Cyclosporine is an immunosuppressant that undergoes extensive hepatic biotransformation to hydroxylated and demethylated metabolites. At present, the CYP3A gene family is thought to be the primary enzyme system responsible for cyclosporine metabolism. The effect of chronic cyclosporine therapy on the suppression of drug metabolism was studied in male and female rats maintained on a low-salt diet. After 28 days of subcutaneous cyclosporine dosing 15 mg/kg, cyclosporine-treated rats had significant renal dysfunction as compared with gender-matched control rats. Creatinine clearance in male cyclosporine-treated rats was reduced by 47% (P < .01) as compared with male controls. Female rats demonstrated a 38% (P < .01) decrease in creatinine clearance as a result of chronic cyclosporine therapy. Despite similar nephrotoxicity, female rats had whole blood cyclosporine levels 48% (P < .01) less than male rats. Immunoblot analysis of hepatic microsomal proteins indicated that chronic cyclosporine treatment decreased the protein levels of P450 3A2 in male rats. This loss was paralleled by reduced production of 6 beta-hydroxytestosterone, the primary product of P450 3A activity, by hepatic microsomes from cyclosporine-treated male rats by 76% (P < .001). In addition, cyclosporine treatment of male rats also reduced the formation of 2 alpha-hydroxytestosterone and 16 alpha-hydroxytestosterone by 81% (P < .01) and 84% (P < .001), respectively. At the end of the study period, steroid 5 alpha-reductase activity in control male rats was only 4% (P < .001) of female counter-parts; however, cyclosporine treatment increased steroid 5 alpha-reductase activity in male rats to 79% (P < .001) of female values. These alterations in testosterone metabolism are consistent with the suppression of the predominately male-associated P450 3A2, P450 2C11 and P450 2C13 isoforms. Levels of 6 alpha-hydroxytestosterone and 7 alpha-hydroxytestosterone were not statistically different between rat groups. Taken together, the steady-state blood levels and metabolism studies suggest that, after chronic cyclosporine treatment, isoforms other than those from the CYP3A family or unidentified members of the CYP3A family are likely responsible for cyclosporine metabolism.