Fig. 3.

Effect of respiratory substrate and cyclosporine A on the Ca²⁺-induced swelling, depolarization, Ca²⁺ release and ROS generation of isolated mitochondria. Experimental conditions were the same as described in Figure 2. Membrane depolarization, Ca²⁺-release and ROS generation were analyzed with TMRE, Rhod 2 and hydroethidine (HE), respectively, and detected with flow cytometry for changes in FL2-H in the gated R1. Mitochondrial swelling was also analyzed simultaneously with these events by SSC. (Left, Control) MPT was induced by addition of 50 µM Ca²⁺ for 5 min at 25°C. Mitochondria were incubated in the standard medium containing 2.5 mM succinate and 0.5 mM Pi before addition of Ca²⁺. (Center, -Succ) The effect of the absence of respiratory substrate on the Ca²⁺-induced MPT. Mitochondria were incubated in standard medium containing 0.5 mM Pi without succinate (Suc). (Right, +CsA) The effect of cyclosporine A on the Ca²⁺-induced MPT. Mitochondria were incubated in the presence of 1 µM CsA in the standard medium containing 2.5 mM succinate and 0.5 mM Pi before addition of Ca²⁺. Similar results were obtained in 3 separate experiments.