Subcellular Pathways of Internalized Neurotrophics

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Neurotrophic factors are messengers in the communication between neurons. They regulate neuronal differentiation and function. Internalization, intracellular transport, signal transduction and eventual degradation are essential steps in trophic signaling. Recent studies have shown that internalized neurotrophins can be sorted into either degradative or recycling pathways. The application's central aim is to elucidate how internalized neurotrophins are sorted into distinct intracellular pathways. This research will be done primarily at the Nencki Institute in Poland as an extension of NIH grant # RO 1 EY 12841. The proposed studies will utilize the developing visual system of chick embryos as a unique invivo model system which allows the introduction of radiolabeled trophic factors into a compartment and the quantification of intracellular pathways. Experiments will focus on the comparison of different subcellular pathways after internalization of neurotrophins by dendrites (retinal ganglion cells) and after internalization exclusively by axon terminals (isthmo-optic neurons). These experiments will show how the route of uptake (dendritic vs. axonal) influences the subsequent signaling and degradation pathways. Autoradiography at the ultrastructural level will be used to identify the subcellular distribution of neurotrophins. Distribution profiles will be compared between neurotrophins. Receptor binding of invivo- transported neurotrophins will be determined by crosslinking and immunoprecipitation with receptor-specific antibodies. The organelles in which internalized neurotrophins accumulate and their pathways and subcellular destinations will be compared and experimentally manipulated by competition with heterologous neurotrophins and inactivation of tyrosine kinase receptors. The combination of molecular, pharmacological and ultrastructural approaches will allow us to answer questions about trafficking of neurotrophins which are crucial to an understanding of how these factors may regulate events as diverse as neuronal survival and synaptic plasticity. Neurodegenerative diseases have been related to deficits in trophic support. Knowledge about the normal trafficking, sorting and recycling of neurotrophic factors will help us to understand pathologic conditions and how exogenous neurotrophins may be used as therapeutic agents.