Recently, we demonstrated that a microsomal enzyme inducer with a steroidal structure, pregnenolone-16 alpha-carbonitrile (PCN), markedly decreased the hepatotoxicity of acetaminophen (AA) in hamsters. Therefore, it was of interest to determine if PCN, as well as another steroid microsomal enzyme inducer, dexamethasone (DEX), would decrease the toxicity of AA in mice, another species sensitive to AA hepatotoxicity. Mice were pretreated with PCN or DEX (100 and 75 mg/kg, ip, for 4 days, respectively) and were given AA (300-500 mg/kg, ip). Twenty-four hours after AA administration, liver injury was assessed by measuring serum activities of sorbitol dehydrogenase and alanine aminotransferase and by histopathological examination. Neither PCN nor DEX protected markedly against AA hepatotoxicity in mice; PCN tended to decrease AA-induced hepatotoxicity, whereas DEX was found to enhance AA-induced hepatotoxicity and it produced some hepatotoxicity itself. DEX decreased the glutathione concentration (36%) in liver and increased the biliary excretion of AA-GSH, which reflects the activation of AA, whereas PCN produced neither effect. Thus, whereas PCN has been shown to markedly decrease the hepatotoxicity of AA in hamsters, apparently by decreasing the isoform of P450 responsible for activating AA to N-acetyl-p-benzoquinoneimine, this does not occur in mice after induction with either PCN or DEX. In contrast, DEX enhances AA hepatotoxicity apparently by decreasing liver GSH levels and increasing the activation of AA to a cytotoxic metabolite.