Profilins are thought to be essential for regulation of actin assembly, However, the functions of profilins in mammalian tissues are not well understood, In mice profilin I is expressed ubiquitously while profilin II is expressed at high levels only in brain, In extracts from mouse brain, profilin I and profilin II can form complexes with regulators of endocytosis, synaptic vesicle recycling and actin assembly, Using mass spectrometry and database searching we characterized a number of ligands for profilin I and profilin II from mouse brain extracts including dynamin I, clathrin, synapsin, Rho-associated coiled-coil kinase, the Rac-associated protein NAP1 and a member of the NSF/sec18 family, In vivo, profilins co-localize with dynamin I and synapsin in axonal and dendritic processes, Our findings strongly suggest that in brain profilin I and profilin II complexes link the actin cytoskeleton and endocytic membrane how directing actin and clathrin assembly to distinct membrane domains.