Cytochrome P-450 (P-450) and associated mono-oxygenase activities were estimated in male and female rat brain microsomes. The P-450 concentration in male rat brain was one-tenth the corresponding hepatic levels, which is considerably higher than earlier reports. A distinct sex-related difference was observed in the levels of total P-450 and mono-oxygenase activities known to be mediated by P-450b,e; the female brain levels were 60% of those in the males. Immunoinhibition and immunoblot studies using antisera to P-450b,e and P-450c,d indicated the presence of multiple forms of P-450, immunologically similar to P-450b,e, P-450c and P-450d in the rat brain. Prior treatment with phenobarbital resulted in two-fold increase of total P-450 and selective induction of aminopyrine N-demethylase (APD) and morphine N-demethylase (MND) activities. Administration of 3-methylcholanthrene, selectively induced the levels of ethoxycoumarin O-deethylase (ECD) and arylhydrocarbon hydroxylase, although the levels of total P-450 were not increased. 3-Methylcholanthrene induction was also accompanied by a shift in the absorption maximum of the reduced carbon monoxide difference spectrum from 452 to 448 nm. Immunocytochemical localization using antibodies to P-450b,e indicated the presence of P-450 predominantly in the neuronal cell bodies and to a lesser extent in the fibre tracts in cerebral cortex, cerebellum, thalamus, hypothalamus, hippocampus and brainstem. These studies indicate that the brain contains significant amounts of P-450, which exists in multiple forms and can be selectively induced by prior exposure to phenobarbital or 3-methylcholanthrene.