Muscle enzyme and fiber type-specific sarcomere protein increases in serum after inertial concentric-eccentric exercise.

Muscle damage induced by inertial exercise performed on a flywheel device was assessed through the serum evolution of muscle enzymes, interleukin 6, and fiber type-specific sarcomere proteins such as fast myosin (FM) and slow myosin (SM). We hypothesized that a model of muscle damage could be constructed by measuring the evolution of serum concentration of muscle proteins following inertial exercise, according to their molecular weight and the fiber compartment in which they are located. Moreover, by measuring FM and SM, the type of fibers that are affected could be assessed. Serum profiles were registered before and 24, 48, and 144 h after exercise in 10 healthy and recreationally active young men. Creatine kinase (CK) and CK-myocardial band isoenzyme increased in serum early (24 h) and returned to baseline values after 48 h. FM increased in serum late (48 h) and remained elevated 144 h post-exercise. The increase in serum muscle enzymes suggests increased membrane permeability of both fast and slow fibers, and the increase in FM reveals sarcomere disruption as well as increased membrane permeability of fast fibers. Consequently, FM could be adopted as a fiber type-specific biomarker of muscle damage.

Fast and slow myosins as markers of muscle injury.

OBJECTIVE: The diagnosis of muscular lesions suffered by athletes is usually made by clinical criteria combined with imaging of the lesion (ultrasonography and/or magnetic resonance) and blood tests to detect the presence of non-specific muscle markers. This study was undertaken to evaluate injury to fast and slow-twitch fibres using specific muscle markers for these fibres. METHODS: Blood samples were obtained from 51 non-sports people and 38 sportsmen with skeletal muscle injury. Western blood analysis was performed to determine fast and slow myosin and creatine kinase (CK) levels. Skeletal muscle damage was diagnosed by physical examination, ultrasonography and magnetic resonance and biochemical markers. RESULTS: The imaging tests were found to be excellent for detecting and confirming grade II and III lesions. However, grade I lesions were often unconfirmed by these techniques. Grade I lesions have higher levels of fast myosin than slow myosin with a very small increase in CK levels. Grade II and III lesions have high values of both fast and slow myosin. CONCLUSIONS: The evaluation of fast and slow myosin in the blood 48 h after the lesion occurs is a useful aid for the detection of type I lesions in particular, since fast myosin is an exclusive skeletal muscle marker. The correct diagnosis of grade I lesions can prevent progression of the injury in athletes undergoing continual training sessions and competitions, thus aiding sports physicians in their decision making.

Disturbances of the sarcoplasmic reticulum and transverse tubular system in 24-h electrostimulated fast-twitch skeletal muscle.

Chronic low-frequency stimulation of rabbit tibialis anterior muscle over a 24-h period induces a conspicuous loss of isometric tension that is unrelated to muscle energy metabolism (J.A. Cadefau, J. Parra, R. Cusso, G. Heine, D. Pette, Responses of fatigable and fatigue-resistant fibres of rabbit muscle to low-frequency stimulation, Pflugers Arch. 424 (1993) 529-537). To assess the involvement of sarcoplasmic reticulum and transverse tubular system in this force impairment, we isolated microsomal fractions from stimulated and control (contralateral, unstimulated) muscles on discontinuous sucrose gradients (27-32-34-38-45%, wt/wt). All the fractions were characterized in terms of calcium content, Ca2+/Mg2+-ATPase activity, and radioligand binding of [3H]-PN 200-110 and [3H]ryanodine, specific to dihydropyridine-sensitive calcium channels and ryanodine receptors, respectively. Gradient fractions of muscles stimulated for 24 h underwent acute changes in the pattern of protein bands. First, light fractions from longitudinal sarcoplasmic reticulum, enriched in Ca2+-ATPase activity, R1 and R2, were greatly reduced (67% and 51%, respectively); this reduction was reflected in protein yield of crude microsomal fractions prior to gradient loading (25%). Second, heavy fractions from the sarcoplasmic reticulum were modified, and part (52%) of the R3 fraction was shifted to the R4 fraction, which appeared as a thick, clotted band. Quantification of [3H]-PN 200-110 and [3H]-ryanodine binding revealed co-migration of terminal cisternae and t-tubules from R3 to R4, indicating the presence of triads. This density change may be associated with calcium overload of the sarcoplasmic reticulum, since total calcium rose three- to fourfold in stimulated muscle homogenates. These changes correlate well with ultrastructural damage to longitudinal sarcoplasmic reticulum and swelling of t-tubules revealed by electron microscopy. The ultrastructural changes observed here reflect exercise-induced damage of membrane systems that might severely compromise muscle function. Since this process is reversible, we suggest that it may be part of a physiological response to fatigue.

