Exercise training improves muscle insulin resistance but not insulin receptor signaling in obese Zucker rats.

Exercise training improves skeletal muscle insulin sensitivity in the obese Zucker rat. The purpose of this study was to investigate whether the improvement in insulin action in response to exercise training is associated with enhanced insulin receptor signaling. Obese Zucker rats were trained for 7 wk and studied by using the hindlimb-perfusion technique 24 h, 96 h, or 7 days after their last exercise training bout. Insulin-stimulated glucose uptake (traced with 2-deoxyglucose) was significantly reduced in untrained obese Zucker rats compared with lean controls (2.2 +/- 0.17 vs. 5.4 +/- 0.46 micromol x g(-1) x h(-1)). Glucose uptake was normalized 24 h after the last exercise bout (4.9 +/- 0.41 micromol x g(-1) x h(-1)) and remained significantly elevated above the untrained obese Zucker rats for 7 days. However, exercise training did not increase insulin receptor or insulin receptor substrate-1 (IRS-1) tyrosine phosphorylation, phosphatidylinositol 3-kinase (PI3-kinase) activity associated with IRS-1 or tyrosine phosphorylated immunoprecipitates, or Akt serine phosphorylation. These results are consistent with the hypothesis that, in obese Zucker rats, adaptations occur during training that lead to improved insulin-stimulated muscle glucose uptake without affecting insulin receptor signaling through the PI3-kinase pathway.     

Acute and chronic leptin treatment mediate contrasting effects on signaling,glucose uptake,and GLUT4 translocation in L6-GLUT4myc myotubes

We have previously shown that in L6-GLUT4myc rat skeletal muscle cells, acute treatment with leptin reduced insulin-stimulated glucose uptake without altering insulin-stimulated GLUT4 translocation. In contrast, we show here that the ability of leptin to increase phosphorylation of its receptor and to reduce insulin-stimulated glucose uptake was lost in cells that were continuously exposed to leptin for 24 h. This desensitization correlated with an increase in expression of suppressor of cytokine signaling-3 (SOCS-3). Time course analysis demonstrated that the transition from acute to chronic effects of leptin occurs after 2 h. The desensitization of leptin action at 24 h was not reversed by 30 min washout period prior to re-exposing cells to leptin. However, despite insulin-stimulated glucose uptake being unaffected upon 24 h preincubation with leptin, a small but significant decrease (37%) in insulin-stimulated GLUT4 translocation and phosphorylation of Akt on T308 was detected. Insulin-stimulated phosphorylation of Akt on S473 or of p38 MAPK were unaffected. These results suggest that the chronic leptin treatment leads to desensitization of leptin signaling yet can simultaneously decrease the ability of insulin to phosphorylate Akt on T308 and translocate GLUT4. However, this does not manifest as a reduction in total glucose uptake into L6 myotubes.     

Reduced insulin-stimulated glucose transport in denervated muscle is associated with impaired Akt-alpha activation

Insulin signaling was examined in muscle made insulin resistant by short-term (24-h) denervation. Insulin-stimulated glucose transport in vitro was reduced by 28% (P < 0.05) in denervated muscle (DEN). In control muscle (SHAM), insulin increased levels of surface-detectable GLUT-4 (i.e., translocated GLUT-4) 1.8-fold (P < 0.05), whereas DEN surface GLUT-4 was not increased by insulin (P > 0.05). Insulin treatment in vivo induced a rapid appearance of phospho[Ser(473)]Akt-alpha in SHAM 3 min after insulin injection. In DEN, phospho[Ser(473)]Akt-alpha also appeared at 3 min, but Ser(473)-phosphorylated Akt-alpha was 36% lower than in SHAM (P < 0. 05). In addition, total Akt-alpha protein in DEN was 37% lower than in SHAM (P < 0.05). Akt-alpha kinase activity was lower in DEN at two insulin levels tested: 0.1 U insulin/rat (-22%, P < 0.05) and 1 U insulin/rat (-26%, P < 0.01). These data indicate that short-term (24-h) denervation, which lowers insulin-stimulated glucose transport, is associated with decreased Akt-alpha activation and impaired insulin-stimulated GLUT-4 appearance at the muscle surface.     

