Intense exercise up-regulates Na+, K+ -ATPase isoform mRNA, but not protein expression in human skeletal muscle

Characterization of expression of, and consequently also the acute exercise effects on, Na+,K+-ATPase isoforms in human skeletal muscle remains incomplete and was therefore investigated. Fifteen healthy subjects (eight males, seven females) performed fatiguing, knee extensor exercise at 40% of their maximal work output per contraction. A vastus lateralis muscle biopsy was taken at rest, fatigue and 3 and 24 h postexercise, and analysed for Na+,K+-ATPase 1, 2, 3, ß1, ß2 and ß3 mRNA and crude homogenate protein expression, using Real-Time RT-PCR and immunoblotting, respectively. Each individual expressed gene transcripts and protein bands for each Na+,K+-ATPase isoform. Each isoform was also expressed in a primary human skeletal muscle cell culture. Intense exercise (352 ± 69 s; mean ±S.E.M.) immediately increased 3 and ß2 mRNA by 2.4- and 1.7-fold, respectively (P < 0.05), whilst 1 and 2 mRNA were increased by 2.5- and 3.5-fold at 24 h and 3 h postexercise, respectively (P < 0.05). No significant change occurred for ß1 and ß3 mRNA, reflecting variable time-dependent responses. When the average postexercise value was contrasted to rest, mRNA increased for 1, 2, 3, ß1, ß2 and ß3 isoforms, by 1.4-, 2.2-, 1.4-, 1.1-, 1.0- and 1.0-fold, respectively (P < 0.05). However, exercise did not alter the protein abundance of the 1–3 and ß1–ß3 isoforms. Thus, human skeletal muscle expresses each of the Na+,K+-ATPase 1, 2, 3, ß1, ß2 and ß3 isoforms, evidenced at both transcription and protein levels. Whilst brief exercise increased Na+,K+-ATPase isoform mRNA expression, there was no effect on isoform protein expression, suggesting that the exercise challenge was insufficient for muscle Na+,K+-ATPase up-regulation.

Depressed Na+-K+-ATPase activity in skeletal muscle at fatigue is correlated with increased Na+-K+-ATPase mRNA expression following intense exercise

We investigated whether depressed muscle Na⁺-K⁺-ATPase activity with exercise reflected a loss of Na⁺-K⁺-ATPase units, the time course of its recovery postexercise, and whether this depressed activity was related to increased Na⁺-K⁺-ATPase isoform gene expression. Fifteen subjects performed fatiguing, knee extensor exercise at ~40% maximal work output per contraction. A vastus lateralis muscle biopsy was taken at rest, fatigue, 3 h, and 24 h postexercise and analyzed for maximal Na⁺-K⁺-ATPase activity via 3-O-methylfluorescein phosphatase (3-O-MFPase) activity, Na⁺-K⁺-ATPase content via [3H]ouabain binding sites, and Na⁺-K⁺-ATPase α1-, α2-, α3-, ß1-, ß2- and ß3-isoform mRNA expression by real-time RT-PCR. Exercise [352 (SD 267) s] did not affect [³H]ouabain binding sites but decreased 3-O-MFPase activity by 10.7 (SD 8)% (P < 0.05), which had recovered by 3 h postexercise, without further change at 24 h. Exercise elevated α1-isoform mRNA by 1.5-fold at fatigue (P < 0.05). This increase was inversely correlated with the percent change in 3-O-MFPase activity from rest to fatigue (%Δ3-O-MFPaserest-fatigue) (r = –0.60, P < 0.05). The average postexercise (fatigue, 3 h, 24 h) {alpha}1-isoform mRNA was increased 1.4-fold (P < 0.05) and approached a significant inverse correlation with %Δ3-O-MFPaserest-fatigue (r = –0.56, P = 0.08). Exercise elevated α2-isoform mRNA at fatigue 2.5-fold (P < 0.05), which was inversely correlated with %Δ3-O-MFPaserest-fatigue (r = –0.60, P = 0.05). The average postexercise α2-isoform mRNA was increased 2.2-fold (P < 0.05) and was inversely correlated with the %Δ3-O-MFPaserest-fatigue (r = –0.68, P < 0.05). Nonsignificant correlations were found between %Δ3-O-MFPaserest-fatigue and other isoforms. Thus acute exercise transiently decreased Na^+-K⁺-ATPase activity, which was correlated with increased Na⁺-K⁺-ATPase gene expression. This suggests a possible signal-transduction role for depressed muscle Na⁺-K⁺-ATPase activity with exercise.