Glycogen depletion and resynthesis during 14 days of chronic low-frequency stimulation of rabbit muscle.

Electro-stimulation alters muscle metabolism and the extent of this change depends on application intensity and duration. The effect of 14 days of chronic electro-stimulation on glycogen turnover and on the regulation of glycogen synthase in fast-twitch muscle was studied. The results showed that macro- and proglycogen degrade simultaneously during the first hour of stimulation. After 3 h, the muscle showed net synthesis, with an increase in the proglycogen fraction. The glycogen content peaked after 4 days of stimulation, macroglycogen being the predominant fraction at that time. Glycogen synthase was determined during electro-stimulation. The activity of this enzyme was measured at low UDPG concentration with either high or low Glu-6-P content. Western blots were performed against glycogen synthase over a range of stimulation periods. Activation of this enzyme was maximum before the net synthesis of glycogen, partial during net synthesis, and low during late synthesis. These observations suggest that the more active, dephosphorylated and very low phosphorylated forms of glycogen synthase may participate in the first steps of glycogen resynthesis before net synthesis is observed, while partially phosphorylated forms are most active during glycogen elongation.

A short training programme for the rapid improvement of both aerobic and anaerobic metabolism.

The aim of this study was to evaluate the changes in aerobic and anaerobic metabolism produced by a newly devised short training programme. Five young male volunteers trained daily for 2 weeks on a cycle ergometer. Sessions consisted of 15-s all-out repetitions with 45-s rest periods, plus 30-s all-out repetitions with 12-min rest periods. The number of repetitions was gradually increased up to a maximum of seven. Biopsy samples of the vastus lateralis muscle were taken before and after training. Performance changes were evaluated by two tests, a 30-s all-out test and a maximal progressive test. Significant increases in phosphocreatine (31%) and glycogen (32%) were found at the end of training. In addition, a significant increase was observed in the muscle activity of creatine kinase (44%), phosphofructokinase (106%), lactate dehydrogenase (45%), 3-hydroxy-acyl-CoA dehydrogenase (60%) and citrate synthase (38%). After training, performance of the 30-s all-out test did not increase significantly, while in the maximal progressive test, the maximum oxygen consumption increased from mean (SD) 57.3 (2.6) ml x min(-1) x kg(-1) to 63.8 (3.0) ml min(-1) x kg(-1), and the maximum load from 300 (11) W to 330 (21) W; all changes were significant. In conclusion, this new protocol, which utilises short durations, high loads and long recovery periods, seems to be an effective programme for improving the enzymatic activities of the energetic pathways in a short period of time.

The distribution of rest periods affects performance and adaptations of energy metabolism induced by high-intensity training in human muscle.