Exercise reverses high-fat diet-induced impairments on compartmentalization and activation of components of the insulin-signaling cascade in skeletal muscle

The aims of this investigation were 1) to determine whether endurance exercise training could reverse impairments in insulin-stimulated compartmentalization and/or activation of aPKCzeta/lambda and Akt2 in skeletal muscle from high-fat-fed rodents and 2) to assess whether the PPARgamma agonist rosiglitazone could reverse impairments in skeletal muscle insulin signaling typically observed after high-fat feeding. Sprague-Dawley rats were placed on chow (NORCON, n = 16) or high-fat (n = 64) diets for 4 wk. During a subsequent 4-wk experimental period, high-fat-fed rats were allocated (n = 16/group) to either sedentary control (HFC), exercise training (HFX), rosiglitazone treatment (HFRSG), or a combination of both exercise training and rosiglitazone (HFRX). Following the 4-wk experimental period, animals underwent hindlimb perfusions. Insulin-stimulated plasma membrane-associated aPKCzeta and -lambda protein concentration, aPKCzeta/lambda activity, GLUT4 protein concentration, cytosolic Akt2, and aPKCzeta/lambda activities were reduced (P < 0.05) in HFC compared with NORCON. Cytosolic Akt2, aPKCzeta, and aPKClambda protein concentrations were not affected in HFC compared with NORCON. Exercise training reversed the deleterious effects of the high-fat diet such that insulin-stimulated compartmentalization and activation of components of the insulin-signaling cascade in HFX were normalized to NORCON. High-fat diet-induced impairments to skeletal muscle glucose metabolism were not reversed by rosiglitazone administration, nor did rosiglitazone augment the effect of exercise. Our findings indicate that chronic exercise training, but not rosiglitazone, reverses high-fat diet induced impairments in compartmentalization and activation of components of the insulin-signaling cascade in skeletal muscle.     

Exercise does not alter subcellular localization, but increases phosphorylation of insulin-signaling proteins in human skeletal muscle

The subcellular localization of insulin signaling proteins is altered by various stimuli such as insulin, insulin-like growth factor I, and oxidative stress and is thought to be an important mechanism that can influence intracellular signal transduction and cellular function. This study examined the possibility that exercise may also alter the subcellular localization of insulin signaling proteins in human skeletal muscle. Nine untrained males performed 60 min of cycling exercise (approximately 67% peak pulmonary O2 uptake). Muscle biopsies were sampled at rest, immediately after exercise, and 3 h postexercise. Muscle was fractionated by centrifugation into the following crude fractions: cytosolic, nuclear, and a high-speed pellet containing membrane and cytoskeletal components. Fractions were analyzed for protein content of insulin receptor, insulin receptor substrate (IRS)-1 and -2, p85 subunit of phosphatidylinositol 3-kinase, Akt, and glycogen synthase kinase-3 (GSK-3). There was no significant change in the protein content of the insulin signaling proteins in any of the crude fractions after exercise or 3 h postexercise. Exercise had no significant effect on the phosphorylation of IRS-1 Tyr612 in any of the fractions. In contrast, exercise increased (P < 0.05) the phosphorylation of Akt Ser473 and GSK-3alpha/beta Ser9/21 in the cytosolic fraction only. In conclusion, exercise can increase phosphorylation of downstream insulin signaling proteins specifically in the cytosolic fraction but does not result in changes in the subcellular localization of insulin signaling proteins in human skeletal muscle. Change in the subcellular protein localization is therefore an unlikely mechanism to influence signal transduction pathways and cellular function in skeletal muscle after exercise.     

Increased submaximal insulin-stimulated glucose uptake in mouse skeletal muscle after treadmill exercise.