Effect of accurate exercise on uncoupling protein3 is a fat metabolism-mediated effect

Human and rodent uncoupling protein (UCP)3 mRNA is upregulated after acute exercise. Moreover, exercise increases plasma levels of free fatty acid (FFA), which are also known to upregulate UCP3. We investigated whether the upregulation of UCP3 after exercise is an effect of exercise per se or an effect of FFA levels or substrate oxidation. Seven healthy untrained men [age: 22.7 ± 0.6 yr; body mass index: 23.8 ± 1.0 kg/m²; maximal O₂ uptake (VO_{2 max}): 3,852 ± 211 ml/min] exercised at 50% VO_{2 max} for 2 h and then rested for 4 h. Muscle biopsies and blood samples were taken before and immediately after 2 h of exercise and 1 and 4 h in the postexercise period. To modulate plasma FFA levels and fat/glucose oxidation, the experiment was performed two times, one time with glucose ingestion and one time while fasting. UCP3 mRNA and UCP3 protein were determined by RT-competitive PCR and Western blot. In the fasted state, plasma FFA levels significantly increased (P < 0.0001) during exercise (293 ± 25 vs. 1,050 ± 127 μmol/l), whereas they were unchanged after glucose ingestion (335 ± 54 vs. 392 ± 74 µmol/l). Also, fat oxidation was higher after fasting (P < 0.05), whereas glucose oxidation was higher after glucose ingestion (P < 0.05). In the fasted state, UCP3L mRNA expression was increased significantly (P < 0.05) 4 h after exercise (4.6 ± 1.2 vs. 9.6 ± 3.3 amol/µg RNA). This increase in UCP3L mRNA expression was prevented by glucose ingestion. Acute exercise had no effect on UCP3 protein levels. In conclusion, we found that acute exercise had no direct effect on UCP3 mRNA expression. Abolishing the commonly observed increase in plasma FFA levels and/or fatty acid oxidation during and after exercise prevents the upregulation of UCP3 after acute exercise. Therefore, the previously observed increase in UCP3 expression appears to be an effect of prolonged elevation of plasma FFA levels and/or increased fatty acid oxidation.

NOS isoform-specific regulation of basal but not exercise-induced mitochondrial biogenesis in mouse skeletal muscle

Nitric oxide is a potential regulator of mitochondrial biogenesis. Therefore, we investigated if mice deficient in endothelial nitric oxide synthase (eNOS-/-) or neuronal NOS (nNOS-/-) have attenuated activation of skeletal muscle mitochondrial biogenesis in response to exercise. eNOS-/-, nNOS-/- and C57Bl6 (CON) mice (16.3 ± 0.2 weeks old) either remained in their cages (basal) or ran on a treadmill (16 m min-1, 5 grade) for 60 min (n = 8 per group) and were killed 6 h after exercise. Other eNOS-/-, nNOS-/- and CON mice exercise trained for 9 days (60 min per day) and were killed 24 h after the last bout of exercise training. eNOS-/- mice had significantly higher nNOS protein and nNOS-/- mice had significantly higher eNOS protein in the EDL, but not the soleus. The basal mitochondrial biogenesis markers NRF1, NRF2α and mtTFA mRNA were significantly (P< 0.05) higher in the soleus and EDL of nNOS-/- mice whilst basal citrate synthase activity was higher in the soleus and basal PGC-1α mRNA higher in the EDL. Also, eNOS-/- mice had significantly higher basal citrate synthase activity in the soleus but not the EDL. Acute exercise increased (P< 0.05) PGC-1α mRNA in soleus and EDL and NRF2α mRNA in the EDL to a similar extent in all genotypes. In addition, short-term exercise training significantly increased cytochrome c protein in all genotypes (P< 0.05) in the EDL. In conclusion, eNOS and nNOS are differentially involved in the basal regulation of mitochondrial biogenesis in skeletal muscle but are not critical for exercise-induced increases in mitochondrial biogenesis in skeletal muscle.