The effect of the distribution of rest periods on the efficacy of interval sprint training is analysed. Ten male subjects, divided at random into two groups, performed distinct incremental sprint training protocols, in which the muscle load was the same (14 sessions), but the distribution of rest periods was varied. The 'short programme' group (SP) trained every day for 2 weeks, while the 'long programme' group (LP) trained over a 6-week period with a 2-day rest period following each training session. The volunteers performed a 30-s supramaximal cycling test on a cycle ergometer before and after training. Muscle biopsies were obtained from the vastus lateralis before and after each test to examine metabolites and enzyme activities. Both training programmes led to a marked increase (all significant, P < 0.05) in enzymatic activities related to glycolysis (phosphofructokinase - SP 107%, LP 68% and aldolase - SP 46%, LP 28%) and aerobic metabolism (citrate synthase - SP 38%, LP 28.4% and 3-hydroxyacyl-CoA dehydrogenase - SP 60%, LP 38.7%). However, the activity of creatine kinase (44%), pyruvate kinase (35%) and lactate dehydrogenase (45%) rose significantly (P < 0.05) only in SP. At the end of the training programme, SP had suffered a significant decrease in anaerobic ATP consumption per gram muscle (P < 0.05) and glycogen degradation (P < 0.05) during the post-training test, and failed to improve performance. In contrast, LP showed a marked improvement in performance (P < 0.05) although without a significant increase in anaerobic ATP consumption, glycolysis or glycogenolysis rate. These results indicate that high-intensity cycling training in 14 sessions improves enzyme activities of anaerobic and aerobic metabolism. These changes are affected by the distribution of rest periods, hence shorter rest periods produce larger increase in pyruvate kinase, creatine kinase and lactate dehydrogenase. However, performance did not improve in a short training programme that did not include days for recovery, which suggests that muscle fibres suffer fatigue or injury.

Contractile activity modifies Fru-2,6-P(2) metabolism in rabbit fast twitch skeletal muscle.

Modification of muscular contractile patterns by denervation and chronic low frequency stimulation induces structural, physiological, and biochemical alterations in fast twitch skeletal muscles. Fructose 2,6-bisphosphate is a potent activator of 6-phosphofructo-1-kinase, a key regulatory enzyme of glycolysis in animal tissues. The concentration of Fru-2,6-P(2) depends on the activity of the bifunctional enzyme, 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFK-2/FBPase-2), which catalyzes the synthesis and degradation of this metabolite. This enzyme has several isoforms, the relative abundance of which depends on the tissue metabolic properties. Skeletal muscle expresses two of these isoforms; it mainly contains the muscle isozyme (M-type) and a small amount of the liver isozyme (L-type), whose expression is under hormonal control. Moreover, contractile activity regulates expression of muscular proteins related with glucose metabolism. Fast twitch rabbit skeletal muscle denervation or chronic low frequency stimulation can provide information about the regulation of this enzyme. Our results show an increase in Fru-2,6-P(2) concentration after 2 days of denervation or stimulation. In denervated muscle, this increase is mediated by a rise in liver PFK-2/FBPase-2 isozyme, while in stimulated muscle it is mediated by a rise in muscle PFK-2/FBPase-2 isozyme. In conclusion, our results show that contractile activity could alter the expression of PFK-2/FBPase-2.

Effect of chronic electrostimulation of rabbit skeletal muscle on calmodulin level and protein kinase activity.

(a) Chronic electrostimulation of fast-twitch skeletal muscles makes them resemble slow-twitch muscles. The involvement of second-messenger cascades in this muscle reprogramming is not well understood. The goal of this study was to examine protein kinase activities and calmodulin levels as a function of the duration of electrostimulation. (b) Fast-twitch rabbit muscle was subjected to continuous low-frequency electrostimulation for 2 weeks. The extensor digitorum longus was taken and examined for calmodulin concentration and cAMP-dependent (PKA). Ca(2+)-phospholipid-dependent (PKC) and Ca(2+)-calmodulin-dependent (CaM kinase or PKB) protein kinase activities. (c) Electrostimulation for 14 days led to a significant increase in total calmodulin level and PKB activity, both rising in the cytosolic fraction. Protein kinase C translocated to the membrane fraction, although total activity did not change. (d) These changes could be related with electrostimulation-induced changes in excitation-contraction coupling.

Effect of summer intermission on skeletal muscle of adolescent soccer players.