The primary purpose of this study was to determine the effect of prior exercise on insulin-stimulated glucose uptake with physiological insulin in isolated muscles of mice. Male C57BL/6 mice completed a 60-min treadmill exercise protocol or were sedentary. Paired epitrochlearis, soleus, and extensor digitorum longus (EDL) muscles were incubated with [3H]-2-deoxyglucose without or with insulin (60 microU/ml) to measure glucose uptake. Insulin-stimulated glucose uptake for paired muscles was calculated by subtracting glucose uptake without insulin from glucose uptake with insulin. Muscles from other mice were assessed for glycogen and AMPK Thr172 phosphorylation. Exercised vs. sedentary mice had decreased glycogen in epitrochlearis (48%, P < 0.001), soleus (51%, P < 0.001), and EDL (41%, P < 0.01) and increased AMPK Thr172 phosphorylation (P < 0.05) in epitrochlearis (1.7-fold), soleus (2.0-fold), and EDL (1.4-fold). Insulin-independent glucose uptake was increased 30 min postexercise vs. sedentary in the epitrochlearis (1.2-fold, P < 0.001), soleus (1.4-fold, P < 0.05), and EDL (1.3-fold, P < 0.01). Insulin-stimulated glucose uptake was increased (P < 0.05) approximately 85 min after exercise in the epitrochlearis (sedentary: 0.266 +/- 0.045 micromol x g(-1) x 15 min(-1); exercised: 0.414 +/- 0.051) and soleus (sedentary: 0.102 +/- 0.049; exercised: 0.347 +/- 0.098) but not in the EDL. Akt Ser473 and Akt Thr308 phosphorylation for insulin-stimulated muscles did not differ in exercised vs. sedentary. These results demonstrate enhanced submaximal insulin-stimulated glucose uptake in the epitrochlearis and soleus of mice 85 min postexercise and suggest that it will be feasible to probe the mechanism of enhanced postexercise insulin sensitivity by using genetically modified mice.     

Short-Term Intensified Cycle Training Alters Acute and Chronic Responses of PGC1α and Cytochrome C Oxidase IV to Exercise in Human Skeletal Muscle

Reduced activation of exercise responsive signalling pathways have been reported in response to acute exercise after training; however little is known about the adaptive responses of the mitochondria. Accordingly, we investigated changes in mitochondrial gene expression and protein abundance in response to the same acute exercise before and after 10-d of intensive cycle training. Nine untrained, healthy participants (mean±SD; VO_2peak 44.1±17.6 ml/kg/min) performed a 60 min bout of cycling exercise at 164±18 W (72% of pre-training VO_2peak). Muscle biopsies were obtained from the vastus lateralis muscle at rest, immediately and 3 h after exercise. The participants then underwent 10-d of cycle training which included four high-intensity interval training sessions (6×5 min; 90–100% VO_2peak) and six prolonged moderate-intensity sessions (45–90 min; 75% VO_2peak). Participants repeated the pre-training exercise trial at the same absolute work load (64% of pre-training VO_2peak). Muscle PGC1-α mRNA expression was attenuated as it increased by 11- and 4- fold (P<0.001) after exercise pre- and post-training, respectively. PGC1-α protein expression increased 1.5 fold (P<0.05) in response to exercise pre-training with no further increases after the post-training exercise bout. RIP140 protein abundance was responsive to acute exercise only (P<0.01). COXIV mRNA (1.6 fold; P<0.01) and COXIV protein expression (1.5 fold; P<0.05) were increased by training but COXIV protein expression was decreased (20%; P<0.01) by acute exercise pre- and post-training. These findings demonstrate that short-term intensified training promotes increased mitochondrial gene expression and protein abundance. Furthermore, acute indicators of exercise-induced mitochondrial adaptation appear to be blunted in response to exercise at the same absolute intensity following short-term training.     

