It has been hypothesized that insulin resistance is mediated by a deficiency of mitochondria in skeletal muscle. In keeping with this hypothesis, high-fat diets that cause insulin resistance have been reported to result in a decrease in muscle mitochondria. In contrast, we found that feeding rats high-fat diets that cause muscle insulin resistance results in a concomitant gradual increase in muscle mitochondria. This adaptation appears to be mediated by activation of peroxisome proliferator-activated receptor (PPAR)␦ by fatty acids, which results in a gradual, posttranscriptionally regulated increase in PPAR ␥ coactivator 1␣ (PGC-1␣) protein expression. Similarly, overexpression of PPAR␦ results in a large increase in PGC-1␣ protein in the absence of any increase in PGC-1␣ mRNA. We interpret our findings as evidence that raising free fatty acids results in an increase in mitochondria by activating PPAR␦, which mediates a posttranscriptional increase in PGC-1␣. Our findings argue against the concept that insulin resistance is mediated by a deficiency of muscle mitochondria.I t has been hypothesized that insulin resistance in patients with impaired or diabetic glucose tolerance is mediated by a deficiency of mitochondria in skeletal muscle (1, 2). The mechanism by which a decrease in mitochondria is proposed to cause insulin resistance is accumulation of intramyocellular lipids caused by a decrease in the capacity to oxidize fat (2). This hypothesis is based on the finding that type 2 diabetics and insulin-resistant individuals with impaired glucose tolerance have Ϸ30% less mitochondria in their muscles than insulinsensitive control subjects (3-7). In support of this concept, recent studies have reported that raising serum free fatty acids (FFA) by a high-fat diet in humans (8), or by feeding mice or rats high-fat diets (8-10), results in decreases in skeletal muscle peroxisome proliferator-activated receptor ␥ coactivator-1␣ (PGC-1␣) mRNA (8-10) and the mRNA levels of various mitochondrial constituents (8). In contrast, a number of earlier studies provided evidence that high-fat diets induce increases in mitochondrial marker enzymes (11-14), and Turner et al. (15) recently reported that a high-fat diet resulted in increases in mitochondrial biogenesis and fatty acid oxidative capacity in skeletal muscle of mice.We have found that raising serum FFA in rats by feeding them a high-fat diet and giving them daily heparin injections results in an increase in muscle mitochondria (16). The initial purpose of the present study was to determine whether the more modest increase in FFA induced by a high-fat diet also results in increased mitochondrial biogenesis with an increase in the capacity of muscle to oxidize fat. We found that a high-fat diet does induce an increase in muscle mitochondria. This finding made it possible to evaluate whether a high-fat diet causes muscle insulin resistance despite increases in mitochondria and fat oxidative capacity.Overexpression of peroxisome proliferator-activated receptor (PPAR)␦ i...

Exercise results in rapid increases in expression of the transcription coactivator peroxisome proliferator-activated receptor ␥ coactivator-1␣ (PGC-1␣) and in mitochondrial biogenesis in skeletal muscle. PGC-1␣ regulates and coordinates mitochondrial biogenesis, and overexpression of PGC-1␣ in muscle cells results in increases in mitochondrial content. In this context, it has been proposed that the increase in PGC-1␣ protein expression mediates the exercise-induced increase in mitochondrial biogenesis. However, we found that mitochondrial proteins with a short half-life increase as rapidly as, or more rapidly than, PGC-1␣ protein. This finding led us to hypothesize that activation, rather than increased expression, of PGC-1␣ mediates the initial phase of the exercise-induced increase in mitochondria. In this study, we found that most of the PGC-1␣ in resting skeletal muscle is in the cytosol. Exercise resulted in activation of p38 MAPK and movement of PGC-1␣ into the nucleus. In support of our hypothesis, binding of the transcription factor nuclear respiratory factor 1 (NRF-1) to the cytochrome c promoter and NRF-2 to the cytochrome oxidase subunit 4 promoter increased in response to exercise prior to an increase in PGC-1␣ protein. Furthermore, exercise-induced increases in the mRNAs of cytochrome c, ␦-aminolevulinate synthase, and citrate synthase also occurred before an increase in PGC-1 protein. Thus, it appears that activation of PGC-1␣ may mediate the initial phase of the exercise-induced adaptive increase in muscle mitochondria, whereas the subsequent increase in PGC-1␣ protein sustains and enhances the increase in mitochondrial biogenesis.

