This competitive renewal grant application of a Javits Award proposes to: i) continue exploring the typology and synaptic connections of the small inhibitory neurons of the cerebellar nuclei to clarify their involvement in the circuitry of the inferior olivary and cerebellar systems; and ii) clarify further the development of morphological and chemical phenotypes of cerebellar neurons in vivo and in vitro, to provide the basis for a better understanding of cell class specific gene expression and of the mechanisms of brain development and plasticity. The project utilizes cerebellar tissues from: a) normal adult animals; b) human brains obtained at autopsy from persons deceased after non-neurologic processes; c) embryonic, mutant and transgenic mice. Specifically, we propose to continue studying: 1) The morphological and chemical phenotypes of small neurons in the cerebellar nuclei of various mammals, including humans; 2) The synaptic connections of these neurons in experimental animals; 3) The changes in these neurons and their connections in murine mutants that have lost Purkinje neurons; 4) The pattern of the L7 gene expression at early stages of cerebellar morphogenesis in normal and transgenic mice, as a means to reveal developmental maps; and 5) The development and experimental manipulation of distinct neuronal populations in dissociated cerebellar cell cultures derived from normal and transgenic mice, with emphasis on the morphological and chemical phenotypes of Purkinje cells and the deep nuclei cells. A battery of classical and modern neuroanatomical and neurocytological procedures will be used. Normal, experimental, mutant and transgenic animals will be studied by quantitative electron microscopy, immuno-light and -electron microscopy, iontophoretic and pressure driven microinjections of molecules to trace neural pathways, classical Golgi impregnations and Golgi/EM impregnations combined with anterograde degeneration. Human autopsy material will be explored by light microscopic immunocytochemistry. Dissociated cerebellar cell cultures will be analyzed by electron microscopy and immunocytochemistry. Mutant mice and other animals will derive from commercial and institutional colonies; transgenic mice will be provided by collaborating molecular genetics laboratories, and human autopsy material will be obtained from approved human tissue banks. The proposed studies will help understand the participation of inhibitory cerebellar neurons in sensorimotor processing and clarify the ontogenetic and evolutionary development and plasticity of the cerebellum. As this center is an amply utilized model system for studies on development and neurospecific gene expression, this research will contribute to advancements not only in the area of a particular neural system, but also in cellular neurobiology in general.
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