Differential effects of low pH on Ca 2+ ‐induced ROS emission from mitochondrial complexes I and III

Mitochondrial emission of reactive oxygen species (ROS) is a critical factor in cardiac ischemic injury. Increased extra‐matrix Ca 2+ , decreased extra‐matrix pH, and altered substrate utilization are factors that modulate ROS emission during ischemia. Here we examined the combined effects of these factors on ROS emission from complexes I and III. Guinea pig heart mitochondria were suspended in experimental buffer (pH 7.15, 6.9, 6.5), pre‐incubated with increasing [CaCl 2 ], and then energized with either pyruvate or succinate followed by adding either rotenone or antimycin A. H 2 O 2 release rate and matrix volume were measured using pyruvate/rotenone or succinate/antimycin A with or without rotenone to mimic conditions that exist during early or late global ischemia, respectively. High buffer CaCl 2 enhanced rotenone or antimycin A‐induced H 2 O 2 release under all conditions. The largest increase in H 2 O 2 release occurred at pH 6.9 or pH 6.5 with pyruvate/rotenone or succinate/antimycin A, respectively. The large increase in H 2 O 2 release was associated with a significant mitochondrial swelling, and both measures were abolished with cyclosporine A, indicating opening of the mitochondrial permeability transition pore. These results highlight the significance of each respiratory complex in Ca 2+ ‐induced ROS emission under different substrate/pH conditions that may mimic early and late ischemia. (NIH and VA)