It is now generally accepted that activation of AMPactivated protein kinase (AMPK) is involved in the stimulation of glucose transport by muscle contractions. However, earlier studies provided evidence that increases in cytosolic Ca 2؉ mediate the effect of muscle contractions on glucose transport. The purpose of this study was to test the hypothesis that both the increase in cytosolic Ca 2؉ E xercise and insulin stimulate glucose transport by separate pathways, and their maximal effects on muscle glucose uptake are additive (1). Both muscle contractions and insulin increase glucose transport by inducing translocation of the GLUT4 isoform of the glucose transporter from intracellular sites to the cell surface (1). Results of early studies suggested that the increase in cytosolic Ca 2ϩ during contractile activity initiates the process that leads to increased muscle glucose transport (2-5). However, because increases in Ca 2ϩ caused the muscles to contract, it was not possible to clearly dissociate the effects of Ca 2ϩ from the metabolic consequences of the contraction-induced decrease in highenergy phosphates. This problem was surmounted in experiments in which caffeine or W-7, agents that release Ca 2ϩ from the sarcoplasmic reticulum, were used to raise cytosolic Ca 2ϩ to levels too low to cause muscle contraction or a decrease in high-energy phosphates (6). In these experiments, glucose transport increased in response to raising cytosolic Ca 2ϩ to subcontraction levels (6). We interpreted this finding as evidence supporting our hypothesis that Ca 2ϩ mediates the effect of exercise on muscle glucose transport.More recent studies have shown that 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) stimulates glucose transport in skeletal muscle (7)(8)(9)(10)(11). AICAR is taken up by cells and converted to the AMP analog ZMP, which mimics the stimulatory effect of AMP on AMP kinase (AMPK) (12). Like the effect of exercise, the stimulation of glucose transport by AICAR is not inhibited by wortmannin and is additive to that of a maximal insulin stimulus (8). AMPK is activated by increases in the AMP-to-ATP ratio and decreases in phosphocreatine, and thus is activated during muscle contractions (12).Although it seems firmly established that activation of AMPK is involved in mediating the stimulation of glucose uptake by muscle contractions, it does not appear to account for all of the increase in glucose transport activity. This is evidenced by the finding of Mu et al. (13) that expression of a dominant inhibitory mutant of AMPK in mouse muscle results in only an ϳ30 -40% decrease in contraction-stimulated glucose transport. In this context, the purpose of the present study was to test the hypothesis that both the increase in cytosolic Ca 2ϩ and the activation of AMPK during muscle contractions are involved in mediating the stimulation of glucose transport by contractile activity. AICAR, 5-aminoimidazole-4-carboxamide ribonucleoside; AMPK, AMP kinase; CAMK, calmodulin-dependent protein kinase; 2-DG, 2-[1,2-3 H]d...

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Previous studies have shown that raising cytosolic calcium in myotubes induces increases in peroxisome proliferator-activated receptor ␥ coactivator-1␣ expression and mitochondrial biogenesis. This finding suggests that the increases in cytosolic calcium in skeletal muscle during exercise may mediate the exercise-induced increase in mitochondria. The initial aim of this study was to determine whether raising calcium in skeletal muscle induces the same adaptations as in myotubes. We found that treatment of rat epitrochlearis muscles with a concentration of caffeine that raises cytosolic calcium to a concentration too low to cause contraction induces increases in peroxisome proliferator-activated receptor ␥ coactivator-1␣ expression and mitochondrial biogenesis. Our second aim was to elucidate the pathway by which calcium induces these adaptations. Raising cytosoliccalciumhasbeenshowntoactivatecalcium/calmodulindependent protein kinase in muscle. In the present study raising cytosolic calcium resulted in increases in phosphorylation of p38 mitogen-activated protein kinase and activating transcription factor-2, which were blocked by the calcium/calmodulindependent protein kinase inhibitor KN93 and by the p38 mitogen-activated protein kinase inhibitor SB202190. The increases in peroxisome proliferator-activated receptor ␥ coactivator-1␣ expression and mitochondrial biogenesis were also prevented by inhibiting p38 activation. We interpret these findings as evidence that p38 mitogen-activated protein kinase is downstream of calcium/calmodulindependent protein kinase in a signaling pathway by which increases in cytosolic calcium lead to increases in peroxisome proliferator-activated receptor ␥ coactivator-1␣ expression and mitochondrial biogenesis in muscle.

The purpose of the present investigation was to explore the effects of exercise and adrenaline on the mRNA expression of PGC-1α, a master regulator of mitochondrial biogenesis, in rat abdominal adipose tissue. We hypothesized that (1) exercise training would increase PGC-1α mRNA expression in association with increases in mitochondrial marker enzymes, (2) adrenaline would increase PGC-1α mRNA expression and (3) the effect of exercise on PGC-1α mRNA expression in white adipose tissue would be attenuated by a β-blocker. Two hours of daily swim training for 4 weeks led to increases in mitochondrial marker proteins and PGC-1α mRNA expression in epididymal and retroperitoneal fat depots. Additionally, a single 2 h bout of exercise led to increases in PGC-1α mRNA expression immediately following exercise cessation. Adrenaline treatment of adipose tissue organ cultures led to dose-dependent increases in PGC-1α mRNA expression. A supra-physiological concentration of adrenaline increased PGC-1α mRNA expression in epididymal but not retroperitoneal adipose tissue. β-Blockade attenuated the effects of an acute bout of exercise on PGC-1α mRNA expression in epididymal but not retroperitoneal fat pads. In summary, this is the first investigation to demonstrate that exercise training, an acute bout of exercise and adrenaline all increase PGC-1α mRNA expression in rat white adipose tissue. Furthermore it would appear that increases in circulating catecholamine levels may be one potential mechanism mediating exercise induced increases in PGC-1α mRNA expression in rat abdominal adipose tissue.

