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00005768-200901000-0001500005768_2009_41_129_gjovaag_mechanical_1miscellaneous-article< 108_0_25_5 >Medicine & Science in Sports & Exercise© 2009 American College of Sports MedicineVolume 41(1)January 2009pp 129-136Effect of Training with Different Mechanical Loadings on MyHC and GLUT4 Changes[BASIC SCIENCES]GJØVAAG, TERJE F.1,2,3; DAHL, HANS A.2,31Oslo University College, Oslo, NORWAY; 2Norwegian School of Sport Sciences, Oslo, NORWAY; and 3Department of Anatomy, Institute of Basic Medical Sciences, University of Oslo, Oslo, NORWAYAddress for correspondence: Terje F. Gjøvaag, Ph.D., Oslo University College, P.o.b. 4 St. Olavs Plass, N-0130 Oslo, Norway; E-mail: .Submitted for publication November 2007.Accepted for publication June 2008.ABSTRACTPurpose: There is an inverse relationship between insulin sensitivity and percentage of myosin heavy chain IIx (MyHC IIx) isoform in sedentary, obese, and type 2 diabetic humans. How different exercise conditions may reduce the proportion of MyHC IIx and in parallel elevate glucose transporter 4 (GLUT4) content is interesting in a therapeutic setting. This study investigates the nature of exercise signals regulating MyHC gene switching and whether it is accompanied by GLUT4 changes.Methods: Thirty-two subjects performed high loading (60% of 1 repetition maximum [RM]) or low loading (30% of 1 RM) elbow extensions in a training apparatus and exercised three times per week for either 5 wk (low volume) or 8 wk (high volume). MyHC and GLUT4 contents in the musculus triceps brachii were measured by Western blotting pre- and posttraining and after 8 wk of detraining.Results: All training regimes resulted in MyHC IIx changes of similar magnitude, and differences in training volume had no effect on the outcome. The reduction in MyHC IIx content after high loading, high volume was similar to low loading, matching volume of training. Thus, there was no effect of training load on MyHC changes. GLUT4 increased more after high than low loading (P < 0.0.1). In addition, the larger increases in the GLUT4 were associated with the larger reductions in MyHC IIx content (r = −0.56, P < 0.01). Detraining returned GLUT4 to baseline, but MyHC IIx content was still higher than baseline (P < 0.01).Conclusion: Magnitude of loading is not important for suppression of MyHC IIx but for increases in GLUT4 content. The GLUT4 content responded, however, more rapidly to detraining than the MyHC IIx and IIa isoforms.There is an ongoing discussion concerning the nature of exercise signals that regulate myosin heavy chain (MyHC) gene switching during long-term training in humans. It has been suggested (1) that training volume is the sole determining factor for myosin heavy chain IIx (MyHC IIx) suppression during repetitive, low force activity, but that the magnitude of mechanical loading is a governing factor for the suppression of MyHC IIx during training with higher loads (1). In contrast, results from other studies suggest that the magnitude of loading is not important for suppression of MyHC IIx in exercising muscle (6). Hence, there is no consensus regarding the magnitude of mechanical loading, whether it is important for fast MyHC isoform shifts (1) or not (6). If the training volume is important for fast MyHC shifts during repetitive training with low mechanical loading (e.g., continuous endurance training), one would expect that a large total volume of this type of training would suppress the MyHC IIx content more than a small total volume of training. In contrast, if the magnitude of mechanical loading is important, one might anticipate that training with a high mechanical load would suppress the MyHC IIx content more than training with less heavy loads despite a matching total volume of training. If loading is of little importance, the implications are that training with low or higher mechanical loading will suppress the MyHC IIx content to similar levels. The response of the MyHC isoforms to different doses of exercise is, however, little investigated in human skeletal muscle, and we will analyze this relationship by comparing the effects of training with high or low relative loading combined with matching high (HV) or low volumes (LV) on MyHC isoform expression in the musculus triceps brachii.Training studies show that long-term endurance training increases insulin sensitivity and glucose transporter 4 (GLUT4) content in humans (15), and it is suggested that the total amount of the GLUT4 protein available may be an