Effects of dynamic ischaemic training on human skeletal muscle dimensions

The effect of training under conditions of local leg ischaemia on muscle area and fibre dimensions was studied in nine males. Leg ischaemia was induced by enclosing the legs in a pressure chamber and sealing the opening with a rubber membrane at the level of the crotch. Air pressure over the legs was 50 mmHg. The subjects performed 16 sessions (45 min) of one-legged supine strenuous ischaemic training during 4 weeks. Exercise intensity was maintained as high as possible during the whole session. The contralateral leg served as a control leg and remained passive during exercise. Before and after the training period, muscle fibre dimensions were determined from biopsy samples taken from the m. vastus lateralis, and leg muscle dimensions were assessed by magnetic resonance imaging (MRI). In the trained leg, mean fibre area increased by 12% (P < 0.05). The MRI-assessed cross-sectional area of the vastus group increased by 4% (P=0.01). In the control leg, mean fibre area and the cross-sectional area of the vastus group were unchanged, while those of the adductor muscle group decreased by 4% (P < 0.05). It is concluded that a short period of strenuous ischaemic endurance training increases the cross-sectional area of the ischaemically trained muscle group, as measured both by MRI and from muscle biopsy samples. In contrast, the adductor muscles in the contralateral thigh showed a decreased cross-sectional area (as assessed by MRI), possibly due to the effects of the strenuous contralateral training, by mechanisms that have yet to be identified. Accepted: 26 December 1999     

Effect of hyperthyroidism on fibre-type composition, fibre area, glycogen content and enzyme activity in human skeletal muscle

Seven hyperthyroid patients were studied by repeated muscle biopsies (vastus lateralis) before and after a period of medical treatment which averaged 10 months. The biopsies were analysed with regard to fibre-type composition, fibre area, capillary density, glycogen content and enzyme activities representing the glycolytic capacity (hexokinase, 6-phosphofructokinase), oxidative capacity (oxoglutarate dehydrogenase, citrate synthase) and Ca2+- and Mg2+-stimulated ATPase in muscle. In the pretreatment biopsy (hyperthyroid state), there was a significantly lower proportion of type I fibres (30% vs. 41%), a higher capillary density (23%), lower glycogen content (33%), and higher hexokinase activity (32%) compared with the post-treatment biopsy. No significant changes in the activity of the remaining enzymes were observed. The present study indicates that hyperthyroidism induces a transformation from type I to type II fibres in human skeletal muscle. The increase in hexokinase activity probably reflects a higher glucose utilization by skeletal muscle in order to compensate partially for the reduced glycogen content.     

Effects of amrinone on shortening velocity and force development in skinned skeletal muscle fibres

Summary The effects of amrinone were studied on single skinned fibres isolated from rat hindlimb muscles. In each fibre a force-velocity relation was determined during maximal calcium activation (pCa=4.45) in control conditions and in the presence of amrinone. The MgATP concentration was 3.93 mm, close to the physiological value. After the experiment the fibre was classified as fast or slow on the basis of its reactivity with anti-myosin monoclonal antibodies. In fast fibres amrinone (3 mm) potentiated isometric tension (P ₀) by 13.8±2.9% (n=13), reduced maximum shortening velocity (V _max ) by 32.6±3.2% and the curvature of the force-velocity relation (a/P ₀) was increased by 98.9±46.0%. All these effects were less pronounced in slow fibres, where V _max was reduced only by 11.4±3.6 (n=16). The effects of amrinone (0.3–6 mm) on the ATPase activity of myofibrils and myosin prepared from fast (tibialis anterior) and slow (soleus) rat skeletal muscles were studied. Amrinone was found to depress Ca–Mg dependent ATPase activity of myofibrillar preparations of the tibialis anterior (up to 16.6±2%) and, to a lesser extent, of the soleus (up to 7.2±1.2%). On the contrary, Ca-stimulated myosin ATPase activity was significantly increased by amrinone in myosin preparations from the tibialis anterior. Experiments were carried out to test whether amrinone (3 mm) might affect the sensitivity of the contractile system to MgATP concentration ([MgATP]). The results obtained showed that (1) the [MgATP] value at which isometric tension reached its maximum was shifted by amrinone from 0.1 mm to 0.3 mm, (2) the slope of the negative relation between [MgATP] and a/P ₀ was made more steep by amrinone, and (3) the Km of the hyperbolic relation between [MgATP] and V _max was increased from 0.39 to 1.71 mm by amrinone, thus indicating a reduced affinity of myosin for MgATP. These results are in accordance with the hypothesis that amrinone exerts a direct effect on the contractile mechanism.     

