Role of actin cytoskeleton in the axonal growthcone during brain development

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During the neuronal network formation, the inhibition of axonal growthcone morphogenesis and its function make it impossible for the brain function to develop normally. In this study, in order to elucidate the regulatory mechanism of actin cytoskeleton, which is directly involved in the morphogenesis and function of axonal growthcones, we focused in the actin filament and analyzed the change in the protein network of actin-binding proteins which regulate the physicochemical and biochemical characters using primary cultured hippocampal neurons. We first analyzed the distribution of drebrin in the axonal growthcone of cultured hippocampal neurons at various developmental stages using immunocytochemistry. In the neuron of stage 2, drebrin was localized at transitional area of growthcones. The immunocytochemistry also showed that Neurabin 1 showed similar localization of in the growthcone to drebrin. We further analyzed localization of various actin-related proteins in the growthcones at stage 2. Microtubules did not invade into the transitional area. In order to discriminate drebrin E from drebrin A, we developed drebrin A-specific antibody. This antibody clearly showed that growthcone only has drebrin E. This antibody further showed that migrating neurons in the adult brain only express drebrin E. We developed the new identification method of migrating neurons in the adult brain. Using this method, we discovered migrating neurons in the piriform cortex in adult rats. We finally showed that drebrin knockdown by RNAi resulted the inhibition of axonal growth.