important factor for governing insulin sensitivity (14). It is also reported that insulin sensitivity is inversely related to the percentage of MyHC IIx muscle fibers in sedentary (12), obese (5,21), and type 2 diabetic humans (5). Consequently, a reduction in proportion of MyHC IIx fibers and an increase in the protein content of GLUT4 would be associated with increased insulin sensitivity and responsiveness in type 2 diabetic individuals (for a review, see (17). Decreased insulin-stimulated glucose transport is one key aspect of insulin resistance, but glucose uptake into skeletal muscle cells by GLUT4 is also stimulated by contractile activity, independent of insulin (for a review, see (18). Chronic training can elevate the GLUT4 content in human skeletal muscle (18) as well as reduce the percentage of MyHC IIx fibers (e.g., 6), and it is suggested that transformation of muscle fiber types from IIx to IIa should be considered when attempting to explain elevations in the GLUT4 content with training (14). In contrast, others have argued that the GLUT4 content may be more related to the activity level of the muscle fiber than to its expression of contractile proteins (8,9). Consequently, there are conflicting views on the association between GLUT4 and MyHC and on their regulation in response to training (e.g., 8,14,23,24).Based on this, we will investigate how different combinations of exercise may change the proportion of MyHC in a favorable direction (MyHC IIx à MyHC IIa) and elevate the GLUT4 content. Thus, in a therapeutic setting, the present study may provide valuable information regarding tailoring of physical activity programs as part of a lifestyle intervention for type 2 diabetic persons. Specifically, we have tested the following hypotheses: 1) Will a large total volume of repetitive training with low mechanical loading (continuous training) suppress the MyHC IIx content more than a small total volume of this type of training? 2) Will a high mechanical load (intermittent training) suppress the MyHC IIx content more than training with less heavy loads but with a matching total volume of training? 3) What type of training is the better to reduce the proportion of MyHC IIx and in parallel increase the GLUT4 content in the exercising muscles?MATERIALS AND METHODSSubjectsIn the present study, we chose to include young, healthy, and lean subjects because the subjects had to tolerate repeated biopsies and intensive training three times per week for maybe eight or more weeks. Before inclusion in the study, 77 subjects completed two questionnaires, one about self-reported health and another about habitual physical activity. One the basis of the results from a 1-RM test of their elbow extensors and the questionnaires, we excluded 37 subjects with a history of regular exercise of the elbow extensors and those with the highest and the lowest 1 RM.The remaining subjects (n = 40), who were untrained in their musculus triceps brachii, were included in the study and were assigned to one of four different training regimes by a random number generator. This randomization resulted in three groups with 9 subjects each (group HI/HV, HI/LV, and LI/LV) and one group with 13 subjects (group LI/HV). The subjects were healthy and were not using any medication for chronic diseases. They had not been engaged in any type of regular exercise of the elbow extensors for at least 6 months before the start of this study. During the study, the subjects were instructed not to engage in physical activities that exercised their triceps muscle other than their prescribed training program. The subjects' adherence to this regimen was controlled by their self-reported training diary. There was a total of eight dropouts from the study (three men and five women), resulting in the following group sizes: HL/HV, n = 6 (1 male and 5 females); HL/LV, n = 5 (2 males and 3 females); LL/HV, n = 13 (6 males and 7 females); and LL/LV, n = 8 (3 males and 5 females). The mean (SE) age, height, bodyweight, and body mass index of the 32 persons who completed this study was 22.0 (0.6) yr, 172.8 (1.3) cm, 64.6 (1.2) kg, and 21.6 (0.3), respectively. Written informed consent was obtained from all subjects, and the study was approved by the Regional Committees for Medical Research Ethics (REK) in Norway.Study designAll subjects performed elbow extensions three times per week on alternating days with both arms simultaneously in a commercial training apparatus (Triceps Extension Machine; Cybex International Inc., Medway, MA). The