Changes in skeletal muscle size, fibre-type composition and capillary supply after chronic venous occlusion in rats

Aim: We have previously shown that surgical occlusion of some veins from skeletal muscle results in muscle hypertrophy without mechanical overloading in the rat. The present study investigated the changes in muscle‐fibre composition and capillary supply in hypertrophied muscles after venous occlusion in the rat hindlimb. Methods: Sixteen male Wistar rats were randomly assigned into two groups: (i) sham operated (sham‐operated group; n = 7); (ii) venous occluded for 2 weeks (2‐week‐occluded group; n = 9). At the end of the experimental period, specimens of the plantaris muscle were dissected from the hindlimbs and subjected to biochemical and histochemical analyses. Results: Two weeks after the occlusion, both the wet weight of plantaris muscle relative to body weight and absolute muscle weight showed significant increases in the 2‐week‐occluded group (～15%) when compared with those in the sham‐operated group. The concentrations of muscle glycogen and lactate were higher in the 2‐week‐occluded group, whereas staining intensity of muscle lipid droplets was lower in the 2‐week‐occluded group than those in the sham‐operated group. The percentage of type I muscle fibre decreased, whereas that of type IIb fibre increased in the 2‐week‐occluded group when compared with the sham‐operated group. Although the expression of vascular endothelial growth factor‐188 mRNA increased, the number of capillaries around the muscle fibres tended to decrease (P = 0.07). Conclusion: Chronic venous occlusion causes skeletal muscle hypertrophy with fibre‐type transition towards faster types and changes in contents of muscle metabolites.     

Potassium-transporting proteins in skeletal muscle: cellular location and fibre-type differences

Potassium (K⁺) displacement in skeletal muscle may be an important factor in the development of muscle fatigue during intense exercise. It has been shown in vitro that an increase in the extracellular K⁺ concentration ([K⁺]_e) to values higher than approx. 10 mm significantly reduce force development in unfatigued skeletal muscle. Several in vivo studies have shown that [K⁺]_e increases progressively with increasing work intensity, reaching values higher than 10 mm . This increase in [K⁺]_e is expected to be even higher in the transverse (T)-tubules than the concentration reached in the interstitium. Besides the voltage-sensitive K⁺ (K_v) channels that generate the action potential (AP) it is suggested that the big-conductance Ca²⁺-dependent K⁺ (K_Ca1.1) channel contributes significantly to the K⁺ release into the T-tubules. Also the ATP-dependent K⁺ (K_ATP) channel participates, but is suggested primarily to participate in K⁺ release to the interstitium. Because there is restricted diffusion of K⁺ to the interstitium, K⁺ released to the T-tubules during AP propagation will be removed primarily by reuptake mediated by transport proteins located in the T-tubule membrane. The most important protein that mediates K⁺ reuptake in the T-tubules is the Na⁺,K⁺-ATPase α₂ dimers, but a significant contribution of the strong inward rectifier K⁺ (Kir2.1) channel is also suggested. The Na⁺, K⁺, 2Cl⁻ 1 (NKCC1) cotransporter also participates in K⁺ reuptake but probably mainly from the interstitium. The relative content of the different K⁺-transporting proteins differs in oxidative and glycolytic muscles, and might explain the different [K⁺]_e tolerance observed.     

