High Ca²⁺ load promotes Hydrogen peroxide generation via activation of α-glycerophosphate dehydrogenase in brain mitochondria
2012
Tretter, Laszlo | Adam-Vizi, Vera
H₂O₂ generation associated with α-glycerophosphate (α-GP) oxidation was addressed in guinea pig brain mitochondria challenged with high Ca²⁺ load (10μM). Exposure to 10μM Ca²⁺ induced an abrupt 2.5-fold increase in H₂O₂ release compared to that measured in the presence of a physiological cytosolic Ca²⁺ concentration (100nM) from mitochondria respiring on 5mM α-GP in the presence of ADP (2mM). The Ca²⁺-induced stimulation of H₂O₂ generation was reversible and unaltered by the uniporter blocker Ru 360, indicating that it did not require Ca²⁺ uptake into mitochondria. Enhanced H₂O₂ generation by Ca²⁺ was also observed in the absence of ADP when mitochondria exhibited permeability transition pore opening with a decrease in the NAD(P)H level, dissipation of membrane potential, and mitochondrial swelling. Furthermore, mitochondria treated with the pore-forming peptide alamethicin also responded with an elevated H₂O₂ generation to a challenge with 10μM Ca²⁺. Ca²⁺-induced promotion of H₂O₂ formation was further enhanced by the complex III inhibitor myxothiazol. With 20mM α-GP concentration, stimulation of H₂O₂ formation by Ca²⁺ was detected only in the presence, not in the absence, of ADP. It is concluded that α-glycerophosphate dehydrogenase, which is accessible to and could be activated by a rise in the level of cytosolic Ca²⁺, makes a major contribution to Ca²⁺-stimulated H₂O₂ generation. This work highlights a unique high-Ca²⁺-stimulated reactive oxygen species-forming mechanism in association with oxidation of α-GP, which is largely independent of the bioenergetic state and can proceed even in damaged, functionally incompetent mitochondria.
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