Mitochondrial Physiology Network 10.9: 17 (2005) - download pdf

Mitochondrial studies in situ reveal a novel mechanism of dysfunction: uncoupling in aged muscle.

CE Amara, DJ Marcinek, KE Conley, SA Jubrias, EG Shankland, Martin J Kushmerick

Dept. Radiol., Univ. Washington, Seattle, WA 98195, USA. - kushmeri@u.washington.edu 

    Studies of mitochondria in biochemically purified organelles or permeabilized fibers in vitro are not always made under conditions mimicking their normal in situ environment of intact muscle.  We developed NMR and optical spectroscopic methods to quantify mitochondrial function directly in intact muscle of humans and mice to establish a “gold standard” by which to identify and quantify abnormal function.  We focused on aging because muscles in the elderly have reduced aerobic ATP synthetic capacity.  We showed a substantial decline in aerobic phosphorylation capacity due to both reduced mitochondrial volume and reduced ATP synthesis capacity per mitochondrial unit.  The latter is an example of dysfunction, not merely reduced function.  Most work on mitochondria in aged muscle focuses on respiration.  However, our recent in vivo experiments revealed significant uncoupling of ATP synthesis and respiration with age in both mouse and human muscle as we now describe.

    In order to evaluate the effect of mitochondrial dysfunction on energetics in intact muscle, the following characteristics need to be quantified:  ATP energy store generated, oxygenation, capacity for ATP synthesis by mitochondria, and coupling of phosphorylation to respiration (P/O ratio).  In aged human and mouse muscle there is little change in total creatine and a tendency for ATP to be reduced with the consequence that the energy stores are maintained with minimal decrement in chemical potential. In young and aged muscle myoglobin is normally not fully saturated indicating that mitochondria function in vivo at very low p_O2 (<8 torr, 1.1 kPa).  The rate of PCr resynthesis following a metabolic perturbation is reduced in aged muscle and phosphorylation is decreased in elderly muscle per unit mitochondrial volume.

    In addition our results also revealed the novel finding that phosphorylation is uncoupled to respiration in aged muscle.  Mouse young adult leg muscle has a P/O = 2.2 whereas it is reduced to 1.1 in 30 month old mice.  In elderly human muscle our recent data reveal the tibialis anterior muscle is normally well-coupled (P/O ~ 2.5) whereas the first dorsal interosseous of the hand can be substantially uncoupled in the same individual with P/O~1.9 for subjects >65 years.  There is also a reduction in resting energy demand because oxygen consumption is not increased with uncoupling.  The differences found in human individuals can be exploited to characterize the underlying mechanisms and their functional consequences.

   To summarize, our work identified uncoupling of oxidative phosphorylation in elderly muscle (a mitochondrial dysfunction) as a new mechanism accounting for the reduced capacity of ATP synthesis characteristic of muscle in elderly subjects.  This mechanism is in addition to the known loss of mitochondrial mass (sarcopenia, reduced mitochondrial volume) and respiratory chain capacity. Our ability to measure coupling in vivo in different human muscles allows us to design new experiments to identify variation in the effects of aging on mitochondrial function.  We are actively exploring the mechanisms involved in this novel observation of uncoupling in both mouse and human muscle and their functional consequences.