Effects of ischaemic training on local aerobic muscle performance in man

The aim of the study was to compare the effects of ischaemic and non-ischaemic training on aerobic performance. In 10 subjects, peak oxygen uptake (peak V_{O 2}) and time to fatigue (TTF) for one-legged exercise were measured before and after 4 weeks (4 times week-1) of one-legged training. Each training session started with one leg training for 45 min with 20% blood-flow reduction induced by local application of a supra-atmospheric external pressure of 50 mmHg (ischaemic leg; I-leg). We have previously shown that this decreases leg blood flow by about 20%. The contralateral leg (nonrestricted-flow leg; N-leg), serving as a control, then trained with an identical power-output profile for 45 min but without flow restriction. In the I-leg the average training-induced increments in TTF and peak V_{O 2} were 27 and 24%, respectively. In the N-trained leg TTF and peak-V_{O 2} increased 10 and 14%, respectively. Both increments were significantly greater (P < 0.05) in the I-trained leg. Moreover, the performance increase in the I-trained leg was exaggerated (P < 0.05) in the ischaemic test condition, i.e. there was a specificity in the training response. In conclusion, ischaemia acts as an additive stimulus to training leading to an exaggerated increase in endurance and peak-V_O2 compared to identical training without blood-flow restriction. The main explanation is probably an enhanced local adaptation in the I-trained leg.     

Effects of age and life-time physical training on fibre composition of slow and fast skeletal muscle in rats

The effects of age and endurance training on muscle fibre characteristics were studied in a slow (m. soleus, MS) and in a fast (m. rectus femoris, MRF) skeletal muscle. Wistar rats at ages of 1, 2, 4, 10, and 24 months were used as experimental animals. The trained rats were put to run on a motor-driven treadmill 5 d/wk beginning from the age of 1 month. The body weights of the animals increased continuously throughout their lives. The muscle weights increased up to the age of 10 months, after which they tended to decrease. The trained adult rats had lower body weights as well as lower muscle weights than the untrained adult rats. The amount of the intramuscular lipid decreased with age, especially during the first months of life. The activity of isocitrate dehydrogenase (ICDH) decreased during the growth period in both muscles and remained more or less constant thereafter, whereas the activity of phosphofructokinase decreased with age only in MS. In MS, the trained animals tended to have higher ICDH activities than the untrained animals. The cross-sectional area of the different fibre types in both muscles increased up to the age of 10 months. The major fibre types, type I in MS and type IIB in MRF, were smaller for trained than untrained rats. The percentage number of the slower fibre types of both muscles — type I in MS and types I and IIA in MRF —increased with advancing age. The muscles of the trained animals contained higher percentages of the slower fibre types than those of the untrained rats. The present paper indirectly supports the opinion that not only the metabolic but also the contractile properties of different muscle cell types can be changed by age and long-term endurance-type physical training.     

Effect of satellite cell ablation on low-frequency-stimulated fast-to-slow fibre-type transitions in rat skeletal muscle

The purpose of this study was to determine whether satellite cell ablation within rat fast-twitch muscles exposed to chronic low-frequency stimulation (CLFS) would limit fast-to-slow fibre-type transitions. Twenty-nine male Wistar rats were randomly assigned to one of three groups. Satellite cells of the left tibialis anterior were ablated by weekly exposure to a 25 Gy dose of γ-irradiation during 21 days of CLFS (IRR-Stim), whilst a second group received only 21 days of CLFS (Stim). A third group received weekly doses of γ-irradiation (IRR). Non-irradiated right legs served as internal controls. Continuous infusion of 5-bromo-2'-deoxyuridine (BrdU) revealed that CLFS induced an 8.0-fold increase in satellite cell proliferation over control (mean ± s.e.m. : 23.9 ± 1.7 versus 3.0 ± 0.5 mm⁻², P < 0.0001) that was abolished by γ-irradiation. M-cadherin and myogenin staining were also elevated 7.7- and 3.8-fold ( P < 0.0001), respectively, in Stim compared with control, indicating increases in quiescent and terminally differentiating satellite cells; these increases were abolished by γ-irradiation. Myonuclear content was elevated 3.3-fold ( P < 0.0001) in Stim, but remained unchanged in IRR-Stim. Immunohistochemical analyses revealed attenuation of fast-to-slow fibre-type transitions in IRR-Stim compared with Stim. Comparable changes were observed at the protein level by SDS-PAGE. It is concluded that although considerable adaptive potential exists within myonuclei, satellite cells play a role in facilitating fast-to-slow fibre-type transitions.     

