Mitochondrial coupling in vivo in mouse skeletal muscle

Abstract

The coupling of mitochondrial ATP synthesis and oxygen consumption (ratio of ATP and oxygen fluxes, P/O) plays a central role in cellular bioenergetics. Reduced P/O values are associated with mitochondrial pathologies that can lead to reduced capacity for ATP synthesis and tissue degeneration. Previous work found a wide range of values for P/O in normal mitochondria. To measure mitochondrial coupling under physiological conditions, we have developed a procedure for determining the P/O of skeletal muscle in vivo. This technique measures ATPase and oxygen consumption rates during ischemia with31P magnetic resonance and optical spectroscopy, respectively. This novel approach allows the independent quantitative measurement of ATPase and oxygen flux rates in intact tissue. The quantitative measurement of oxygen consumption is made possible by our ability to independently measure the saturations of hemoglobin (Hb) and myoglobin (Mb) from optical spectra. Our results indicate that the P/O in skeletal muscle of the mouse hindlimb measured in vivo is 2.16 ± 0.24. The theoretical P/O for resting muscle is 2.33. Systemic treatment with 2,4-dinitrophenol to partially uncouple mitochondria does not affect the ATPase rate in the mouse hindlimb but nearly doubles the rate of oxygen consumption, reducing in vivo P/O to 1.37 ± 0.22. These results indicate that only a small fraction of the oxygen consumption in resting mouse skeletal muscle is nonphosphorylating under physiological conditions, suggesting that mitochondria are more tightly coupled than previously thought.