Characteristics of α-glycerophosphate-evoked H2O 2 generation in brain mitochondria

Abstract

Characteristics of reactive oxygen species (ROS) production in isolated guinea-pig brain mitochondria respiring on α-glycerophosphate (α-GP) were investigated and compared with those supported by succinate. Mitochondria established a membrane potential (Δψm) and released H 2O2 in parallel with an increase in NAD(P)H fluorescence in the presence of α-GP (5-40 mm). H2O2 formation and the increase in NAD(P)H level were inhibited by rotenone, ADP or FCCP, respectively, being consistent with a reverse electron transfer (RET). The residual H2O2 formation in the presence of FCCP was stimulated by myxothiazol in mitochondria supported by α-GP, but not by succinate. ROS under these conditions are most likely to be derived from α-GP-dehydrogenase. In addition, huge ROS formation could be provoked by antimycin in α-GP-supported mitochondria, which was prevented by myxothiazol, pointing to the generation of ROS at the quinol-oxidizing center (Qo) site of complex III. FCCP further stimulated the production of ROS to the highest rate that we observed in this study. We suggest that the metabolism of α-GP leads to ROS generation primarily by complex I in RET, and in addition a significant ROS formation could be ascribed to α-GP-dehydrogenase in mammalian brain mitochondria. ROS generation by α-GP at complex III is evident only when this complex is inhibited by antimycin. © 2007 The Authors.