Clotrimazole, an N-substituted imidazole, is a widely used topical agent for the treatment of superficial fungal infections. In this study, the effect of application of clotrimazole to the skin of neonatal rats on the induction response of the cytochrome P-450-dependent monooxygenase system in epidermis and fiver has been examined. A single topical application of clotrimazole (10 mg/100 g) to rats resulted in a 53% increase in hepatic cytochrome P-450 content. Clotrimazole treatment also resulted in significant induction of epidermal 7-ethoxycoumarin-O-deethylase activity. Hepatic pnitrophenol hydroxylase, an enzyme, catalyzed principally by the ethanol inducible cytochrome P-450 isozyme, was also significantly induced (58%) by topically applied clotrimazole. This enzyme activity was undetectable in epidermal microsomes. Further characterization of the cytoehrome P-450 isozymes induced in fiver by clotrimazole treatment was based on monoclonal antibodies (MAbs) raised against purified rat liver cytochrome P-450 isozymes induced by phenobarbital (MAb 2-66-3) and ethanol (MAh 1-98-1). Hepatic microsomes prepared from clotrimazole-treated rats showed significant immunoreactivity on Western blot with both the MAbs whereas no reactivity occurred in epidermal microsomes. Our data indicate that topical application of clotrimazole to rats results in the induetion of selected cytoehrome P-450 isozyme(s) in fiver and epidermis which may have implications for the therapeutic use of this compound.