Abstract
Fuel substrate supply and oxidative phosphorylation are key determinants of muscle performance. Numerous studies of mammalian mitochondria are carried out (i) with substrate supply that limits electron flow, and (ii) far below physiological temperature. To analyze potentially implicated biases, we studied mitochondrial respiratory control in permeabilized mouse myocardial fibers using high-resolution respirometry. The capacity of oxidative phosphorylation at 37 °C was nearly two-fold higher when fueled by physiological substrate combinations reconstituting tricarboxylic acid cycle function, compared with electron flow measured separately through NADH to Complex I or succinate to Complex II. The relative contribution of the NADH pathway to physiological respiratory capacity increased with a decrease in temperature from 37 to 25 °C. The apparent excess capacity of cytochrome c oxidase above physiological pathway capacity increased sharply under hypothermia due to limitation by NADH-linked dehydrogenases. This mechanism of mitochondrial respiratory control in the hypothermic mammalian heart is comparable to the pattern in ectotherm species, pointing towards NADH-linked mt-matrix dehydrogenases and the phosphorylation system rather than electron transfer complexes as the primary drivers of thermal sensitivity at low temperature. Delineating the link between stress and remodeling of oxidative phosphorylation is important for understanding metabolic perturbations in disease evolution and cardiac protection.
Highlights
Contractile activity in cardiac muscle mainly depends on mitochondrial energy transformed by oxidative phosphorylation (OXPHOS)
Mitochondrial respiration in the living cell is supported by fuel substrates supplying electrons to multiple dehydrogenases followed by convergent electron entry into the Q-junction
Physiological respiratory capacity is underestimated in isolated mitochondria and permeabilized fibers when using simple substrate combinations such as pyruvate&malate supporting the NADH-pathway (N), or a single substrate for the succinate-pathway (S)
Summary
Contractile activity in cardiac muscle mainly depends on mitochondrial (mt) energy transformed by oxidative phosphorylation (OXPHOS). Fuel substrates supporting convergent electron transfer at the Q-junction enhance respiratory capacity, as shown when succinate is added to NADH-linked substrates, reconstituting physiological tricarboxylic acid cycle function with combined NS-pathway flux. This effect of succinate varies depending on species, strains, organ and experimental conditions; stimulation is 1.6 to 2.0-fold in rat heart22, 23, 1.2 to 1.8-fold in rat skeletal muscle24–26, 1.4-fold in mouse skeletal muscle[27], and 1.3 to 2.1-fold in human skeletal muscle (reviewed by Gnaiger[28]). Our results provide new perspectives on evolutionary temperature adaptation, suggesting that key control of energy transformation and survival at low temperatures might not primarily be exerted by electron transfer complexes but by (i) upstream elements of the electron transfer system (ETS) including transport of substrates across the inner mitochondrial membrane and matrix dehydrogenases, and (ii) downstream elements of the phosphorylation system
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