Exercise, muscle glycogen and insulin action

The aim of this thesis was to investigate the influence of muscle glycogen concentration on whole body insulin stimulated glucose uptake in humans and to examine the potential signalling mechanisms responsible for enhanced insulin action in the post exercise period. Untrained male subjects were conditioned to achieve a range of muscle glycogen concentrations via acute exercise or a combination of exercise and diet. The influence of muscle glycogen content on whole body insulin stimulated glucose uptake was determined via hyperinsulinaemic / euglycaemic clamps conducted at rest, 30 min after exercise or 24 hours after exercise. Muscle glycogen content did not influence insulin mediated glucose disposal either 30 min or 24 hrs after exercise when compared with basal. Conventional insulin signalling to muscle glucose uptake and signalling through the p38 MAPK cascade was also largely unaltered by glycogen concentration. Muscle glycogen synthesis was significantly increased in heavily but not moderately glycogen depleted muscle 30 min after exercise. Enhanced muscle glycogen synthesis occurred in line with a significant increase in insulin stimulated GSK-3 serine phosphorylation. This finding suggests that enhanced insulin sensitivity of muscle glycogen synthesis following glycogen depleting exercise may be mediated via a pathway involving alterations in insulin stimulated GSK-3 phosphorylation. In summary, whilst glycogen influences insulin mediated GSK-3 phosphorylation and glycogen synthesis, the findings of the present series of investigations suggest that the role of muscle glycogen in the process of insulin stimulated glucose uptake may not be as important as previously theorised.

Effect of resistance exercise contraction mode and protein supplementation on members of the STARS signalling pathway

The striated muscle activator of Rho signalling (STARS) pathway is suggested to provide a link between external stress responses and transcriptional regulation in muscle. However, the sensitivity of STARS signalling to different mechanical stresses has not been investigated. In a comparative study, we examined the regulation of the STARS signalling pathway in response to unilateral resistance exercise performed as either eccentric (ECC) or concentric (CONC) contractions as well as prolonged training; with and without whey protein supplementation. Skeletal muscle STARS, myocardian-related transcription factor-A (MRTF-A) and serum response factor (SRF) mRNA and protein, as well as muscle cross-sectional area and maximal voluntary contraction, were measured. A single-bout of exercise produced increases in STARS and SRF mRNA and decreases in MRTF-A mRNA with both ECC and CONC exercise, but with an enhanced response occurring following ECC exercise. A 31% increase in STARS protein was observed exclusively after CONC exercise (P < 0.001), while pSRF protein levels increased similarly by 48% with both CONC and ECC exercise (P < 0.001). Prolonged ECC and CONC training equally stimulated muscle hypertrophy and produced increases in MRTF-A protein of 125% and 99%, respectively (P < 0.001). No changes occurred for total SRF protein. There was no effect of whey protein supplementation. These results show that resistance exercise provides an acute stimulation of the STARS pathway that is contraction mode dependent. The responses to acute exercise were more pronounced than responses to accumulated training, suggesting that STARS signalling is primarily involved in the initial phase of exercise-induced muscle adaptations.

Can exercise positively influence the intervertebral disc?

To better understand what kinds of sports and exercise could be beneficial for the intervertebral disc (IVD), we performed a review to synthesise the literature on IVD adaptation with loading and exercise. The state of the literature did not permit a systematic review; therefore, we performed a narrative review. The majority of the available data come from cell or whole-disc loading models and animal exercise models. However, some studies have examined the impact of specific sports on IVD degeneration in humans and acute exercise on disc size. Based on the data available in the literature, loading types that are likely beneficial to the IVD are dynamic, axial, at slow to moderate movement speeds, and of a magnitude experienced in walking and jogging. Static loading, torsional loading, flexion with compression, rapid loading, high-impact loading and explosive tasks are likely detrimental for the IVD. Reduced physical activity and disuse appear to be detrimental for the IVD. We also consider the impact of genetics and the likelihood of a ‘critical period’ for the effect of exercise in IVD development. The current review summarises the literature to increase awareness amongst exercise, rehabilitation and ergonomic professionals regarding IVD health and provides recommendations on future directions in research.

Circulating leukocyte heat shock protein 70 (HSP70) and oxidative stress markers in rats after a bout of exhaustive exercise

