The role of oxygen in the induction of mitochondrial permeability transitions was studied. Oxygen consumption, swelling, membrane potential and calcium transport were recorded simultaneously in isolated rat liver mitochondria. Oxygen depletion was accomplished by saturating the medium with N2 and allowing either mitochondrial respiration or glucose/glucose oxidase to consume the residual oxygen. Upon anaerobiosis, mitochondria were supplemented with 500 microM ATP to support succinate-driven membrane potential. Under these conditions, 100 microM Ca2+ induced cyclosporin A-sensitive permeability transitions. To eliminate the possible inhibition of permeability transition by high concentrations of adenine nucleotides, anaerobic mitochondria were also energized by the combination of 20 microM ADP and phosphoenolpyruvate/pyruvate kinase. These mitochondria also underwent Ca2+-induced permeability transition. Under both of these conditions, namely the addition of ATP as a single or through actions of pyruvate kinase, the respiratory components were totally reduced. Thus, oxygen is not a necessary factor for mitochondria to undergo permeability transitions.