Calcium Regulation of Muscle Mitochondrial Function: Dependence on Substrate and Respiratory State

Presentation Number: SUN 251
Date of Presentation: April 2nd, 2017

William I Sivitz*1, Brian D Fink2, Fan Bai3 and Liping Yu3
1University of Iowa and the Iowa City VAMC, Iowa City, IA, 2Iowa City VAMC, Iowa City, IA, 3University of Iowa, Iowa City, IA


Acute calcium influx to mitochondria is known to enhance respiration by triggering substrate utilization and is critical for muscle contraction. However, the effect of calcium under differential respiratory states (i.e. state 4 to 3) is not clear. Corresponding effects on ATP synthesis, reactive oxygen (ROS), substrate dependency, and opening of the mitochondrial permeability transition pore (MTP) are also not well defined. Here we examined skeletal muscle mitochondrial function on different substrates and utilized an ADP recycling technique to assess the effect of calcium on mitochondrial function at clamped external ADP and clamped membrane potential. All studies were carried out using gradient purified and calcium depleted mitochondria isolated from mouse hindlimb skeletal muscle. O2 flux in mitochondria energized by the complex I substrates glutamate plus malate was significantly enhanced by 500 nM calcium at clamped [ADP] from 4 to 256 micromolar, but most substantially at clamped [ADP] concentrations generating intermediate respiratory states between state 3 and 4. In contrast no effect was observed for mitochondria energized by the complex II substrate, succinate, with or without rotenone (to block reverse electron transport to complex I). The stimulatory effect of calcium on respiration and ATP production in the presence of all concentrations of clamped [ADP] was most prominent at calcium of approximately 450 nM to 900 nM but decreased markedly and became inhibitory at calcium levels of 10 micromolar or higher. Interestingly, membrane potential increased up to 10 micromolar calcium and did not decrease until calcium was raised to greater levels. In the absence of added ADP (state 4 respiration), respiration actually increased as calcium increased up to 50 micromolar while potential decreased. Higher levels of calcium were positively associated with ROS production. In summary, the stimulatory effect of calcium on mitochondrial function is substrate dependent consistent with known effects on NADH dehydrogenase activity, is most prominent at intermediate respiratory states between 3 and 4, and most evident at nM concentrations of calcium. Higher calcium concentrations inhibit respiration under conditions other than state 4. Potential is maintained more than respiration as calcium concentrations are increased before eventual collapse of mitochondrial respiration at high calcium, likely associated with opening of the MTP.


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