Caspase activation is indispensable for the proper execution of apoptosis. However, to date, little is known about other possible physiologic functions for this class of enzymes in addition to their well-defined role in apoptosis. In this report, we described an action of caspase-3 involving cell dispersion that is independent of cell death. Using an in vitro neuronal model system consisting of PC12 cells, we observed a transient activation of caspase-3 both in situ and by Western blot analysis that was evident at 1 h following plating, was maximal by 3 h, and was attenuated by 24 h. Preincubation of PC12 cells with either the caspase-3 inhibitor, DEVD, or antisense caspase-3 oligonucleotides caused cells to be more rounded in appearance and led to a failure of cells to disperse properly. Additional experiments demonstrated a possible target for caspase cleavage to be the cytoskeletal protein, tau. These data suggest a requirement for caspase activation and subsequent disassembly of the cytoskeleton during cell dispersion and represent a novel role for caspases that may allow for proper migration of neurons to target locations during development.