Specific Aims A growth cone is the motile tip of agrowing axonor dendrite. Properassemblyof neural circuits depends on navigation by growth cones during development. Fivebehaviors of growth conesarecritical in forming neural circuits: migration, turning, branching, retraction, and transformation to synaptogenesis. A growth conedetectsenvironmental cues and responds by regulating these five behaviors. In addition to improving our understanding of howthe brain develops, webelieve elucidating the mechanismsof growth cone guidance will help in designing strategies to promoterepair andregeneration of damaged nervous tissue. We will pursue two aims to test our main hypothesis that neurotrophins modulatethe responsesof growth conesto other guidance cues viacytoplasmic signaling pathways that regulate thedynamics of cytoskeletal components: 1. Elucidate the role of neurotrophins in regulating growth coneresponsesto multiple guidance cues. Wewilltest the hypothesis that the p75neurotrophin receptor is involved in neurotrophin regulation of growth cone behavior. Wewill test the hypothesis that neurotrophin regulation of growth cone behaviors involves binding to p75,activation of protein kinase A (PKA)andinhibition of p75-mediated activation of RhoA GTPase. 2. Elucidate the cytoskeletal changes and interactions that accompanygrowthcone collapse and retraction. Wewilltestthe hypothesis that myosin II contractility drives growth conecollapse, retraction and cytoskeletalrearrangements. These studies willinvolve tissue culture of sensory,retinal andciliary neurons from chick embryosandsensory and retinal neurons from mice with mutant p75and trkB genes.Explants of neural tissues will bepreparedand growth conebehaviors willbe analyzed by video microscopy and quantitative fluorescence microscopy. Responses of growth conesto severalmolecular guidance cues will bedetermined, and the activities of PKAand Rhofamily GTPases will bemeasured. Cytoskeletalorganization willbe analyzed in growth cones treatedwith neurotrophins andwith negativeguidance cues.
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