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- Bai 2016 PLOS ONE + (Oxygen consumption by isolated mitochondri … Oxygen consumption by isolated mitochondria is generally measured during state 4 respiration (no ATP production) or state 3 (maximal ATP production at high ADP availability). However, mitochondria ''in vivo'' do not function at either extreme. Here we used ADP recycling methodology to assess muscle mitochondrial function over intermediate clamped ADP concentrations. In so doing, we uncovered a previously unrecognized biphasic respiratory pattern wherein O<sub>2</sub> flux on the complex II substrate, succinate, initially increased and peaked over low clamped ADP concentrations then decreased markedly at higher clamped concentrations. Mechanistic studies revealed no evidence that the observed changes in O<sub>2</sub> flux were due to altered opening or function of the mitochondrial permeability transition pore or to changes in reactive oxygen. Based on metabolite and functional metabolic data, we propose a multifactorial mechanism that consists of coordinate changes that follow from reduced membrane potential (as the ADP concentration in increased). These changes include altered directional electron flow, altered NADH/NAD<sup>+</sup> redox cycling, metabolite exit, and OAA inhibition of succinate dehydrogenase. In summary, we report a previously unrecognized pattern for complex II energized O<sub>2</sub> flux. Moreover, our findings suggest that the ADP recycling approach might be more widely adapted for mitochondrial studies.st that the ADP recycling approach might be more widely adapted for mitochondrial studies.)
