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.
OBJECTIVE-Heat treatment and overexpression of heat shock protein 72 (HSP72) have been shown to protect against high-fat diet-induced insulin resistance, but little is known about the underlying mechanism or the target tissue of HSP action. The purpose of this study is to determine whether in vivo heat treatment can prevent skeletal muscle insulin resistance. RESEARCH DESIGN AND METHODS-MaleWistar rats were fed a high-fat diet (60% calories from fat) for 12 weeks and received a lower-body heat treatment (41°C for 20 min) once per week.RESULTS-Our results show that heat treatment shifts the metabolic characteristics of rats on a high-fat diet toward those on a standard diet. Heat treatment improved glucose tolerance, restored insulin-stimulated glucose transport, and increased insulin signaling in soleus and extensor digitorum longus (EDL) muscles from rats fed a high-fat diet. Heat treatment resulted in decreased activation of Jun NH 2 -terminal kinase (JNK) and inhibitor of B kinase (IKK-), stress kinases implicated in insulin resistance, and upregulation of HSP72 and HSP25, proteins previously shown to inhibit JNK and IKK- activation, respectively. Mitochondrial citrate synthase and cytochrome oxidase activity decreased slightly with the high-fat diet, but heat treatment restored these activities. Data from L6 cells suggest that one bout of heat treatment increases mitochondrial oxygen consumption and fatty acid oxidation.CONCLUSIONS-Our results indicate that heat treatment protects skeletal muscle from high-fat diet-induced insulin resistance and provide strong evidence that HSP induction in skeletal muscle could be a potential therapeutic treatment for obesityinduced insulin resistance. Diabetes 58:567-578, 2009
In the present study, myosin heavy chain (MHC) content per half sarcomere, an estimate of the number of cross bridges available for force generation, was determined in rat diaphragm muscle (Dia(m)) fibers expressing different MHC isoforms. We hypothesize that fiber-type differences in maximum specific force [force per cross-sectional area (CSA)] reflect the number of cross bridges present per CSA. Studies were performed on single, Triton X-100-permeabilized rat Dia(m) fibers. Maximum specific force was determined by activation of single Dia(m) fibers in the presence of a high-calcium solution (pCa, -log Ca(2+) concentration of 4.0). SDS-PAGE and Western blot analyses were used to determine MHC isoform composition and MHC content per half sarcomere. Differences in maximum specific force across fast MHC isoforms were eliminated when controlled for half-sarcomere MHC content. However, the force produced by slow fibers remained below that of fast fibers when normalized for the number of cross bridges available. On the basis of these results, the lower force produced by slow fibers may be due to less force per cross bridge compared with fast fibers.
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.
/calmodulindependent protein kinase (CAMK)-dependent signaling pathways. However, recent studies provide evidence suggesting that contraction-stimulated glucose transport in slow-twitch skeletal muscle is mediated through an AMPK-independent pathway. The purpose of the present study was to test the hypothesis that contraction-stimulated glucose transport in rat slow-twitch soleus muscle is mediated by an AMPK-independent/Ca 2ϩ -dependent pathway. Caffeine, a sarcoplasmic reticulum (SR) Ca 2ϩ -releasing agent, at a concentration that does not cause muscle contractions or decreases in high-energy phosphates, led to an ϳ2-fold increase in 2-deoxyglucose (2-DG) uptake in isolated split soleus muscles. This increase in glucose transport was prevented by the SR calcium channel blocker dantrolene and the CAMK inhibitor KN93. Conversely, 5-aminoimidazole-4-carboxamide-1--D-ribofuranoside (AICAR), an AMPK activator, had no effect on 2-DG uptake in isolated split soleus muscles yet resulted in an ϳ2-fold increase in the phosphorylation of AMPK and its downstream substrate acetyl-CoA carboxylase. The hypoxia-induced increase in 2-DG uptake was prevented by dantrolene and KN93, whereas hypoxia-stimulated phosphorylation of AMPK was unaltered by these agents. Tetanic muscle contractions resulted in an ϳ3.5-fold increase in 2-DG uptake that was prevented by KN93, which did not prevent AMPK phosphorylation. Taken in concert, our results provide evidence that hypoxia-and contraction-stimulated glucose transport is mediated entirely through a Ca 2ϩ -dependent mechanism in rat slowtwitch muscle. in vitro; rodent; metabolism; exercise EXERCISE AND INSULIN both stimulate glucose transport, their maximal effects on muscle glucose transport are additive, and hypoxia appears to mimic this effect of exercise (11). Numerous studies have provided evidence that 5Ј-AMP-activated protein kinase (AMPK) is involved in mediating the stimulation of muscle glucose uptake by contractions and hypoxia (3, 8, 16, 18 -20). However, the activation of AMPK accounts for only part of the increase in glucose transport induced by muscle contractions. This is evidenced by the finding of Mu et al. (19) that the expression of a dominant negative AMPK mutant resulted in only a modest decrease in contractionstimulated glucose transport in mouse skeletal muscle and by the more recent report from Richter's group (14) that AMPK␣1 or -␣2 knockout mice displayed normal increases in contraction-mediated muscle glucose transport. Furthermore, increasing cytosolic Ca 2ϩ concentration to a level too low to cause muscle contractions or a decrease in high-energy phosphates (ϳP) induces an increase in muscle glucose transport, providing evidence that the increases in cytosolic Ca 2ϩ during excitation-contraction coupling are involved in mediating the stimulation of glucose transport by contractions (25).In support of this possibility, we found that the increases in glucose transport induced by a subcontraction concentration of the sarcoplasmic reticulum (SR) Ca 2ϩ -rel...