training groups differed in terms of the type of training, the relative load applied, the total number of repetitions per session, and the total number of weeks in training (Fig. 1).FIGURE 1. The study design. The training mode (elbow extensions), velocity of movement (paced by a metronome), range of motion (defined by the training apparatus), frequency of training, and number of recovery days between training sessions were similar for all subjects. HL indicates high loading; LL, low loading; HV, high volume; LV, low volume.Training loadThe loading during training was calculated as a percentage of the subjects' 1 RM of the elbow extensors. Relative to each other, the subjects performed either high loading (HL) or low loading (LL) training. One repetition maximum was tested once every week, and the training load was adjusted progressively to keep the loading at the designated percentage of 1 RM.HL intermittent training (60% of 1 RM) was 20 repetitions during 30 s (paced by a metronome) followed by 30-s recovery. A total of 100 repetitions completed one set, and the subjects performed four sets per session with 5-min recovery between sets. The subjects performed a total of 400 repetitions per session.LL continuous training (30% of 1 RM) was 40 repetitions per minute (i.e., the same pace as intermittent training) for a total of 800 repetitions per session. There were no recovery periods during the training session.Training volume and durationThe total volume of training is calculated as the total weight lifted per session times the total number of sessions. The total volume was matched between the two LV groups and the two HV groups. The LV subjects trained for a mean (SE) of 5 (1) weeks, whereas the HV subjects trained for a mean of 8 (0.8) wk.DetrainingAfter completion of the training period, the subjects went through a period of detraining lasting 8 wk. During this period, the subjects were instructed to refrain from regular training of the elbow extensors.Biopsy protocolThe subjects reported to the laboratory after an overnight fast. Needle biopsies were taken from the lateral head of musculus triceps brachii of the nondominant arm before training (Pre-T), after termination of the training period (Post-T), and 8 wk after the Post-T biopsy (De-T). Post-T biopsies were taken 5 d after the last exercise session.Analysis of GLUT4Muscle membrane preparations (red blood cells, contractile proteins, and insoluble materials removed) were prepared as described previously (10). Total protein concentration of the samples was determined using the bicinchoninic acid (Micro-BCA; Pierce, Rockford, IL) protein assay. The membrane preparations were diluted with sample buffer to give equal amounts of protein. The samples were then heated for 4 min at 100°C in a water bath. Proteins were separated using a 4% stacking and 12% separating gel (Mini-Protean 3; BioRad Laboratories, Hercules, CA). Electrophoresis was run at 150 V (V) for ∼60 min. Separated proteins were transferred from gels to 0.2 μm PVDF membranes (BioRad Laboratories). Protein transfer was done at 100 V for 70 to 80 min (Mini Trans Blot system, BioRad Laboratories). The PVDF membranes were subsequently blocked with skimmed milk powder in 0.1% TBS-Tween (TBS-T) on a shaker over night at 4°C. After blocking, PVDF membranes were incubated at room temperature with a GLUT4 antibody (1:3000, # 4670-1704; Biogenesis Ltd., Poole, UK). After incubation with primary antibody, membranes were washed in TBS-T and incubated with an HRP-conjugated secondary antibody (NA 934; Amersham, Piscataway, NJ) for 60 min. After incubation with the secondary antibody, membranes were repeatedly washed with TBS-T before detection of protein bands. Membranes were subsequently incubated with an ECL + reagent (RPN 2132; Amersham), and proteins bands were visualized on x-ray films.Separation of MyHC isoformsHomogenization and electrophoretic separation of myosin heavy chains (MyHC) were performed on biopsy samples (4). Total protein concentration of the samples was determined using the bicinchoninic acid (Micro-BCA; Pierce) protein assay. After electrophoretic separation of MyHC proteins, protein bands were visualized by Coomassie staining. Identification of MyHC isoforms was confirmed by Western blotting (3). Antibodies used were anti-MyHC IIa + IIx or anti-MyHC I + IIa (SC-71 and BF-35, respectively, kindly provided by Prof. S. Schiaffino, University of Padua, Italy) and anti-MyHC I antibody (NCL-MyHC; Novacastra Laboratories, Newcastle upon Tyne, UK). A typical response