Figure 1.

Calcium uptake (A) and calcium-stimulated ROS (H₂O₂) generation (B) in mitochondria isolated from rat liver. (A) Mitochondria (0.5 mg protein/ml) were energized with 5 mM succinate (+2 µM rotenone) in a standard medium consisting of 125 mM sucrose, 65 mM KCl, and 10 mM HEPES-KOH pH 7.4, at 30°C; calcium (Ca²⁺) was determined with arsenazo III as described in Materials and methods. (B) Mitochondria were incubated in the standard medium at 30°C and H₂O₂ was determined with the Amplex Red probe/HRP as described in Materials and methods. Additions: standard medium + Amplex Red probe/HRP (base line); Mitochondria (M) + 2 µM rotenone + 100 µM EGTA + 5 mM succinate (Suc) (Control); M + 2 µM rotenone + Suc + 10–250 µM calcium (Ca²⁺). Bars are average ± SEM and traces are representative experiments.