On the Mechanism(s) of Membrane Permeability Transition in Liver Mitochondria of Lamprey, Lampetra fluviatilis L.: Insights from Cadmium
Figure 5
The lamprey liver mitochondrial respiration depending on the season, the substrate used and the presence of Cd2+ and/or cyclosporine A. The rates of LLM respiration (oxygen consumption rates) were monitored polarographically at 25°C with a Clark oxygen electrode in a thermostatic closed chamber of 1.5 mL with magnetic stirring. The mitochondria (1 mg of protein/mL) were added into a medium containing 250 mM sucrose, 3 mM MgCl2, 3 mM KH2PO4, 3 mM Tris-HCl, pH 7.3 ((a) and (b), winter LLM) or 100 mM sucrose, 40 mM KCl, 3 mM MgCl2, 3 mM KH2PO4, and 20 mM Tris-HCl, pH 7.3 (±50 M of EGTA and/or 1 mg/mL BSA) ((c), spring LLM). 5 mM pyruvate or glutamate and 1 or 5 mM malate or 5 mM succinate (±5 M of rotenone) were added to the incubation medium for energization of the mitochondria. The concentration of oligomycin, if added, was 1–5 g/mL. The basal respiration rate (in the presence of substrates), in state 3 (substrates and 50 M ADP), in state 4 (after exhausting of ADP) and uncoupled respiration rate (2, 4-DNP was added in state 4) were estimated. The respiration rates (ngatom O/min/mg of protein), respiratory control ratios [VO2(3)/VO2(4)], ADP/O2, and the phosphorylation rate were calculated from polarographic curves. Typical traces for three independent mitochondrial preparations are shown.