Insulin-stimulated hydrogen peroxide reversibly inhibits protein-tyrosine phosphatase 1b in vivo and enhances the early insulin action cascade

The insulin signaling pathway is activated by tyrosine phosphorylation of the insulin receptor and key post-receptor substrate proteins and balanced by the action of specific protein-tyrosine phosphatases (PTPases). PTPase activity, in turn, is highly regulated in vivo by oxidation/reduction reactions involving the cysteine thiol moiety required for catalysis. Here we show that insulin stimulation generates a burst of intracellular H(2)O(2) in insulin-sensitive hepatoma and adipose cells that is associated with reversible oxidative inhibition of up to 62% of overall cellular PTPase activity, as measured by a novel method using strictly anaerobic conditions. The specific activity of immunoprecipitated PTP1B, a PTPase homolog implicated in the regulation of insulin signaling, was also strongly inhibited by up to 88% following insulin stimulation. Catalase pretreatment abolished the insulin-stimulated production of H(2)O(2) as well as the inhibition of cellular PTPases, including PTP1B, and was associated with reduced insulin-stimulated tyrosine phosphorylation of its receptor and high M(r) insulin receptor substrate (IRS) proteins. These data provide compelling new evidence for a redox signal that enhances the early insulin-stimulated cascade of tyrosine phosphorylation by oxidative inactivation of PTP1B and possibly other tyrosine phosphatases.     

Antioxidant enzyme systems in rat liver and skeletal muscle. Influences of selenium deficiency, chronic training, and acute exercise

The influences of selenium deficiency (Se-D), chronic training, and an acute bout of exercise on hepatic and skeletal muscle antioxidant enzymes, i.e., superoxide dismutase (SOD), catalase, and glutathione peroxidase (GPX), as well as glutathione S-transferase (GST) and tissue lipid peroxidation, were investigated in post-weaning male Sprague-Dawley rats. Se-D per se depleted GPX in both liver and skeletal muscle but had no effect on SOD or catalase activity. One hour of treadmill running (20 m/min, 0% grade and 27 m/min, 15% grade for untrained and trained rats, respectively) significantly elevated hepatic catalase and cytosolic SOD activity; more prominent activations were found in the Se-D or untrained rats, whereas skeletal muscle antioxidant enzymes were little affected. Ten weeks of training (1 h/day, 5 days/week at 27 m/min, 15% grade) increased hepatic mitochondrial SOD by 23% (P less than 0.05) in Se-D rats. Both hepatic mitochondrial and cytosolic GPX were decreased by training whereas GPX was increased twofold in skeletal muscle mitochondria. Se-independent GPX was elevated by training only in the skeletal muscle mitochondria of Se-D rats. Lipid peroxidation (malondialdehyde formation) was increased by an acute bout of exercise in hepatic mitochondria of the untrained rats and in skeletal muscle mitochondria of the Se-D rats. These data indicate that antioxidant enzymes in liver and skeletal muscle are capable of adapting to selenium deficiency and exercise to minimize oxidative injury caused by free radicals.     

Activation pattern of MAPK signaling in the hearts of trained and untrained rats following a single bout of exercise.

Since exercise training causes cardiac hypertrophy and a single bout induces mechanical stress to the heart, the present study aimed to characterize the activation patterns of multiple MAPK signaling pathways in the heart after a single bout of exercise or chronic exercises. The hearts of untrained rats received 5, 15, and 30 min of treadmill running exercise (Ex5 to Ex30) and rested for 0.5, 1, 3, 6, 12, and 24 h (PostEx0.5 to PostEx24) before subjecting them to the following different experiments. Activation of MAPKs (ERK, JNK, and p38) and MAPKKs (MEK1/2, SEK, and MKK3/6) increased immediately after acute exercise in a time-dependent manner, with ERK, JNK, and p38 peaking at Ex15, Ex15, and Ex30, respectively. Expression of immediate early genes (c-fos, c-jun, and c-myc) was augmented and activator protein-1 DNA binding activity was enhanced in untrained rats immediately after a single bout of exercise. The elevated levels of MAPKs declined to the resting levels within 24 h after exercise. In another set of experiments, following 4, 8, and 12 wk of exercise training, the rats exhibited significant cardiac hypertrophy by week 12. Activation of MAPKs in the 4-wk-trained rats increased after a 30-min single bout of exercise but decreased in the 8-wk group. Finally, the activity of MAPKs signaling in the 12-wk-trained rats exposed to an acute bout of exercise was unaltered. We conclude that exercise induces the activation of multiple MAPK (ERK, JNK, and p38) pathways in the heart, an effect that gradually declines with the development of exercise-induced cardiac hypertrophy.     

