Figure 10.

Calcium decreases mitochondrial membrane potential (ΔΨ). 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, 150 nM TMRE (Ex: 550 nm, Em: 575 nm). Each run was performed with a baseline reading of buffer. Mitochondria, calcium and the substrates/inhibitors were added as indicated. An increase in TMRE fluorescence, as a measure of loss of mitochondrial membrane potential (ΔΨ), was observed when mitochondria treated with calcium compared to no calcium (slope of TMRE traces between 3 and 5 min). Additionally, this real-time loss of ΔΨ was calcium dose-dependent; and was sustained in the presence of ADP (slope of TMRE traces between 5 and 7 min). Additionally, a sharp bell shaped ADP response loss was evident with increasing calcium dose due to reflective prior loss of ΔΨ in presence of calcium. n = 3 per group.