INCREASE IN LIVER MICROSOMAL GLUTATHIONE-S-TRANSFERASE ACTIVITY BY PHENOBARBITAL TREATMENT OF RATS - POSSIBLE INVOLVEMENT OF OXIDATIVE ACTIVATION VIA CYTOCHROME-P450

The possible involvement of oxidative activation of liver microsomal g lutathione (GSH) S-transferase by the cytochrome P450 system was inves tigated. When rats were given phenobarbital (PB) intraperitoneally for 3 days, liver microsomal GSH S-transferase activity was stimulated 1. 3-1.4-fold and the effect of PB on the transferase was potentiated by combination with a catalase inhibitor, 3-amino-1,2,4-triazole. Immunob lotting of microsomal proteins from PB-treated rats with anti-microsom al GSH S-transferase antibody after SDS-PAGE showed the presence of a dimer of the transferase. When microsomal suspensions prepared from PB -treated rats were placed on ice without GSH, the microsomal GSH S-tra nsferase activity gradually increased with time and reached 200% of th e initial level at 3 hr when activation of the transferase by N-ethylm aleimide was lost. The time-dependent increase in GSH S-transferase ac tivity in PB-treated microsomes was prevented by addition of 0.1 mM GS H. The increase in microsomal GSH S-transferase activity by NADPH was depressed by cytochrome P450 inhibitors such as SKF 525-A (2-diethylam inoethyl-2,2-diphenylvalerate), metyrapone or isoniazid in agreement w ith the concomitant decrease in generation of hydrogen peroxide in mic rosomes. These results indicate that the increase in GSH S-transferase activity in liver microsomes by PB treatment of rats is due to the ox idative modification of the enzyme by reactive oxygen species which ar e concomitantly increased following induction of cytochrome P450.