Activation of protein kinase C (PKC) produced a novel and rapid formation of lamellae over large surfaces of dendrites in cultured hippocampal neurons. This action was dendrite-specific, involved a postsynaptic locus of activation of PKC and required actin polymerization, but not activation of Erk. Over-expression of active Rho-A GTPase converted a mature, highly branched and spiny neuron into a primitive, non-branching, aspiny neuron. Overexpression of active Rac1 caused massive lamellae formation in the transfected neurons. These morphologically aberrant neurons retained synaptic connectivity with adjacent, normal neurons, as well as the ability to form lamellae in response to PKC. On the other hand, transfection with a dominant negative Rac-N17 or a toxin C3, Rho-A-inactivating plasmid blocked lamellae formation by PKC, but did not prevent PKC-induced plasticity of synaptic currents. These data indicate that PKC activates two independent molecular pathways, one of which involves Rac1 and Rho-A, to produce massive actin-based structural plasticity in dendrites and spines.