Protein tyrosine phosphatase regulation in fibroblasts from patients with an insulin receptor gene mutation

Tyrosine phosphorylation of the insulin receptor is the initial event following receptor binding to insulin, and it induces further tyrosine phosphorylation of various intracellular molecules. This signaling is countered by protein tyrosine phosphatases (PTPases), which reportedly are associated with insulin resistance that can be reduced by regulation of PTPases. Protein tyrosine phosphatase 1B (PTP1B) and leukocyte antigen-related PTPase (LAR) are the PTPases implicated most frequently in insulin resistance and diabetes mellitus. Here, we show that PTP1B and LAR are expressed in human fibroblasts, and we examine the regulation of PTPase activity in fibroblasts from patients with an insulin receptor gene mutation as an in vitro model of insulin resistance. Total PTPase activity was significantly lower in the cytosolic and membrane fractions of fibroblasts with mutations compared with controls (p<0.05). Insulin stimulation of fibroblasts with mutations resulted in a significantly smaller increase in PTP1B activity compared with stimulation of wild-type fibroblasts (p<0.05). This indicates that insulin receptor gene mutations blunt increases in PTPase activity in response to insulin, possibly via a negative feedback mechanism. Our data suggest that the PTPase activity in patients with insulin receptor gene mutation and severe insulin resistance may differ from that in ordinary type 2 diabetes.     

Voluntary exercise training enhances glucose transport but not insulin signaling capacity in muscle of hypertensive TG(mREN2)27 rats.

Male heterozygous TG(mREN2)27 rats (TGR) overexpress a murine renin transgene, display marked hypertension, and have insulin resistance of skeletal muscle glucose transport and insulin signaling. We have shown previously that voluntary exercise training by TGR improves insulin-mediated skeletal muscle glucose transport (Kinnick TR, Youngblood EB, O'Keefe MP, Saengsirisuwan V, Teachey MK, and Henriksen EJ. J Appl Physiol 93: 805-812, 2002). The present study evaluated whether this training-induced enhancement of muscle glucose transport is associated with upregulation of critical insulin signaling elements, including insulin receptor substrate-1 (IRS-1), phosphatidylinositol 3-kinase, Akt, and glycogen synthase kinase-3. TGR remained sedentary or ran spontaneously in activity wheels for 6 wk, averaging 7.1 +/- 0.8 km/day by the end of week 3 and 4.3 +/- 0.5 km/day over the final week of training. Exercise training reduced total abdominal fat by 20% (P < 0.05) in TGR runners (2.64 +/- 0.01% of body weight) compared with sedentary TGR controls (3.28 +/- 0.01%). Insulin-stimulated (2 mU/ml) glucose transport activity in soleus muscle was 36% greater in TGR runners compared with sedentary TGR controls. However, the protein expression and functionality of tyrosine phosphorylation of insulin receptor and IRS-1, IRS-1 associated with the p85 regulatory subunit of phosphatidylinositol 3-kinase, and Ser473 phosphorylation of Akt were not altered by exercise training. Only insulin-stimulated glycogen synthase kinase-3beta Ser9 phosphorylation was increased (22%) by exercise training. These results indicate that voluntary exercise training in TGR can enhance insulin-mediated glucose transport in skeletal muscle, as well as reduce total abdominal fat mass. However, this adaptive response in muscle occurs independently of modifications in the proximal elements of the insulin signaling cascade.     