Exercise and training effects on ceramide metabolism in human skeletal muscle

In rat skeletal muscle prolonged exercise affects the content and composition of ceramides, but in human skeletal muscle no data are available on this compound. Our aim was to examine the content of ceramide- and sphingomyelin fatty acids and neutral, Mg(2+)-dependent sphingomyelinase activity in skeletal muscle in untrained and trained subjects before and after prolonged exercise. Healthy male subjects were recruited into an untrained (n = 8, VO2,max 3.8 +/- 0.2 1 min1) and a trained (n = 8, Vo2,max 5.1 +/- 0.1 1 min2) group. Before and after a 3-h exercise bout (58 +/- 1% VO2,max) a muscle biopsy was excised from the vastus lateralis. Ceramide and sphingomyelin were isolated using thin-layer chromatography. The content of individual ceramide fatty acids and sphingomyelin fatty acids was measured by means of gas-liquid chromatography. The activity of neutral, Mg(2+)-dependent sphingomyelinase was measured using N-[14CH3]-sphingomyelin as a substrate. Prior to exercise, the muscle total ceramide fatty acid content in both groups was similar (201 +/- 18 and 197 +/- 9 nmol g(-1) in the untrained and trained group, respectively) and after exercise a 25% increase in the content was observed in each group. At rest, the muscle total sphingomyelin fatty acid content was higher in untrained than in trained subjects (456 +/- 10, 407 +/- 7 nmol g(-1), respectively; P < 0.05). After exercise a 20% increase (P < 0.05) in total sphingomyelin was observed only in the trained subjects. The muscle neutral, Mg(2+)-dependent sphingomyelinase activity was similar in the two groups at rest and a similar reduction was observed after exercise in both groups (untrained from 2.19 +/- 0.08 to 1.78 +/- 0.08 and trained from 2.31 +/- 0.12 to 1.80 +/- 0.09 nmol (mg protein) (-1) h(-1); P < 0.05 in each case). In conclusion, we have reported, for the first time, the values for ceramide fatty acid content and neutral, Mg2(+)-dependent sphingomyelinase activity in human skeletal muscle. The results indicate that acute prolonged exercise affects ceramide metabolism in human skeletal muscle both in untrained and in trained subjects and this may influence muscle cell adaptation and metabolism.     

Effects of hypobaric hypoxia on histochemical fibre-type composition and myosin heavy chain isoform component in the rat soleus muscle

Histochemical fibre-type composition and myosin heavy chain isoform component in the soleus muscle were studied in normoxic rats at postnatal ages of 5, 10, 15, and 20 weeks and in rats exposed to hypobaric hypoxia (460 torr) for 5 weeks from postnatal ages of 5, 10, and 15 weeks. The increase in the percentage of type I fibres and the concomitant decrease in that of type IIa fibres in the soleus muscle of normoxic rats were observed until 15 weeks of age. On the other hand, no change in the fibre-type composition of the muscle during postnatal development was observed in hypoxic rats, irrespective of the age at which they were exposed to hypoxia. The changes in the myosin heavy chain isoform component (MHC I and MHC IIa) of the muscle during postnatal development and by hypoxia corresponded well with those in the muscle fibre-type composition. It is concluded that hypobaric hypoxia inhibits the growth-related shift of muscle fibre-types from type IIa to type I and of myosin heavy chain isoforms from MHC IIa to MHC I in the rat soleus muscle, and that there are no changes in the muscle fibre-type composition or the myosin heavy chain isoform component caused by hypoxia after the shifts in these parameters which occur during postnatal development are completed.     