The present study examined Ca(2+) sensitivity of diaphragm muscle (Dia(m)) fibers expressing different myosin heavy chain (MHC) isoforms. We hypothesized that Dia(m) fibers expressing the MHC(slow) isoform have greater Ca(2+) sensitivity than fibers expressing fast MHC isoforms and that this fiber-type difference in Ca(2+) sensitivity reflects the isoform composition of the troponin (Tn) complex (TnC, TnT, and TnI). Studies were performed in single Triton-X-permeabilized Dia(m) fibers. The Ca(2+) concentration at which 50% maximal force was generated (pCa(50)) was determined for each fiber. SDS-PAGE and Western analyses were used to determine the MHC and Tn isoform composition of single fibers. The pCa(50) for Dia(m) fibers expressing MHC(slow) was significantly greater than that of fibers expressing fast MHC isoforms, and this greater Ca(2+) sensitivity was associated with expression of slow isoforms of the Tn complex. However, some Dia(m) fibers expressing MHC(slow) contained the fast TnC isoform. These results suggest that the combination of TnT, TnI, and TnC isoforms may determine Ca(2+) sensitivity in Dia(m) fibers.
(PD), the clinical literature remains equivocal. We, therefore, sought to address the relationship between insulin resistance and nigrostriatal dopamine (DA) in a preclinical animal model. High-fat feeding in rodents is an established model of insulin resistance, characterized by increased adiposity, systemic oxidative stress, and hyperglycemia. We subjected rats to a normal chow or high-fat diet for 5 wk before infusing 6-hydroxydopamine (6-OHDA) into the medial forebrain bundle. Our goal was to determine whether a high-fat diet and the resulting peripheral insulin resistance would exacerbate 6-OHDA-induced nigrostriatal DA depletion. Prior to 6-OHDA infusion, animals on the high-fat diet exhibited greater body weight, increased adiposity, and impaired glucose tolerance. Two weeks after 6-OHDA, locomotor activity was tested, and brain and muscle tissue was harvested. Locomotor activity did not differ between the groups nor did cholesterol levels or measures of muscle atrophy. High-fat-fed animals exhibited higher homeostatic model assessment of insulin resistance (HOMA-IR) values and attenuated insulin-stimulated glucose uptake in fast-twitch muscle, indicating decreased insulin sensitivity. Animals in the high-fat group also exhibited greater DA depletion in the substantia nigra and the striatum, which correlated with HOMA-IR and adiposity. Decreased phosphorylation of HSP27 and degradation of IB␣ in the substantia nigra indicate increased tissue oxidative stress. These findings support the hypothesis that a diet high in fat and the resulting insulin resistance may lower the threshold for developing PD, at least following DA-specific toxin exposure. dopamine; diabetes; substantia nigra; striatum; insulin resistance CLINICAL STUDIES SUGGEST A LINK between type 2 diabetes (T2D) and Parkinson's disease (PD) (30,46), and between fat intake or adiposity and PD (1, 31, 34). Moreover, it was reported over 40 years ago that greater than 50% of PD patients exhibit abnormal glucose tolerance (4, 10) or diabetes (36). Despite this information, very little is known regarding the relationship of these diseases and the impact of comorbidity on their pathogenesis. By 2025, T2D is estimated to impact 300 million individuals (47), with the elderly at greatest risk (54), the population also at greatest risk for neurodegenerative diseases like PD. For these reasons, understanding the potential for T2D, obesity, high dietary fat intake, and insulin resistance to contribute to PD is critical. Although the exact cause of PD is unknown, various environmental factors such as aging, diet, and environmental toxin exposure have been implicated in contributing to its development (29, 34, 51). The idea that "multiple hits" play a role in PD degeneration is supported by the fact that 80% of dopamine (DA)-producing neurons must be lost for symptoms to appear (50). While diabetes and PD do not invariably coincide, several studies suggest that obesity may potentiate neuronal dysfunction or even neurodegeneration (reviewed in Ref. 11). High-f...
Clinical studies have indicated a link between Parkinson's disease (PD) and Type 2 Diabetes. Although preclinical studies have examined the effect of high-fat feeding on dopamine function in brain reward pathways, the effect of diet on neurotransmission in the nigrostriatal pathway, which is affected in PD and parkinsonism, is less clear. We hypothesized that a high-fat diet, which models early-stage Type 2 Diabetes, would disrupt nigrostriatal dopamine function in young adult Fischer 344 rats. Rats were fed a high fat diet (60% calories from fat) or a normal chow diet for 12 weeks. High fat-fed animals were insulin resistant compared to chow-fed controls. Potassium-evoked dopamine release and dopamine clearance were measured in the striatum using in vivo electrochemistry. Dopamine release was attenuated and dopamine clearance was diminished in the high-fat diet group compared to chow-fed rats. Magnetic resonance imaging indicated increased iron deposition in the substantia nigra of the high fat group. This finding was supported by alterations in the expression of several proteins involved in iron metabolism in the substantia nigra in this group compared to chow-fed animals. The diet-induced systemic and basal ganglia-specific changes may play a role in the observed impairment of nigrostriatal dopamine function.
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