Effects of high-fat diet and exercise training on intracellular glucose metabolism in rats

We examined the effects of high-fat diet (HFD) and exercise training on insulin-stimulated whole body glucose fluxes and several key steps of glucose metabolism in skeletal muscle. Rats were maintained for 3 wk on either low-fat (LFD) or high-fat diet with or without exercise training (swimming for 3 h per day). After the 3-wk diet/exercise treatments, animals underwent hyperinsulinemic euglycemic clamp experiments for measurements of insulin-stimulated whole body glucose fluxes. In addition, muscle samples were taken at the end of the clamps for measurements of glucose 6-phosphate (G-6-P) and GLUT-4 protein contents, hexokinase, and glycogen synthase (GS) activities. Insulin-stimulated glucose uptake was decreased by HFD and increased by exercise training (P < 0.01 for both). The opposite effects of HFD and exercise training on insulin-stimulated glucose uptake were associated with similar increases in muscle G-6-P levels (P < 0.05 for both). However, the increase in G-6-P level was accompanied by decreased GS activity without changes in GLUT-4 protein content and hexokinase activities in the HFD group. In contrast, the increase in G-6-P level in the exercise-trained group was accompanied by increased GLUT-4 protein content and hexokinase II (cytosolic) and GS activities. These results suggest that HFD and exercise training affect insulin sensitivity by acting predominantly on different steps of intracellular glucose metabolism. High-fat feeding appears to induce insulin resistance by affecting predominantly steps distal to G-6-P (e.g., glycolysis and glycogen synthesis). Exercise training affected multiple steps of glucose metabolism both proximal and distal to G-6-P. However, increased muscle G-6-P levels in the face of increased glucose metabolic fluxes suggest that the effect of exercise training is quantitatively more prominent on the steps proximal to G-6-P (i.e., glucose transport and phosphorylation).     

Effect of acute exercise and exercise training on VEGF splice variants in human skeletal muscle

The present study investigated the effect of an acute exercise bout on the mRNA response of vascular endothelial growth factor (VEGF) splice variants in untrained and trained human skeletal muscle. Seven habitually active young men performed one-legged knee-extensor exercise training at an intensity corresponding to approximately 70% of the maximal workload in an incremental test five times/week for 4 wk. Biopsies were obtained from the vastus lateralis muscle of the trained and untrained leg 40 h after the last training session. The subjects then performed 3 h of two-legged knee-extensor exercise, and biopsies were obtained from both legs after 0, 2, 6, and 24 h of recovery. Real-time PCR was used to examine the expression of VEGF mRNA containing exon 1 and 2 (all VEGF isoforms), exon 6 or exon 7, and VEGF(165) mRNA. Acute exercise induced an increase (P < 0.05) in total VEGF mRNA levels as well as VEGF(165) and VEGF splice variants containing exon 7 at 0, 2, and 6 h of recovery. The increase in VEGF mRNA was higher in the untrained than in the trained leg (P < 0.05). The results suggest that in human skeletal muscle, acute exercise increases total VEGF mRNA, an increase that appears to be explained mainly by an increase in VEGF(165) mRNA. Furthermore, 4 wk of training attenuated the exercise-induced response in skeletal muscle VEGF(165) mRNA.     

Overexpression of protein tyrosine phosphatase-alpha (PTP-alpha) but not PTP-kappa inhibits translocation of GLUT4 in rat adipose cells

Protein tyrosine phosphatases (PTPases) are likely to play important roles in insulin action. We recently demonstrated that the nontransmembrane PTPase PTP1B can act as a negative modulator of insulin-stimulated translocation of GLUT4. We now examine the role of PTP-alpha and PTP-kappa (two transmembrane PTPases) in this metabolic action of insulin. Rat adipose cells were transfected with either PTP-alpha or PTP-kappa and effects of these PTPases on the translocation of a cotransfected epitope-tagged GLUT4 were studied. Cells overexpressing wild-type PTP-alpha had significantly lower levels of cell surface GLUT4 in response to insulin and a threefold decrease in insulin sensitivity when compared with control cells expressing only tagged GLUT4. Co-overexpression of PTP-alpha and PTP1B did not have additive effects, suggesting that these PTPases share common substrates. Cells overexpressing either wild-type PTP-kappa or catalytically inactive mutants of PTP-alpha had dose-response curves similar to those of control cells. Since overexpression of PTP-alpha, but not PTP-kappa, had effects on translocation of GLUT4, our data suggest that PTPalpha may be a specific negative modulator of insulin-stimulated glucose transport.     