of GLUT4 and MyHC responses after training and subsequent detraining is shown in Figure 2.FIGURE 2. Electrophoretic separation of MyHC isoforms and immunoblot of the GLUT4 expression in the musculus triceps brachii. Typical response of myosin heavy chain (MyHC) isoforms and glucose transporter 4 (GLUT4) after HL training and subsequent detraining in one subject.QuantificationDigitized scans of Coomassie-stained SDS gels (MyHC) and x-ray films (GLUT4) were quantified by image analysis software (TotalLab; Nonlinear Dynamics, Newcastle upon Tyne, UK). All samples were run in duplicate and averaged. GLUT4 values are reported as mean integrated optical density (IOD) units. IOD is calculated as the sum of pixel gray level values within a defined area divided by the number of pixels within the same area. We did not use a control sample to correct for differences between gels, but before quantification of IOD levels, each digitized scan was calibrated by adjusting the OD levels of the scanned image to the expected values of a an optical density control scale (Kodak control scale T-14; Kodak, Rochester, NY). MyHC isoforms were identified as MyHC I, IIa, or IIx, and each isoform was expressed as a percentage of the total MyHC content detected in the samples.Statistical analysesThe variables were tested by two-way repeated-measures ANOVA with time as within-subjects variables, load and volume as between-subjects factors, and gender as a covariate. Significant main effects were subsequently analyzed by univariate analysis with an LSD post hoc test to examine differences between the training groups. Linear regression was used to investigate the relationships between the variables. All calculations were done in SPSS statistical program (SPSS Inc., Chicago, IL). Statistical significance was set at P < 0.05.RESULTSMain resultsLong-term training increased the GLUT4 content by 64% (Pre-T vs Post-T; P < 0.001), whereas the reduced percentage of the MyHC IIx isoform was accompanied by a reciprocal increase of the MyHC IIa isoform (Post-T vs Pre-T; P < 0.001 for both; Table 1). After detraining, the MyHC IIx and IIa isoform contents were reversed (Post-T vs De-T; P < 0.01 for both), but the De-T contents of both isoforms were still different compared with their respective Pre-T values (both P < 0.01). In contrast, GLUT4 levels had returned to Pre-T levels.TABLE 1. The content of MyHC isoforms and GLUT4 in the musculus triceps brachii after training and detraining.Group comparisonsThe changes in GLUT4 content were clearly dependent on the magnitude of loading during training. In this respect, the GLUT4 content increased more after training with high loading (groups HL/ HV and HL/LV) than after training with low loading (groups LL/HV and LL/LV) (128% vs 13%, respectively; P < 0.01).In contrast to the GLUT4 changes, the type of training did not influence the outcome of MyHC IIx and IIa isoform changes. In this respect, no significant differences between the training groups were observed for training-induced changes (Post-T values − Pre-T values) in the MyHC IIa or IIx contents (Figs. 3A and B). Thus, all training regimens regardless of mechanical loading and total training volume resulted in changes of similar magnitudes with a general reduction of the MyHC IIx content and a reciprocal increase in the MyHC IIa content.FIGURE 3. Changes in MyHC IIa (A) and MyHC IIx (B) content after long-term training and detraining in relation to different training groups. (Filled bars) Training changes (Post-T − Pre-T values). (Open bars) Detraining changes (De-T − Post-T values). Values are means (SE). aP < 0.05; LL/HV versus LL/LV.CorrelationsThere were no significant correlations between MyHC isoforms and GLUT4 contents Pre-T, Post-T, or De-T. The changes in MyHC IIa and IIx content after detraining were inversely related to the magnitude of their changes after training (r = −0.65 and −0.56, respectively, P < 0.001, for both). Thus, the larger changes in MyHC IIa and IIx after detraining were observed in the subjects with the larger changes after training.The protein content of GLUT4 increased with an increasing Post-T percentage of MyHC IIa in the triceps muscle (Fig. 4) but only for the subjects that performed intermittent training with HL (r = 0.73, P < 0.01). On the other hand, the GLUT4 protein content after continuous LL training was similar over the whole range of Post-T MyHC IIa content. There was an inverse relationship between the training-induced reductions (Post-T − Pre-T values) in MyHC IIx content and