Ca2+-activated myosin-ATPases, creatine and adenylate kinases regulate mitochondrial function according to myofibre type in rabbit

Mitochondrial respiration rates and their regulation by ADP, AMP and creatine, were studied at different free Ca2+ concentrations (0.1 versus 0.4 mu m) on permeabilized fibre bundles of rabbit skeletal muscles differing in their myosin heavy chain profiles. Four fibre bundle types were obtained: pure types I and IN, and mixed types IIax (approximately 50% IIa and 50% IIx fibres) and IIb+ (60% IIb fibres, plus IIx and IIa). At rest, pure type I fibres displayed a much higher apparent K,,, for ADP (212 pm) than Ilx fibres (8 pm). Within the IIax and IIb+ mixed fibre bundle types, two K ADP values were observed (70 mu m and 5 tim). Comparison between pure IIx and mixed types indicates that the intermediate Km of 70 pm most probably corresponds to the mitochondrial affinity for ADP in IIa fibres, the lowest K-m for ADP (5 mu m) corresponding to IIx and Ilb types. Activation of mitochondrial creatine and adenylate kinase reactions stimulated mitochondrial respiration only in type I and Ilax fibre bundles, indicating an efficient coupling between both kinases and ADP rephosphorylation in type I and, likely, IIa fibres, since no effect was observed in pure IIx fibres. Following Ca2+ induced activation of myosin-ATPase, an increase in mitochondrial sensitivity to ADP of 45% and 250% was observed in type Ilax and I bundles, respectively, an effect mostly prevented by addition of vanadate, an inhibitor of myosin-ATPase. Ca2+ -induced activation of myosin-ATPase also prevented the stimulation of respiration rates by creatine and AMP in I and Ilax bundles. In addition to differential regulation of mitochondrial respiration and energy transfer systems at rest in I and IIa versus IIx and Ilb muscle fibres, our results indicate a regulation of phosphotransfer systems by Ca2+ via the stimulation of myosin-ATPases in type I and IIa fibres of rabbit muscles.