The regeneration of reduced glutathione in rat forebrain mitochondria identifies metabolic pathways providing the NADPH required

Metabolic pathways underlying the regeneration of reduced glutathione were investigated in acutely isolated metabolically active mitochondria from rat forebrain. The application of hydrogen peroxide to the organelles was acco mpanied by a transient increase in glutathione disulfide. The recovery of r educed glutathione was significantly improved in the presence of alternativ ely succinate, malate, citrate, isocitrate, or beta-hydroxybutyrate. Inhibi tion of succinate dehydrogenase by malonate abolished the beneficial effect of succinate on the reduction of glutathione disulfide but did not influen ce the effect of isocitrate, Fluorocitrate, an inhibitor of aconitase, bloc ked the effect exerted by citrate but did not inhibit the effects of malate or beta-hydroxybutyrate. Uncoupling of the respiratory chain by carbonyl c yanide m-chlorophenylhydrazone prevented the beneficial effect of beta-hydr oxybutyrate but did not abolish the improved reduction of mitochondrial glu tathione disulfide in the presence of malate and isocitrate. These results suggest that NADP(+)-dependent isocitrate dehydrogenase as well as malic en zyme and nicotinamide nucleotide transhydrogenase contribute to the regener ation of NADPH required for the reduction of glutathione disulfide in brain mitochondria. (C) 1999 Elsevier Science Ireland Ltd. All rights reserved.