PMC:509410 / 21171-23557 JSONTXT

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Mice were fed either a standard chow with 4% (w/w) fat content (Harlan Teklad, Harlan, Indianapolis, Indiana, United States) or a high-fat diet containing 35% (w/w) fat content (product F3282, Bioserv, Frenchtown, New Jersey, United States) as indicated. For ligand experiments, we synthesized the GW501516 compound and mice were orally gavaged daily (10 mg/kg or vehicle alone).\n\nGene expression analysis and physiological studies\nMouse EST clones were obtained from ATCC (Manassas, Virginia, United States), verified by sequencing, and used as Northern probes. Antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, California, United States). Total muscle protein extracts (Lin et al. 2002) and nuclear proteins (Wang et al. 2003) were prepared as described.\nPrior to the exercise performance test, the mice were accustomed to the treadmill (Columbus Instruments, Columbus, Ohio, United States) with a 5-min run at 7 m/min once per day for 2 d. The exercise test regimen was 10 m/min for the first 60 min, followed by 1 m/min increment increases at 15-min intervals. Exhaustion was defined when mice were unable to avoid repetitive electrical shocks.\n\nMuscle fiber typing and mitochondrial DNA isolation\nMuscle fiber typing was essentially performed using metachromatic dye–ATPase methods as described (Ogilvie and Feeback 1990). Muscle mitochondria were isolated (Scholte et al. 1997). Mitochondrial DNA was prepared and analyzed on 1% agarose gel.\n\nStatistical analysis\nNumber of mice for each group used in experiments is indicated in figure legends. Values are presented as mean ± SEM. 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