DEPENDENCE OF H2O2 FORMATION BY RAT-HEART MITOCHONDRIA ON SUBSTRATE AVAILABILITY AND DONOR AGE

We have examined the substrate specificity and inhibitor sensitivity o f H2O2 formation by rat heart mitochondria. Active H2O2 production req uires both a high fractional reduction of Complex I (indexed by NADH/N AD(+) + NADH ratio) and a high membrane potential, Delta psi. These co nditions are achieved with supraphysiological concentrations of succin ate. With physiological concentrations of NAD-linked substrates, rates of H2O2 formation are much lower (less than 0.1% of respiratory chain electron flux) but may be stimulated by the Complex III inhibitor ant imycin A, but not by myxothiazol. Addition of Mn2+ to give 10 nmol/mg of mitochondrial protein enhances H2O2 production with all substrate c ombinations, possibly by repleting mitochondrial superoxide dismutase with this cation. Contrary to previously published work, no increased activity of H2O2 production was found with heart mitochondria from sen escent (24 month) rats, relative to young adults (6 month).