Long-term calorie restriction reduces proton leak and hydrogen peroxide production in liver mitochondria

1 Department of Molecular Biosciences, School of Veterinary Medicine, University of California, Davis, California; 2 Department of Biochemistry, Microbiology, and Immunology, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada; and 3 Department of Medicine, University of Wisconsin Medical School and Veterans Administration Geriatric Research, Education and Clinical Center, Madison, Wisconsin Submitted 17 August 2004 ; accepted in final form 15 November 2004 Calorie restriction (CR) without malnutrition increases maximal life span in diverse species. It has been proposed that reduction in energy expenditure and reactive oxygen species (ROS) production could be a mechanism for life span extension with CR. As a step toward testing this theory, mitochondrial proton leak, H 2 O 2 production, and markers of oxidative stress were measured in liver from FBNF 1 rats fed control or 40% CR diets for 12 or 18 mo. CR was initiated at 6 mo of age. Proton leak kinetics curves, generated from simultaneous measures of oxygen consumption and membrane potential, indicated a decrease in proton leak after 18 mo of CR, while only a trend toward a proton leak decrease was observed after 12 mo. Significant shifts in phosphorylation and substrate oxidation curves also occurred with CR; however, these changes occurred in concert with the proton leak changes. Metabolic control analysis indicated no difference in the overall pattern of control of the oxidative phosphorylation system between control and CR animals. At 12 mo, no significant differences were observed between groups for H 2 O 2 production or markers of oxidative stress. However, at 18 mo, protein carbonyl content was lower in CR animals, as was H 2 O 2 production when mitochondria were respiring on either succinate alone or pyruvate plus malate in the presence of rotenone. These results indicate that long-term CR lowers mitochondrial proton leak and H 2 O 2 production, and this is consistent with the idea that CR may act by decreasing energy expenditure and ROS production. energy expenditure; aging; oxidative stress; food restriction Address for reprint requests and other correspondence: J. J. Ramsey, Dept. of Molecular Biosciences, School of Veterinary Medicine, Univ. of California, 1 Shields Ave., Davis, CA 95616. (E-mail: jjramsey{at}ucdavis.edu )