Acyl-coenzyme A binding protein expression is fibre-type specific in rat skeletal muscle but not affected by moderate endurance training

The metabolic active form of free fatty acids, long-chain acyl-coenzyme A (lc-acyl-CoA), binds to its 10-kDa binding protein with high affinity. In the present study, we investigated the content of lc-acyl-CoA binding protein (ACBP) in different skeletal muscle fibre types. Soleus had the highest expression of ACBP (0.33+/-0.02 microg mg protein(-1)) and the content was as high as in heart muscle. The content in mixed gastrocnemius (0.27+/-0.02 microg mg protein(-1)), extensor digitorum longus (0.21+/-0.01 microg mg protein(-1)) and white gastrocnemius (0.16+/-0.01 microg mg protein(-1)) were lower than in soleus and differed from each other ( P<0.05). The ACBP content correlated positively with the fraction of myosin heavy chain I in the skeletal muscles (Spearman rank correlation rho=0.90; P<0.0001), and negatively with the myosin heavy chain IIB fraction ( rho=-0.92; P<0.0001). The content of ACBP also correlated with 3-hydroxy-acyl-CoA dehydrogenase (HAD) and citrate synthase (CS). Five weeks of endurance training increased HAD and CS activities in soleus and mixed gastrocnemius but did not affect the ACBP content. These findings demonstrate that ACBP expression is fibre-type specific in skeletal muscles and correlates with beta-oxidative potential. Training-induced increase in oxidative capacity was not paralleled by an increase in ACBP content.     

Fatiguability and fibre composition of human skeletal muscle.

The fatiguability of the quadriceps muscle was investigated in 10 male subjects (25-40 yrs), with inter-individual differences in fibre composition of their vastus lateralis muscles (range 25-65% fast twitch, FT, muscle fibres). Fatiguability was assessed as the decline in maximal force (in % of initial values) with 50 repeated isokinetic knee-extensions at fast angular velocity (3.14 rad/s). Each contraction lasted 0.5 s and the rest periods were about 0.7 s. Every subject was tested on two occasions and the standard deviation for a single determination of fatiguability was 1.4%. The decline in force after 50 contractions was on the average about 45%. The individual values varied, however, and a linear correlation was present between fatiguability and % FT fibres (r = 0.86, p less than 0.01). It was concluded that development of fatigue in human skeletal muscle performing repeated fast dynamic contractions with maximal effort was most marked in muscles with a high proportion FT fibres. This finding was in conformity with earlier results from animal skeletal muscle preparations.     

Effects of cooling on human skin and skeletal muscle

To investigate the effects of cooling on local temperature and circulation in the skin and skeletal muscle at different cooling temperatures. Ten male subjects (mean age 24.9 years) participated in this study. Intramuscular temperatures were measured by inserting two 22-gauge temperature probes (needle length; 8 and 18 mm) into the ankle dorsiflexors, while skin temperature was measured using a thermocouple attached to the leg skin anteriorly. Near-infrared spectroscopy was also used to evaluate the concentration changes in oxygenated, deoxygenated, and total hemoglobin/myoglobin in local skin and skeletal muscle. These measurements were simultaneously performed during the 10-min noncooling, 30-min cooling (cooling pad temperature; 0, 10, or 20°C), and 60-min recovery periods. Under all cooling conditions, skin and intramuscular temperatures decreased during cooling (P < 0.01) and began to increase after the cooling pad was removed. However, these values did not return to baseline values during the recovery period (P < 0.01). Moreover, tissue temperatures tended to show lower values during cooling at lower cooling temperatures. All hemoglobin/myoglobin concentrations also showed a concomitant significant decrease during cooling under three cooling conditions (P < 0.01); the oxygenated and total hemoglobin/myoglobin concentrations did not return to the exact values before cooling during the recovery period. This study suggested that the rate of decrease in tissue temperature depends on the cooling temperature and the effects of cooling on tissue temperatures and circulation tend to be maintained during 60 min post-cooling period despite the cooling temperature.     