the increases (Post-T − Pre-T values) in GLUT4 content after training (r = −0.56, P < 0.01), meaning that the larger increases in GLUT4 content were observed in the subjects with the larger reductions in MyHC IIx content.FIGURE 4. Changes in GLUT4 content in relation to Post-T MyHC IIa content. (Closed circles) HL; (open circles) LL. IOD indicates integrated optical density (the integral of the mean optical density over the area of each band). Correlation is for HL only.Total training volumeTotal mean (SE) training volume for the LV groups (HL/LV and LL/LV) was 107 (2.9) tons, whereas the HV groups (HL/HV and LL/HV) lifted an average of 179 (1.9) tons (HV vs LV; P < 0.001).DISCUSSIONEffects of trainingIn the present study, both high loading (HL) and low loading (LL) training-induced transformation of muscle fiber types from MyHC IIx to MyHC IIa, but only HL training increased the GLUT4 expression in the musculus triceps brachii. The relationship between GLUT4 and MyHC isoform expression is, however, not completely resolved, and it has been suggested that transformation of muscle fiber types from MyHC IIx to MyHC IIa should be considered when attempting to explain elevations in the GLUT4 content with training (14). Interestingly, we observed that the larger increases in the GLUT4 content occurred in the individuals with the larger reductions in the MyHC IIx content. It is, however, unlikely that the MyHC IIx directly determines the GLUT4 expression, and others have argued that the GLUT4 content may be more related to the activity level of the muscle fiber than to its expression of contractile proteins (8,9).Exercise-induced adaptation of contractile and metabolic properties in muscle may be regulated by different signaling pathways (e.g., 20). Previous research on signaling pathways have, however, suggested that metabolic properties and MyHC isoforms are independently regulated and that metabolic changes can occur without transformation of fast MyHC isoforms (23). In contrast, Russel et al. (24) argues that increased expression of the transcriptional coactivator PGC-1 (peroxisome proliferator-activated receptor-γ coactivator-1) might be linked to MyHC transformations and increases in GLUT4 content after endurance training. Thus, there are seemingly conflicting views on the association between GLUT4 and MyHC gene expression in muscle. If, however, multiple signaling pathways are activated during training, this may possibly integrate the metabolic changes with the MyHC transformations that are observed after training (29). This would ensure a timely match of contractile and metabolic properties to meet the altered workload imposed by chronic training (29). However, the exact signaling mechanisms linking the different types of neuromuscular activity to altered gene expression remain to be elucidated.The fact that most subjects in the present study experienced a reduction in the percentage of MyHC IIx fibers and a reciprocal increase in the percentage of MyHC IIa fibers and GLUT4 content (Table 1) is interesting also of therapeutic reasons. Although we trained young, healthy, and lean subjects, our findings may have implications for a diseased population. In this regard, it is reported that type 2 diabetes and insulin resistance in humans are associated with a greater percentage of MyHC IIx muscle fibers, and it is suggested that a reduction in proportion of MyHC IIx fibers and an increase in the proportion of MyHC IIa fibers would increase GLUT4 levels and increase muscle insulin sensitivity and responsiveness in type 2 diabetic individuals (17). Thus, to look at a possible association between the transformation of the MyHC IIx isoform to the MyHC IIa isoform and GLUT4 expression, we plotted the training-induced changes in GLUT4 levels in relation to the Post-T content of MyHC IIa (Fig. 4). Interestingly, there were only minor and quite similar increases in individual GLUT4 protein content after training with low mechanical loading, regardless of the individual Post-T MyHC IIa content (Fig 4; open circles). Evidently, this type of training is unable to stimulate to increases in the GLUT4 protein content, and given this scenario, there seems to be a relatively uniform expression of GLUT4 regardless of the percentage of muscle fibers expressing MyHC IIa. Previous investigations have argued that continuous training provides the highest stimulus for inducing the GLUT4 expression in skeletal muscle (26), but the present study show much larger increases in the individual GLUT4 content after