Monoamine oxidases A/B (EC 1.4.3.4, MAO), flavoenzymes located on the outer mitochondrial membrane, catalyze the oxidative deamination of biogenic amines, such as dopamine, serotonin, and norepinephrine. In this study, we examined whether the H2O2 formed during the two-electron oxidation of tyramine [4-(2-aminoethyl)phenol] (a substrate for monoamine oxidases A/B) may contribute to the intramitochondrial steady-state concentration of H2O2 ([H2O2]ss) and, thus, be involved in the oxidative impairment of mitochondrial matrix components. Supplementation of intact, coupled rat brain mitochondria with benzylamine, beta-phenylethylamine, or tyramine showed initial rates of H2O2 production ranging from 0.4- to 1.6 nmol H2O2/min/mg protein. ESR analysis of the oxidative deamination of tyramine by intact rat brain mitochondria revealed the formation of hydroxyl (HO.) and carbon-centered radical adducts--the latter probably originating by the (HO.-)-mediated oxidation of mannitol. The signals were substantially enhanced upon addition of FeSO4 and were abolished by catalase. The intramitochondrial [H2O2]ss calculated in terms of glutathione peroxidase activity during the metabolism of tyramine was 48-fold higher (7.71 +/- 0.25 x 10(-7) M) than that obtained during the oxidation of succinate via complex II in the presence of antimycin A (1.64 +/- 0.2 x 10(-8) M). Oxidative damage to the brain mtDNA was assessed by single strand breakage. The ratio of nicked DNA for the preparations treated with tyramine and those without the amine was 1.5 +/- 0.29 (n = 4), 2.12 +/- 0.28 (n = 8, P < or = 0.05), and 3.12 +/- 0.69 (n = 3, P < or = 0.05) at 15, 30, and 60 min, respectively . Preincubation of mitochondria with tranylcypromine (trans-2-phenylcyclopropylamine), an inhibitor to MAO A/B, abolished mtDNA oxidative damage. Catalase inhibited mtDNA strand breakage by approximately 60%. Incubation of intact, coupled rat brain mitochondria with chlorodinitrobenzene (CDNB) depleted mitochondrial GSH by 72%. Tyramine-dependent damage of mtDNA was decreased by 68% in CDNB-treated mitochondria (with 28% remaining GSH). The [H2O2]ss was slightly increased in CDNB-treated mitochondria: 1.38- and 1.28-fold increase during the oxidation of succinate in the presence of antimycin A and during the oxidation of tyramine, respectively. These results suggest that the H2O2 generated during the MAO-catalyzed oxidation of biogenic amines and possibly certain neurotransmitters at the outer mitochondrial membrane contributes to the intramitochondrial [H2O2]ss and may cause oxidative damage to mtDNA. This is effected by the intramitochondrial concentration of GSH and might have potential implications for aging and neurodegenerative processes.