Myofibrillar disruption following acute concentric and eccentric resistance exercise in strength-trained men

We have previously quantified the extent of myofibrillar disruption which occurs following an acute bout of resistance exercise in untrained men, however the response of well-trained subjects is not known. We therefore recruited six strength-trained men, who ceased training for 5 days and then performed 8 sets of 8 uni-lateral repetitions, using a load equivalent to 80% of their concentric (Con) 1-repetition maximum. One arm performed only Con actions by lifting the weight and the other arm performed only eccentric actions (Ecc) by lowering it. Needle biopsy samples were obtained from biceps brachii of each arm approximately 21 h following exercise, and at baseline (i.e., after 5 days without training), and subsequently analyzed using electron microscopy to quantify myofibrillar disruption. A greater (P < or = 0.05) proportion of disrupted fibres was found in the Ecc arm (45 +/- 11%) compared with baseline values (4 +/- 2%), whereas fibre disruption in the Con arm (27 +/- 4%) was not different (P > 0.05) from baseline values. The proportion of disrupted fibres and the magnitude of disruption (quantified by sarcomere counting) was considerably less severe than previously observed in untrained subjects after an identical exercise bout. Mixed muscle protein synthesis, assessed from approximately 21-29 h post-exercise, was not different between the Con- and Ecc-exercised arms. We conclude that the Ecc phase of resistance exercise is most disruptive to skeletal muscle and that training attenuates the severity of this effect. Moreover, it appears that fibre disruption induced by habitual weightlifting exercise is essentially repaired after 5 days of inactivity in trained men.     

Brief calorie restriction increases Akt2 phosphorylation in insulin-stimulated rat skeletal muscle

Skeletal muscle insulin sensitivity improves with short-term reduction in calorie intake. The goal of this study was to evaluate changes in the abundance and phosphorylation of Akt1 and Akt2 as potential mechanisms for enhanced insulin action after 20 days of moderate calorie restriction [CR; 60% of ad libitum (AL) intake] in rat skeletal muscle. We also assessed changes in the abundance of SH2 domain-containing inositol phosphatase (SHIP2), a negative regulator of insulin signaling. Fisher 344 x Brown Norway rats were assigned to an AL control group or a CR treatment group for 20 days. Epitrochlearis muscles were dissected and incubated with or without insulin (500 microU/ml). Total Akt serine and threonine phosphorylation was significantly increased by 32 (P < 0.01) and 30% (P < 0.005) in insulin-stimulated muscles from CR vs. AL. Despite an increase in total Akt phosphorylation, there was no difference in Akt1 serine or Akt1 threonine phosphorylation between CR and AL insulin-treated muscles. However, there was a 30% decrease (P < 0.05) in Akt1 abundance for CR vs. AL. In contrast, there was no change in Akt2 protein abundance, and there was a 94% increase (P < 0.05) in Akt2 serine phosphorylation and an increase of 75% (P < 0.05) in Akt2 threonine phosphorylation of insulin-stimulated CR muscles compared with AL. There was no diet effect on SHIP2 abundance in skeletal muscle. These results suggest that, with brief CR, enhanced Akt2 phosphorylation may play a role in increasing insulin sensitivity in rat skeletal muscles.     

Effect of endurance and sprint exercise on the sensitivity of glucose metabolism to insulin in the epitrochlearis muscle of sedentary and trained rats