intermittent training. In the present study, a high mechanical loading is necessary to stimulate to increases in the GLUT4 protein content; thus, the activity level of the muscle fibers is clearly important (8). During HL training, however, there seems to be an increase in GLUT protein content that is related to an increasing percentage of MyHC IIa muscle fibers. Thus, there are larger increases of GLUT4 in the individuals with a high Post-T percentage of MyHC IIa (Fig. 4; closed circles). It is argued that it is difficult for many patients with type 2 diabetes to engage in endurance-type training programs (13), and studies on a disease population using a resistance type of exercise are few. Holten et al. (13), however, found that resistance training for 30 min, three times per week, increased insulin action and increased the protein contents of GLUT4 in patients with type 2 diabetes. Because they did not determine the percentage of MyHC IIa and IIx in their biopsies, it is not possible to tell whether these changes were accompanied by transformation of fast MyHC isoforms. In our study, both training regimes reduced the MyHC IIx content in exercising muscles to the same extent (Fig. 3B), but HL training increased the GLUT4 content more than LL training. Thus, the present results support the view that a resistance type of training with HL may represent an attractive alternative for patients with type 2 diabetes (13). Sedentary and obese persons may, however, have less tolerance to high training loads; thus, the work/rest protocol during exercise needs to be carefully designed.Given that the proportion of MyHC IIx muscle fibers in many muscles may be small, it is relevant to discuss if a reduced proportion of MyHC IIx have a physiological role for individuals with insulin resistance and impaired glucose metabolism. In this regard, Bandyopadhyay et al. (5) showed that the percent composition of MyHC IIx fibers in insulin-resistant subjects (∼16% MyHC IIx) was higher than in lean, healthy subjects (∼5% MyHC IIx). Furthermore, Venojärvi et al. (27) investigated the role of muscle fiber type composition in regulation of the glucose metabolism in subjects with impaired glucose tolerance (IGT). The subjects were divided in two groups with either a high proportion of MyHC I (IGT_slow) or a high proportion of MyHC II isoforms (IGT_fast). The actual MyHC IIx content in the musculus vastus lateralis was ∼17% (IGT_slow) and ∼23% (IGT_fast), respectively, that is, a difference of 6% in the MyHC IIx content. Despite this small difference, Venojärvi et al. (27) argued that the difference between the proportions of MHC I and MHC II isoforms could partially explain why the IGT_fast subjects were more insulin-resistant than the IGT_slow subjects. Correspondingly, Hedman et al. (11) observed that muscle morphology explained 16% of the variation in insulin sensitivity in older men. Interestingly, half of this variation was attributed exclusively to the proportion of MyHC IIx fibers in the muscle. In summary, these data underscore the importance of further investigations on the association between GLUT4 expression and the content of fast MyHC isoforms.Given that a transformation of MyHC IIx to MyHC IIa fibers may be interesting also in a therapeutic setting, it is interesting to investigate the nature of exercise signals regulating MyHC gene switching. There is, however, limited information on MyHC changes after different training loadings and concerning a putative role of the training volume per se on MyHC adaptations. In the present study, we find that training with a high mechanical load does not suppress the MyHC IIx expression more than training with less heavy loads but with a matching total volume of training (Fig. 3B), and we believe our findings support the view that the mechanical load is of little importance in this matter (6). Admittedly, our subjects have not lifted as heavy loads as the subjects in other resistance training studies (e.g., 1,19,25), and consequently one may ask if those results are comparable to ours. In a resistance training study with three loading regimes (3-5 RM, 9-11 RM, and 20-28 RM) and similar volumes of training, the magnitude of reductions in MyHC IIx content was the same for all loading conditions (6). In the present study, the two high loading (HL) groups performed repeated series of 20 repetitions, and most of the subjects were close to fatigue at the end of each of the series. Thus, the HL type of training seems comparable to the 20- to 28-RM loading regime previously