Revised Abstract: Neuronal migration is abnormal in human fetuses carried by mothers using drugs of abuse, which has been replicated in animal models. The understanding of mechanisms leading to defective neuronal migration requires investigation of mechanisms underlying normal neuronal migration. The secreted guidance cue Slit is a protein important for controlling axon projection and neuronal migration in the mammalian brain. Its axonal targets include axons from dopaminergic neurons in the basal forebrain and the retinal ganglionic cells, whereas its neuronal targets include neuronal precursor cells in the embryonic striatal primordium and those in the subventricular zone in the forebrain. Studies of mechanisms of Slit signaling will further our fundamental understanding of axon guidance and neuronal migration, shed light on the projection of dopaminergic axons and migration of neurons in the striatum, and provide a basis for further studies of the effect of addictive substances on neuronal migration. This application will focus on studying signal transduction mechanisms mediating cellular responses to Slit, a diffusible chemorepellent for projecting axons and migrating neurons. Our preliminary studies have led to a hypothetical signal transduction pathway for Slit. It begins with the binding of Slit to the extracellular part of the transmembrane receptor Roundabout (Robo), which increases the interaction of Robo with intracellular proteins and inactivates Cdc42, a member of the Rho family of small GTPases, and WISH, a regulator of actin polymerization. Slit-Robo interaction is hypothesized to decrease the activity of N-WASP, an activator of the Arp2/3 complex, which initiates actin polymerization. This sequence of activation and inactivation will result in repulsion. We will test the proposed pathway by investigating the biochemical and functional relationship of these molecules.