The molecular mechanisms underlying directed motility of growth cones have not been determined, The role of myosin-V, an unconventional myosin, in growth cone dynamics was examined by chromophore-assisted laser inactivation (CALI), CALI of purified chick brain myosin-V absorbed onto nitrocellulose-coated cover slips inhibited the ability of myosin-V to translocate actin filaments. CALI of myosin-V in growth cones of chick dorsal root ganglion neurons resulted in rapid filopodial retraction, The rate of filopodial extension was significantly decreased, whereas the rate of filopodial retraction was not affected, which suggests a specific role for myosin-V in filopodial extension.