A growth cone is the motile tip of a growing axon or dendrite. Proper assembly of neural circuits depends on navigation by growth cones during development. Five behaviors of growth cones are critical in forming neural circuits: migration, turning, branching, retraction, and transformation to synaptogenesis. A growth cone detects environmental cues and responds by regulating these five behaviors. In addition to improving our understanding of how the brain develops, we believe elucidating the mechanisms of growth cone guidance will help in designing strategies to promote repair and regeneration of damaged nervous tissue. We will pursue two aims to test our main hypothesis that neurotrophins modulate the responses of growth cones to other guidance cues via cytoplasmic signaling pathways that regulate the dynamics of cytoskeletal components: (1) Elucidate the role of neurotrophins in regulating growth cone responses to multiple guidance cues. We will test the hypothesis that the p75 neurotrophin receptor is involved in neurotrophin regulation of growth cone behavior. We will test the hypothesis that neurotrophin regulation of growth cone behaviors involves binding to p75, activation of protein kinase A (PKA) and inhibition of p75-mediated activation of RhoA GTPase. (2) Elucidate the cytoskeletal changes and interactions that accompany growth cone collapse and retraction. We will test the hypothesis that myosin II contractility drives growth cone collapse, retraction and cytoskeletal rearrangements. We will test the hypothesis that neurotrophins stabilize actin filaments against collapse by preventing loss of actin monomer from filament pointed ends. These studies will involve tissue culture of sensory, retinal and ciliary neurons from chick embryos and sensory and retinal neurons from mice with mutant p75 and trkB genes. Explants of neural tissues will be prepared and growth cone behaviors will be analyzed by video microscopy and quantitative fluorescence microscopy. Responses of growth cones to several molecular guidance cues will be determined, and the activities of PKA and Rho family GTPases will be measured. Cytoskeletal organization will be analyzed in growth cones treated with neurotrophins and with negative guidance cues.
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