A novel method to measure oxidative stress resulting from exhaustive exercise in rats is presented. In this new procedure we evaluated the erythrocyte antioxidant enzymes, catalase ( CAT) and glutathione reductase (GR), the plasma oxidative attack markers, reactive carbonyl derivatives (RCD) and thiobarbituric reactive substances (TBARS). Muscular tissue damage was evaluated by monitoring plasma creatine kinase (CK) and plasma taurine ( Tau) concentrations. Also, we monitored total sulphydryl groups (TSG) and uric acid (UA), and the level of the 70 kDa heat shock protein (HSP70) in leukocytes as a marker of oxidative stress. In the study we found a correspondence between erythrocyte CAT and GR activities and leukocyte HSP70 levels, principally 3 h after the acute exercise, and this suggested an integrated mechanism of antioxidant defense. The increase in levels of plasma Tau was coincident with the increasing plasma levels of CK and TBARS, principally after two hours of exercise. Thus tissue damage occurred before the expression of any anti-oxidant system markers and the monitoring of Tau, CK or TBARS may be important for the estimation of oxidative stress during exhaustive exercise. Furthermore, the integrated analyses could be of value in a clinical setting to quantify the extent of oxidative stress risk and reduce the need to perform muscle biopsies as a tool of clinical evaluation.

Effects of Exercise Training on Hippocampus Concentrations of Insulin and IGF-1 in Diabetic Rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Maximal lactate steady state in rats submitted to swimming exercise

The higher concentration during exercise at which lactate entry in blood equals its removal is known as 'maximal lactate steady state' (MLSS) and is considered an important indicator of endurance exercise capacity. The aim of the present study was to determine MLSS in rats during swimming exercise. Adult male Wistar rats, which were adapted to water for 3 weeks, were used. After this, the animals were separated at random into groups and submitted once a week to swimming sessions of 20 min, supporting loads of 5, 6, 7, 8, 9 or 10% of body wt. for 6 consecutive weeks. Blood lactate was determined every 5 min to find the MLSS. Sedentary animals presented MLSS with overloads of 5 and 6% at 5.5 mmol/l blood lactate. There was a significant (P < 0.05) increase in blood lactate with the other loads. In another set of experiments, rats of the same strain, sex and age were submitted daily to 60 min of swimming with an 8% body wt. overload, 5 days/week, for 9 weeks. The rats were then submitted to a swimming session of 20 min with an 8% body wt. overload and blood lactate was determined before the beginning of the session and after 10 and 20 min of exercise. Sedentary rats submitted to the same acute exercise protocol were used as a control. Physical training did not alter the MLSS value (P < 0.05) but shifted it to a higher exercise intensity (8% body wt. overload). Taken together these results indicate that MLSS measured in rats in the conditions of the present study was reproducible and seemed to be independent of the physical condition of the animals. © 2001 Elsevier B.V. All rights reserved.

The monosodium glutamate (MSG) obese rat as a model for the study of exercise in obesity

Obesity is an increasing problem in several countries, leading to health problems. Physical exercise, in turn, can be used effectively by itself or in combination with dietary restriction to trigger weight loss. The present study was designed to evaluate the effects of aerobic exercise training on lipid profile of obese male Wistar rats in order to verify if this model may be of value for the study of exercise in obesity. Obesity was induced by MSG administration (4mg/g, each other day, from birth to 14 days old) After 14 from drug administration, the rats were separated into two groups: MSG-S (sedentary) and MSG-T (exercise trained). Exercise training consisted in 1h/day, 5 days/week, with an overload of 5% bw, for 10 weeks. Rats of the same age and strain, receiving saline at birth, were used as control (C), and subdivided into two groups: C-S and C-T. At the end of the experimental period, MSG-T and C-T rats showed similar blood lactate and muscle glycogen responses to exercise training and acute exercise. MSG-S rats showed significantly higher carcass fat, serum triacylglycerol, serum insulin and liver total fat than C-S rats. On the other hand, MSG-T rats had lower carcass fat, serum triacylglycerol and liver total fat than MSG-S rats. There were no statistical differences in food intake and serum free fatty acids among the groups studied. These data indicate that this model may be of value for the study of exercise effects on tissue and circulating lipid profile in obesity.

Exercise at anaerobic threshold intensity and insulin secretion by isolated pancreatic islets of rats

To evaluate the effect of acute exercise and exercise training at the anaerobic threshold (AT) intensity on aerobic conditioning and insulin secretion by pancreatic islets, adult male Wistar rats were submitted to the lactate minimum test (LMT) for AT determination. Half of the animals were submitted to swimming exercise training (trained), 1 h/day, 5 days/week during 8 weeks, with an overload equivalent to the AT. The other half was kept sedentary. At the end of the experimental period, the rats were submitted to an oral glucose tolerance test and to another LMT. Then, the animals were sacrificed at rest or immediately after 20 minutes of swimming exercise at the AT intensity for pancreatic islets isolation. At the end of the experiment mean workload (% bw) at AT was higher and blood lactate concentration (mmol/L) was lower in the trained than in the control group. Rats trained at the AT intensity showed no alteration in the areas under blood glucose and insulin during OGTT test. Islet insulin content of trained rats was higher than in the sedentary rats while islet glucose uptake did not differ among the groups. The static insulin secretion in response to the high glucose concentration (16.7 mM) of the sedentary group at rest was lower than the sedentary group submitted to the acute exercise and the inverse was observed in relation to the trained groups. physical training at the AT intensity improved the aerobic condition and altered insulin secretory pattern by pancreatic islets. © 2010 Landes Bioscience.

