Inhibition of brain mitochondrial respiration by dopamine: involvement of H₂O₂ and hydroxyl radicals but not glutathione–protein–mixed disulfides

Abstract

Examination of the downstream mediators responsible for inhibition of mitochondrial respiration by dopamine (DA) was investigated. Consistent with findings reported by others, exposure of rat brain mitochondria to 0.5 mM DA for 15 min at 30°C inhibited pyruvate/glutamate/malate-supported state-3 respiration by 20%. Inhibition was prevented in the presence of pargyline and clorgyline demonstrating that mitochondrial inhibition arose from products formed following MAO metabolism and could include hydrogen peroxide (H 2 O 2 ), hydroxyl radical, oxidized glutathione (GSSG) or glutathione-protein mixed disulfides (PrSSG). As with DA, direct incubation of intact mitochondria with H 2 O 2 (100 lM) significantly inhibited state-3 respiration. In contrast, incubation with GSSG (1 mM) had no effect on O 2 consumption. Exposure of mitochondria to 1 mM GSSG resulted in a 3.3-fold increase in PrSSG formation compared with 1.4-and 1.5-fold increases in the presence of 100 lM H 2 O 2 or 0.5 mM DA, respectively, suggesting a dissociation between PrSSG formation and effects on respiration. The lack of inhibition of respiration by GSSG could not be accounted for by inadequate delivery of GSSG into mitochondria as increases in PrSSG levels in both membranebound (2-fold) and intramatrix (3.5-fold) protein compartments were observed. Furthermore, GSSG was without effect on electron transport chain activities in freeze-thawed brain mitochondria or in pig heart electron transport particles (ETP). In contrast, H 2 O 2 showed differential effects on inhibition of respiration supported by different substrates with a sensitivity of succinate > pyruvate/malate > glutamate/malate. NADH oxidase and succinate oxidase activities in freeze-thawed mitochondria were inhibited with IC 50 approximately 2-3-fold higher than in intact mitochondria. ETPs, however, were relatively insensitive to H 2 O 2 . Co-administration of desferrioxamine with H 2 O 2 had no effect on complex I-associated inhibition in intact mitochondria, but attenuated inhibition of rotenone-sensitive NADH oxidase activity by 70% in freezethawed mitochondria. The results show that DA-associated inhibition of respiration is dependent on MAO and that H 2 O 2 and its downstream hydroxyl radical rather than increased GSSG and subsequent PrSSG formation mediate the effects.

Mitochondrial Dysfunction and Oxidative Damage in parkin-deficient Mice

“…This may suggest a connection between the mitochondrial dysfunction and our findings of increased extracellular DA in the striatum of parkinϪ/Ϫ mice. Elevated extracellular DA has been shown to create an oxidizing environment both in vitro and in vivo (62,63). This environment results in ROS damage and mitochondrial dysfunction analogous to our findings (64).…”

Dopaminergic midbrain neurons are the prime target for mitochondrial DNA deletions

“…It is well recognized that dopamine metabolism leads to the production of ROS [8]. Interestingly, in the context of the presumed role of complex I in PD, dopamine and some of its metabolites can act as inhibitors of complex I [18,24]. It has also been shown that neuromelanin, which is a byproduct of dopamine metabolism, induces oxidative stress in mitochondria through the release of iron [35].…”

Neurodegeneration and neuroprotection in Parkinson disease

“…Perhaps most importantly, DA itself can block complex 1 activity, 80 although it may be that the depletion of ATP is more directly due to monoamine oxidase conversion of DA to its metabolite DOPAC, which engenders production of hydrogen peroxide. 81 MPTP, via a combination of these effects, induces a range of oxidative reactions, including nitration reactions via peroxynitrite, which has been found to react with alphasynuclein 82 and tyrosine hydroxylase. 83 While the 6-hydroxydopamine (6-OHDA) model, which is selectively neurotoxic for DA neurons, 84 suggests that the DA uptake transporter and oxyradical products derived from DA metabolism may initiate selective SN degeneration, the role for oxidized cytosolic DA initially came from work with methamphetamine toxicity.…”