The effects of two types of acute exercise (1 h treadmill running at 20 m.min-1, or 6 x 10-s periods at 43 m.min-1, 0 degree inclination), as well as two training regimes (endurance and sprint) on the sensitivity of epitrochlearis muscle [fast twitch (FT) fibres] to insulin were measured in vitro in rats. The hormone concentration in the incubation medium producing the half maximal stimulation of lactate (la) production and glycogen synthesis was determined and used as an index of the muscle insulin sensitivity. A single period of moderate endurance as well as the sprint-type exercise increased the sensitivity of la production to insulin although the rate of la production enhanced markedly only after sprint exercise at 10 and 100 microU.ml-1 of insulin. These effects persisted for up to 2 h after the termination of exercise. Both types of exercise significantly decreased the muscle glycogen content, causing a moderate enhancement in the insulin-stimulated rates of glycogen synthesis in vitro for up to 2 h after exercise. However, a significant increase in the sensitivity of this process to insulin was found only in the muscle removed 0.25 h after the sprint effort. Training of the sprint and endurance types increased insulin-stimulated rates of glycolysis 24 h after the last period of exercise. The sensitivity of this process to insulin was also increased at this instant. Both types of training increased the basal and maximal rates of glycogen synthesis, as well as the sensitivity of this process to insulin at the 24th h following the last training session.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of tumor necrosis factor-α on the phosphorylation of tyrosine kinase receptors is associated with dynamic alterations in specific protein-tyrosine phosphatases

Tumor necrosis factor-α (TNF-α) can modulate the signalling capacity of tyrosine kinase receptors; in particular, TNF-α has been shown to mediate the insulin resistance associated with animal models of obesity and noninsulin-dependent diabetes mellitus. In order to determine whether the effects of TNF-α might involve alterations in the expression of specific protein-tyrosine phosphatases (PTPases) that have been implicated in the regulation of growth factor receptor signalling, KRC-7 rat hepatoma cells were treated with TNF-α, and changes in overall tissue PTPase activity and the abundance of three major hepatic PTPases (LAR, PTP1B, and SH-PTP2) were measured in addition to effects of TNF-α on ligand-stimulated autophosphorylation of insulin and epidermal growth factor (EGF) receptors and insulin-stimulated insulin receptor substrate-1 (IRS-1) phosphorylation. TNF-α caused a dose-dependent decrease in insulin-stimulated IRS-1 phosphorylation and EGF-stimulated receptor autophosphorylation to 47–50% of control. Overall PTPase activity in the cytosol fraction did not change with TNF-α treatment, and PTPase activity in the particulate fraction was decreased by 55–66%, demonstrating that increases in total cellular PTPase activity did not account for the observed alterations in receptor signalling. However, immunoblot analysis showed that TNF-α treatment resulted in a 2.5-fold increase in the abundance of SH-PTP2, a 49% decrease in the transmembrane PTPase LAR, and no evident change in the expression of PTP1B. These data suggest that at least part of the TNF-α effect on pathways of reversible tyrosine phosphorylation may be exerted through the dynamic modulation of the expression of specific PTPases. Since SH-PTP2 has been shown to interact directly with both the EGF receptor and IRS-1, increased abundance of this PTPase may mediate the TNF-α effect to inhibit signalling through these proteins. Furthermore, decreased abundance of the LAR PTPase, which has been implicated in the regulation of insulin receptor phosphorylation, may account for the less marked effect of TNF-α on the autophosphorylation state of the insulin receptor while postreceptor actions of insulin are inhibited. J. Cell. Biochem. 64:117–127. © 1997 Wiley-Liss, Inc.     

Response of muscle protein turnover to insulin after acute exercise and training.

To determine whether the enhanced insulin-sensitivity of glucose metabolism in muscle after acute exercise also extends to protein metabolism, untrained and exercise-trained rats were subjected to an acute bout of exercise, and the responses of protein synthesis and degradation to insulin were measured in epitrochlearis muscles in vitro. Acute exercise of both untrained and trained rats decreased protein synthesis in muscle in the absence or presence of insulin, but protein degradation was not altered. Exercise training alone had no effect on protein synthesis or degradation in muscle in the absence or presence of insulin. Acute exercise or training alone enhanced the sensitivities of both protein synthesis and degradation to insulin, but the enhanced insulin-sensitivities from training alone were not additive to those after acute exercise. These results indicate that: a decrease in protein synthesis is the primary change in muscle protein turnover after acute exercise and is not altered by prior exercise training, and the enhanced insulin-sensitivities of metabolism of both glucose and protein after either acute exercise or training suggest post-binding receptor events.