-To discover the alterations in lipid metabolism linked to postexercise hypotriglyceridemia, we measured lipid kinetics, lipoprotein subclass distribution and lipid transfer enzymes in seven healthy, lean, young men the day after 2 h of cycling and rest. Compared with rest, exercise increased fatty acid rate of appearance and whole body fatty acid oxidation by ϳ65 and 40%, respectively (P Ͻ 0.05); exercise had no effect on VLDL-triglyceride (TG) secretion rate, increased VLDL-TG plasma clearance rate by 40 Ϯ 8%, and reduced VLDL-TG mean residence time by ϳ40 min and VLDL-apolipoprotein B-100 (apoB-100) secretion rate by 24 Ϯ 8% (all P Ͻ 0.05). Exercise also reduced the number of VLDL but almost doubled the number of IDL particles in plasma (P Ͻ 0.05). Muscle lipoprotein lipase content was not different after exercise and rest, but plasma lipoprotein lipase concentration increased by ϳ20% after exercise (P Ͻ 0.05). Plasma hepatic lipase and lecithin:cholesterol acyltransferase concentrations were not affected by exercise, whereas cholesterol ester transfer protein concentration was ϳ10% lower after exercise than after rest (P ϭ 0.052). We conclude that 1) greater fatty acid availability after exercise does not stimulate VLDL-TG secretion, probably because of the increase in fatty acid oxidation and possibly also fatty acid use for restoration of tissue TG stores; 2) reduced secretion of VLDL-apoB-100 lowers plasma VLDL particle concentration; and 3) increased VLDL-TG plasma clearance maintains low plasma TG concentration but is not accompanied by similar increases in subsequent steps of the delipidation cascade. Acutely, therefore, the cardioprotective lowering of plasma TG and VLDL concentrations by exercise is counteracted by a proatherogenic increase in IDL concentration.lipoprotein; fatty acid; stable isotope; metabolism ALTERATIONS IN LIPOPROTEIN METABOLISM, manifested by increased plasma triglyceride (TG) and low-density lipoprotein (LDL) cholesterol and decreased high-density lipoprotein (HDL) cholesterol concentrations, are associated with a greater risk for the development of cardiovascular disease (CVD) (27), the leading cause of death in the US (31). Exercise has long been recognized as an effective means to improve insulin sensitivity (12, 36) and prevent CVD; in fact, one-third of CVD-related deaths are attributable to physical inactivity (35).The cardioprotective effects of regular exercise are, to some extent at least, related to its effects on plasma lipoprotein profile: exercise lowers plasma TG concentration, raises plasma HDL cholesterol concentration, and possibly also reduces total and LDL cholesterol concentrations (53). The hypotriglyceridemic effect appears to be the most powerful and was first recognized four decades ago (26). The immediate effect of exercise on plasma TG concentration remains controversial: some investigators report a decrease during and immediately after exercise (39), whereas others show no change (3,7,9). However, there is compelling evidence for a "delayedonset" hypo...

High-fat (HF) diets can induce insulin resistance (IR) by altering skeletal muscle lipid metabolism. An imbalance between fatty acid (FA) uptake and oxidation results in intramuscular lipid accumulation, which can impair the insulin-signaling cascade. Adiponectin (Ad) is an insulin-sensitizing adipokine known to stimulate skeletal muscle FA oxidation and reduce lipid accumulation. Evidence of Ad resistance has been shown in obesity and following chronic HF feeding and may contribute to lipid accumulation observed in these conditions. Whether Ad resistance precedes and is associated with the development of IR is unknown. We conducted a time course HF feeding trial for 3 days, 2 wk, or 4 wk to determine the onset of Ad resistance and identify the ensuing changes in lipid metabolism and insulin signaling leading to IR in skeletal muscle. Ad stimulated FA oxidation (+28%, P < or = 0.05) and acetyl-CoA carboxylase phosphorylation (+34%, P < or = 0.05) in control animals but failed to do so in any HF-fed group (i.e., as early as 3 days). By 2 wk, plasma membrane FA transporters and intramuscular diacylglycerol (DAG) and ceramide were increased, and insulin-stimulated phosphorylation of both protein kinase B and protein kinase B substrate 160 was blunted compared with control animals. After 4 wk of HF feeding, maximal insulin-stimulated glucose transport was impaired compared with control. Taken together, our results demonstrate that an early loss of Ad's stimulatory effect on FA oxidation precedes an increase in plasmalemmal FA transporters and the accumulation of intramuscular DAG and ceramide, blunted insulin signaling, and ultimately impaired maximal insulin-stimulated glucose transport in skeletal muscle induced by HF diets.