Effects of two high-intensity intermittent training programs interspaced by detraining on human skeletal muscle and performance

Summary The purpose of this study was to investigate the effects of repeated high-intensity intermittent training programs interspaced by detraining on human skeletal muscle and performances. First, nineteen subjects were submitted to a 15-week cycle ergometer training program which involved both continuous and high-intensity interval work patterns. Among these 19 subjects, six participated in a second 15-week training program after 7 weeks of detraining. Subjects were tested before and after each training program for maximal aerobic power and maximal short-term ergocycle performances of 10 and 90 s. Muscle biopsy from the vastus lateralis before and after both training programs served for the determination of creatine kinase (CK), hexokinase, phosphofructokinase (PFK), lactate dehydrogenase (LDH), malate dehydrogenase, 3-hydroxyacyl-CoA dehydrogenase (HADH) and oxoglutarate dehydrogenase (OGDH) activities. The first training program induced significant increases in all performances and enzyme activities but not in CK. Seven weeks of detraining provoked significant decreases in maximal aerobic power and maximal 90 s ergocycle performance. While the interruption of training had no effect on glycolytic enzyme markers (PFK and LDH), oxidative enzyme activities (HADH and OGDH) declined. These results suggest that a fairly long interruption in training has negligeable effects on glycolytic enzymes while a persistent training stimulus is required to maintain high oxidative enzyme levels in human skeletal muscle. The degree of adaptation observed after the second training program confirms that the magnitude of the adaptive response to exercise-training is limited.     

Blockades of mitogen-activated protein kinase and calcineurin both change fibre-type markers in skeletal muscle culture

Activation of either the calcineurin or the extracellular signal-regulated kinase (ERK1/2) pathway increases the percentage of slow fibres in vivo suggesting that both pathways can regulate fibre phenotypes in skeletal muscle. We investigated the effect of calcineurin blockade with cyclosporin A and mitogen-activated protein kinase kinase (MEK1/2) blockade with U0126 upon myosin heavy chain (MHC) isoform mRNA levels and activities of metabolic enzymes after 1 day, 3 days and 7 days of treatment in primary cultures of spontaneously twitching rat skeletal muscle. U0126 treatment significantly decreased MHC Ibeta mRNA levels and significantly increased MHC IIX, MHC IIB, embryonal MHC and perinatal MHC mRNA levels when compared to control. In addition, U0126 treatment significantly increased lactate dehydrogenase, creatine kinase, hexokinase, malate dehydrogenase and beta-hydroxyacyl-CoA dehydrogenase activities above control values while a significant reduction in the percentage of pyruvate dehydrogenase in the active form was also observed. Calcineurin blockade significantly decreased both MHC Ibeta and embryonal mRNA levels below control and significantly increased MHC IIX mRNA levels. Significant increases in the activities of both lactate dehydrogenase and creatine kinase above control values were also seen following cyclosporin A treatment. In conclusion, the results suggest that calcineurin upregulates slow-fibre genes and suppresses fast-fibre genes. Similarly, the ERK1/2 pathway upregulates slow-fibre MHC and suppresses fast-fibre MHC isoforms. However, the effect on enzyme activities is not fibre-type specific. The effect of U0126 on the percentage of pyruvate dehydrogenase in the active form suggests that the ERK1/2 pathway may also be involved in regulation of the phosphorylation state of this enzyme.