will be analyzed by determining the developmental profile of accumulation of various myelin related lipid and protein components. This will be performed in whole culture, in clone-resolved microwells and in individual oligodendrocytes using a variety of immunological techniques to identify and quantitate components. We will also analyze the regulation of myelin basic protein gene expression in normal and mutant cultures using both Northern hybridization with a recombinant cDNA probe and in vitro translation and immunoprecipitation to quantitate MBP-specific mRNA, and using both immunoblotting and immunofluorescence with polyclonal and monoclonal antibodies to quantitate MBP-related polypeptides. The results of this analysis will provide a detailed description of oligodendrocyte ontogeny and differentiation at the cellular and molecular levels in primary culture and will reveal how this process is regulated by genetic and epigenetic factors. This will provide important insight into the process of brain development and the effect thereon of birth defects, and will also help to elucidate the pathoetiology of demyelinating diseases such as multiple sclerosis and the factors controlling remyelination in the central nervous system.