mentioned (6). In addition, the two low loading (LL) groups in the present study performed elbow extensions at 30% of the subjects' 1 RM, but the type of changes in MyHC content after LL training was similar to the type of changes observed during training with much higher mechanical loadings (e.g., [1]). Consequently, the combined evidence from the present and other studies (6) does not support the view (1) that the magnitude of mechanical loading governs the suppression of the MyHC IIx content in exercising human muscles. Interestingly, we also find that that during repetitive, low force training (LL groups), the magnitude of reductions in MyHC IIx content was similar after either a high volume (HV) or a low volume (LV) of training (Fig. 3B). The total volume during HV training was on average ∼70% higher than the volume during LV training; hence, we find no support for the view of that the training volume is the sole determining factor for MyHC IIx suppression during repetitive, low force activity (1). Because the magnitude of reductions in MyHC IIx content after low force, low volume (LL/LV) training was similar to the changes after a much larger total volume (LL/HV), our results may indicate that the downregulation of MyHC IIx is not related to the total volume of training but perhaps to whether MyHC IIx fibers are recruited or not during training.Effects of detrainingIn the present study, detraining for 8 wk only partly reversed the training-induced changes in MyHC IIx and IIa content in the musculus triceps brachii. The De-T MyHC IIx and IIa contents were not fully restored to their respective Pre-T levels; hence, there was no indication of a MyHC IIx overshoot as reported in previous studies (1,2).Because we detrained our subjects for a shorter period than the previously mentioned studies (8 vs 12 wk, respectively), we may have missed an overshoot because it may need more than 8 wk to develop. In this respect, serial biopsies of well-trained endurance athletes have been performed 1, 2, 3, 8, and 12 wk after cessation of an endurance training program (7). Endurance training reduced the proportion of MyHC IIx, whereas detraining progressively increased the MyHC IIx content. Twelve weeks of detraining did, however, not elevate the IIx content any further than at 8 wk. In addition, a MyHC IIx overshoot has not been confirmed by other resistance studies (16,25) exercising the musculus vastus lateralis and with similar or longer training and detraining periods as the previously mentioned studies (1,2). Consequently, a possible induction of an overshoot in MyHC IIx expression after shorter or longer periods (8-32 wk) of detraining is still an open question. The GLUT4 content responded, however, more rapidly to detraining than MyHC IIx and IIa isoforms, and after 8 wk of detraining, GLUT4 levels had returned to baseline (Table 1). Other studies of endurance-trained athletes confirm our findings and show a significant reduction in the GLUT4 content after 6 (28) or 10 d (22) of detraining. Thus, muscle GLUT4 content starts to decline rapidly after cessation of contractile activity and has returned to baseline after several weeks of detraining. Consequently, to maintain a favorable adaptation in type 2 diabetic persons, a break from training should not be too long.To summarize, within the range of mechanical loading used in this study, the magnitude of mechanical loading is not important for suppression of MyHC IIx synthesis, and a large training volume do not suppress the MyHC IIx content more than a lesser volume of training. An HL is, however, more effective than a lower loading in increasing the GLUT4 content. In addition, GLUT4 increases correlates with the MyHC IIa content in the trained triceps brachii; thus, the suppression of MyHC IIx and the concomitant increase in MyHC IIa and GLUT4 after intermittent training may represent a favorable adaptation in a therapeutic setting. Finally, 8 wk of detraining is sufficient to return training-induced changes in GLUT4 content to baseline, whereas the changes in MyHC IIx and IIa contents are not fully reversed.We thank all our subjects for their time, effort, and patience during this study and Dr. Helge Dyre Meen for help in obtaining the biopsies. We also thank Prof. Schiaffino for kindly supplying us with MyHC antibodies (SC-71 and BF-35). This work was supported by grants from the Oslo University College, the Norwegian School of Sport Sciences, and The Research Council of Norway. 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