Figure 9.

Calcium decreases mitochondrial ROS/RNS levels. Mitochondria (100 μg) were added to a constantly stirred cuvette in a total volume of 2 ml of 125 mM KCl buffer at 37°C containing 1 mM L-Arg, 10 μM H₂DCF-DA (Ex: 485 nm, Em: 530 nm). Each run was performed with a baseline reading of buffer. Mitochondria, calcium and the substrates/inhibitors were added as indicated under identical mitochondrial bioenergetics conditions utilized in Figure 3. The slope of DCF fluorescence was quantified for the respective conditions including baseline and expressed as arbitrary units. A significant decrease in DCF fluorescence, as a measure of free radical generation, was observed in mitochondria treated with calcium (500 nmol/mg mitochondrial protein) compared control conditions. The decrease in DCF fluorescence was calcium dose-dependent and was observed throughout with additions of calcium, ADP, oligomycin, FCCP and rotenone but not with succinate addition. Data represent group means ± SD; n = 3 per group. One-Way ANOVA followed by Fisher's post-hoc test; ^*p < 0.05 compared to 0nmol/mg calcium; @p < 0.05 compared to 200 nmol/mg calcium.