BACKGROUND: To study the effect of some weeks of rest on three groups of adolescent soccer players, who had undergone systematic training for eleven months. EXPERIMENTAL DESIGN: Retrospective and comparative investigation; duration 4-8 weeks. SETTING: young amateur players from a Spanish football club were examined at the beginning and at the end of the summer rest period. PARTICIPANTS: 37 young soccer players aged 14, 15 and 16 years old. They were members of three football teams. INTERVENTIONS: during the rest period they were free from any training program. MEASURES: biopsies of M. vastus lateralis were taken immediately after training and after the summer holidays. The type, percentage and diameter of the fibers, as well as the enzymes of glycogen metabolism (glycogen synthase and glycogen phosphorylase), glycolysis (phosphofructokinase and lactate dehydrogenase), oxidative metabolism (succinate dehydrogenase and citrate synthase) and creatine kinase and transaminase (aspartate and alanine aminotransferase) were studied. RESULTS: Detraining had an adaptation effect, decreasing the cross-sectional area of type I and type II fibers, and decreasing the activities of creatine kinase, citrate synthase, phosphofructokinase, lactate dehydrogenase and aspartate aminotransferase. CONCLUSIONS: The results can help trainers to plan the length of the rest period between training.

Effect of chronic alcoholism on human muscle glycogen and glucose metabolism.

To determine the effect of alcohol on carbohydrate metabolism, 48 human muscle biopsies from chronic alcoholics were studied. The level of glycogen and the activities of the enzymes catalyzing glycogen and glucose metabolism were analyzed. Chronic alcohol intake produced an increase in glycogen concentration and a decrease in pyruvate kinase activity before the first signs of myopathy appeared. When myopathy was present, glycogen decreased. These changes may contribute to the decline in skeletal muscle performance in these patients.

Responses of fatigable and fatigue-resistant fibres of rabbit muscle to low-frequency stimulation.

This study investigates early adaptive responses of fast-twitch muscle to increased contractile activity by low-frequency stimulation. Changes in metabolite levels and activities of regulatory enzymes of carbohydrate metabolism were investigated in rabbit tibialis anterior muscle after 24 h of stimulation. In addition, changes elicited during a 5-min lasting acute stimulation experiment were compared between 24-h-prestimulated and contralateral control muscles. Stimulation for 5 min reduced energy-rich phosphates and glycogen, and increased lactate in the control muscle. A transient elevation of fructose 2,6-bisphosphate demonstrated that activation of phosphofructokinase 2 was an immediate response to contractile activity. Prestimulated muscles displayed nearly normal values for ATP, phosphocreatine and glycogen, and did not augment lactate. Increased activities of hexokinase and phosphofructokinase 2 and permanently elevated levels of fructose 2,6-bisphosphate pointed to enhanced glycolysis with glucose as the main fuel in the prestimulated muscle. Isometric tension of the control muscle decreased rapidly a few minutes after the onset of stimulation. In the prestimulated muscles, tension was almost stable, but amounted to only 30%-40% of the initial tension of the control muscle. In view of the fibre type distribution of rabbit tibialis anterior, these findings suggested that a large fibre fraction of the prestimulated muscle, possibly the glycolytic type IID fibres, did not contract. Therefore, the possibility must be considered that the metabolite pattern of the 24-h-stimulated muscle primarily reflected metabolic activities of the contracting, less fatigable fibres, most likely type IIA and type I fibres.(ABSTRACT TRUNCATED AT 250 WORDS)

Glucose 1,6-bisphosphate and fructose 2,6-bisphosphate in muscle from healthy humans and chronic alcoholic patients.

Chronic alcohol intake produces an increase in the concentration of glucose 1,6-bisphosphate and fructose 2,6-bisphosphate in human muscle before the first sign of myopathy appears. When myopathy was present both sugars decreased to the levels of healthy humans. These changes could contribute to the decline in skeletal muscle performance.