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Heat stress (HS) stimulates heat shock protein (HSP) 72 mRNA expression, and the period after an increase in HSP72 protein is characterized by enhanced glucose metabolism in skeletal muscle. We have hypothesized that, prior to an increase in the level of HSP72 protein, HS activates glucose metabolism by acutely stimulating 5′-AMP-activated protein kinase (AMPK). Rat epitrochlearis muscle was isolated and incubated either with or without HS (42°C) for 10 and 30 min. HS for 30 min led to an increase in the level of Hspa1a and Hspa1b mRNA but did not change the amount of HSP72 protein. However, HS for both 10 and 30 min led to a significant increase in the rate of 3-O-methyl-d-glucose (3MG) transport, and the stimulatory effect of 3MG transport was completely blocked by cytochalasin B. HS-stimulated 3MG transport was also inhibited by dorsomorphin but not by wortmannin. HS led to a decrease in the concentration of ATP, phosphocreatine, and glycogen, to an increase in the level of phosphorylation of AMPKα Thr172, and to an increase in the activity of both AMPKα1 and AMPKα2. HS did not affect the phosphorylation status of insulin receptor signaling or Ca2+/calmodulin-dependent protein kinase II. These results suggest that HS acts as a rapid stimulator of insulin-independent glucose transport, at least in part by stimulating AMPK via decreased energy status. Although further research is warranted, heat treatment of skeletal muscle might be a promising method to promote glucose metabolism acutely.

Physical exercise has potent therapeutic and preventive effects against metabolic disorders. A number of studies have suggested that 5'-AMP-activated protein kinase (AMPK) plays a pivotal role in regulating carbohydrate and lipid metabolism in contracting skeletal muscles, while several genetically manipulated animal models revealed the significance of AMPK-independent pathways. To elucidate significance of AMPK and AMPK-independent signals in contracting skeletal muscles, we conducted a metabolomic analysis that compared the metabolic effects of 5-aminoimidazole-4-carboxamide-1-β-D-ribonucleoside (AICAR) stimulation with the electrical contraction ex vivo in isolated rat epitrochlearis muscles, in which both α1- and α2-isoforms of AMPK and glucose uptake were equally activated. The metabolomic analysis using capillary electrophoresis time-of-flight mass spectrometry detected 184 peaks and successfully annotated 132 small molecules. AICAR stimulation exhibited high similarity to the electrical contraction in overall metabolites. Principal component analysis (PCA) demonstrated that the major principal component characterized common effects whereas the minor principal component distinguished the difference. PCA and a factor analysis suggested a substantial change in redox status as a result of AMPK activation. We also found a decrease in reduced glutathione levels in both AICAR-stimulated and contracting muscles. The muscle contraction-evoked influences related to the metabolism of amino acids, in particular, aspartate, alanine, or lysine, are supposed to be independent of AMPK activation. Our results substantiate the significance of AMPK activation in contracting skeletal muscles and provide novel evidence that AICAR stimulation closely mimics the metabolomic changes in the contracting skeletal muscles.

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