INACTIVATION OF CYTOCHROMES P450 2B PROTECTS AGAINST COCAINE-MEDIATEDTOXICITY IN RAT-LIVER SLICES

Mechanism-based inactivators of rat liver cytochrome P450 2B1 and 2B2 were used to evaluate the role of these enzymes in the hepatotoxicity of cocaine. Loss of liver microsomal androstenedione 16 beta-hydroxyla tion was monitored to determine the extent of P450 2B1/2 inactivation by chloramphenicol (CAP) or its 2B-selective analogue, N-(2-p-nitrophe nethyl)chlorofluoroacetamide (pNO(2)C1FA). The effect of P450 2B1/2 in activation on cocaine-mediated hepatotoxicity was assessed in rat live r slices. Exposure of slices from phenobarbital-induced Lewis rats to CAP concentrations ranging from 100 to 500 mu M resulted in a concentr ation-dependent decrease in P450 2B activity and a corresponding decre ase in cytotoxicity as measured by K+ loss following exposure to 1 mM cocaine. Treating slices from PR-induced rats with 250 mu M pNO(2)C1FA protected slices against cocaine-mediated cytotoxicity after exposure to 500 mu M cocaine. In vivo administration of 300 mg/kg CAP or 200 m g/kg pNO(2)C1FA to phenobarbital-induced Lewis rats decreased androste nedione 16 beta-hydroxylation to 30 or 39% of control, respectively, a nd blocked cocaine-mediated K+ loss in rat liver slices. Rat liver mic rosomes from animals treated with either CAP or pNO(2)C1FA displayed a pproximately 40% of the control rate of cocaine N-demethylation. Exper iments with phenobarbital-treated Munich Wistar (WM) rats, which lack 2B2, revealed similar rates of microsomal N-demethylation and comparab le in vitro hepatotoxicity to Lewis rats. The capacity of a specific P 450 2B1/2 inactivator to protect against cocaine-mediated hepatotoxici ty both in vivo and in vitro and the results with the WM rats support the identification of P450 2B1 as a major cocaine bioactivating form. (C) 1994 Academic Press, Inc.