Glucose homeostasis in type 1 diabetic rats after acute physical activity

Physical activity is considered an extremely effective therapy in cases of type 1 diabetes (DM-1), as it promotes glucose uptake independent of insulin action. However, there are few studies on the effect of a single session of exercise on glucose uptake in DM-1 (i.e., in the absence of insulin). Therefore, the purpose of this study was to assess the effect of a single exercise session on glucose homeostasis in DM-1 rats. For this purpose, 30 male rats were divided into three groups: sedentary control (SC), sedentary diabetic (SD), and exercise diabetic (ED). DM was induced by administration of alloxan and identified by the value of fasting glucose. The physical activity consisted of a single swimming session at the anaerobic threshold intensity for diabetic rats (3.5% body weight overload) for 30 min. The oral glucose tolerance test (OGTT) was performed immediately after the physical activity. The animals were sacrificed 48 hr after the OGTT, and samples were taken from the blood, liver, gastrocnemius, and mesenteric and subcutaneous adipose tissue. We observed that DM caused significant reduction in body weight. A single session of physical activity did not modify the response to the OGTT or glucose. However, it resulted in increased HDL cholesterol and hepatic glycogen content. These results suggest that, despite not having an effect on glucose homeostasis, acute physical activity performed at anaerobic threshold intensity leads to beneficial changes in the context of type 1 diabetes.

Physical exercise modulates peripheral levels of brain-derived neurotrophic factor (BDNF): A systematic review of experimental studies in the elderly

The objective of this study was to conduct a systematic review of studies that analyzed the effect of physical exercise on the peripheral levels of BDNF in elderly individuals. Method: We conducted a search in PsycINFO, Biological Abstracts, Pubmed, Web of Science, and Science Direct from 1990 to 2011, using the following keywords: physical exercise , physical activity , physical therapy , training , BDNF , neuroplasticity , neurotrophins , neuroplasticity proteins , aged , older , elderly The articles were considered for inclusion in the review if they were studies with elderly, assessed peripheral (serum and/or plasma) BDNF and evaluated an acute exercise or chronic exercise (training). Results: Five randomized controlled trial and one randomized non-controlled trial studies were analyzed. Five out of six studies reported a significantly higher BDNF response to aerobic acute exercise and to aerobic or strength training program in healthy elderly and elderly with different pathologies. Conclusion: It was not possible to establish a recommendation protocol for the type and intensity of physical exercise required to produce an increase in levels BDNF. However, physical exercise, particularly, moderate-intensity exercises seem to be more effective to promote increase the peripheral levels of BDNF in the elderly. © 2012 Elsevier B.V.

Influence of chronic exercise on serum cortisol levels in older adults

The circulating level of cortisol is regulated by the hypothalamic-pituitary-adrenal axis through a neuroendocrine feedback circuit. This circuit can be activated by physiological stimuli such as stress, diseases, and exercise. High levels of serum cortisol hormone normally occur as a byproduct of aging, and can cause several types of damage to the organism and exacerbate immunosenescence. There is a great deal of variability in the cortisol response with regard to type, intensity, volume, and frequency of exercise. However, these relationships have been extensively studied with respect to the acute effects of exercise. Despite the well-known effects of acute exercise on cortisol response, it is unclear how it is affected by chronic exercise and the aging process. Therefore, the aim of this study was to conduct a review of studies that attempt to analyze the influence of chronic exercise on serum cortisol hormone in older people. In order to accomplish this goal, a review from 1970 to June 2012 period was performed using the following databases: Biological Abstracts, PsycINFO, PubMed/Medline, and the Web of Science. Eight articles met the criteria used in this study. Based on the included articles, chronic exercise may influence the serum levels of cortisol levels in older people. Despite this evidence, these results may not be generalized to the entire population of older people, given the few number of studies and especially because the studies showed diversity in variables and methodologies. © 2013 European Group for Research into Elderly and Physical Activity (EGREPA).