Rho-GDIs are a family of Rho GDP-dissociation inhibitors that are critical in modulating the activity of the small GTPases, Cdc42 and RhoA. Two Rho-GDI isoforms are expressed in the brain, Rho-GDIgamma and Rho-GDIalpha. Here, we describe the expression of both of these isoforms in the developing and mature brain. The mRNA expression patterns of Rho-GDIgamma and Rho-GDIalpha were almost identical in the brain with expression in the developing and mature cerebral cortex, striatum, and hippocampus. In addition, we generated mice with targeted deletions of Rho-GDIgamma that are viable and fertile and have no obvious phenotypic abnormalities. Mutant brains looked histologically normal and demonstrated normal patterns of dendritogenesis and neuronal layering as determined by Golgi staining. Mutant mice had normal sleep/wake patterns and sleep EEGs and showed normal hippocampal-dependent learning as assayed by the Morris water maze task. Based on the co-expression of Rho-GDIalpha and Rho-GDIgamma in identical populations of cells in the brain, the lack of phenotype caused by targeted deletion of Rho-GDIgamma may not be surprising given that Rho-GDIalpha may compensate for the loss of Rho-GDIgamma. Whether deletion of both Rho-GDIalpha and Rho-GDIgamma, thereby eliminating all GDI activity in the brain, would produce an observable phenotype remains to be determined.