The structure of the paramyosin core in molluscan thick filaments

Summary The thick filaments of molluscan muscles have been examined by electron microscopy and X-ray diffraction in order to test whether the structure of the paramyosin core is crystalline and not helicoidal. In accurately cut transverse sections of the white adductor muscle of the oyster the thick filaments are uniformly stained but, when the sections are tilted, parallel striations are revealed. Essentially all the filaments can be shown to exhibit striations which can only be explained in terms of a crystalline structure. The crystallinity is, however, not perfect since the striations although often straight are more frequently curved.Two sets of striations of different spacing can be detected in each filament on tilting to either side of the filament axis. Their spacings when compared to the Bear-Selby neta spacing obtained from X-ray diffraction patterns of the embedded muscle indicate that the striations are seen when the filaments are viewed down the and 301 planes of the Bear-Selby net.Striations have been observed in thick filaments from other catch muscles, the white adductor of the clam, the smooth adductor of the scallop and the anterior byssus retractor of the mussel as well as in a non-catch muscle, the red adductor of the clam. In all cases the spacing of the striations was similar indicating that, contrary to previous reports, when the conditions are the same thea spacing of the Bear-Selby net is approximately constant from one muscle to another and hence that the paramyosin structure in all these muscles is similar.     

Ultrastructural changes in myocardial cells of rats fed a low protein diet

 Ultrastructural changes in the ventricular myocardial cells in rats fed a low protein diet were examined by electron microscopy. The most striking changes were observed in the I-band region of the sarcomeres, which occurred very occasionally in myofibrils. In the sarcomere affected the I-band region was often fractured and/or disintegrated on one side, leaving an extended space, while the opposing I-band region disappeared along with dislocation of the intact A-band toward the adjacent Z-line. This dislocation was presumably attributed to the elasticity of titins connecting between the end of thick filaments and the Z-line. Fractured I-band regions were often accompanied by the dilated sarcoplasmic reticulum in the close vicinity of them. In some myofibrils the streaming and/or disruption of the Z-line were occasionally observed where disarrangement of thick and thin myofilaments were usually present. The study suggests that the fracture of the I-band region, consisting of actin and titin filaments, and the streaming of the Z-line of myofibrils are due to a proteolytic action of calpain and/or cathepsin L, which are activated by leaked Ca²⁺ ion and/or by modification of internal circumstances of the cytoplasm induced by a low protein diet, thus resulting in a low cardiac output. Received: 22 April 1998 / Accepted: 7 September 1998     

Calpain 3 participates in sarcomere remodeling by acting upstream of the ubiquitin-proteasome pathway

Mutations in the non-lysosomal cysteine protease calpain 3 cause limb-girdle muscular dystrophy type 2A (LGMD2A). Our previous studies of the calpain 3 knockout mouse (C3KO) suggested a role for calpain 3 in sarcomere formation and remodeling. Calpain 3 may mediate remodeling by cleavage and release of myofibrillar proteins, targeting them for ubiquitination and proteasomal degradation. Loss of proper protein turnover may be the basis for this muscle disease. To test this hypothesis in vivo, we used an experimental model of hindlimb unloading and reloading that has been shown to induce sarcomere remodeling. We showed that the rate of atrophy and especially the rate of growth are decreased in C3KO muscles under conditions promoting sarcomere remodeling. In wild-type mice, an elevated level of ubiquitinated proteins was observed during muscle reloading, which is presumably necessary to remove atrophy-specific and damaged proteins. This increase in ubiquitination correlated with an increase in calpain 3 expression. C3KO muscles did not show any increase in ubiquitination at the reloading stage, suggesting that calpain 3 is necessary for ubiquitination and that it acts upstream of the ubiquitination machinery. We found upregulation of heat shock proteins in C3KO muscles following challenge with a physiological condition that requires highly increased protein degradation. Furthermore, old C3KO mice show evidence of insoluble protein aggregate formation in skeletal muscles. These studies suggest that accumulation of aged and damaged proteins can lead to cellular toxicity and a cell stress response in C3KO muscles, and that these characteristics are pathological features of LGMD2A.     

Properties of soleus muscle Z-lines and induced Z-line analogs revealed by dissection with Ca2+-activated neutral protease

Rat soleus muscle Z-lines and Z-line anomalies induced by neostigmine methyl sulfate (NMS) and cat soleus muscle Z-lines and Z-line anomalies induced by tenotomy were examined by electron microscopy before and after dissection of muscle fibers with Ca2+-activated neutral protease (CAF) to elucidate structural properties of Z-lines and related Z-line-type structures. In both normal and treated muscles, interdigitation of thin (6-7 nm) filaments, which were continuous with I-filaments (actin) from adjacent sarcomeres, was observed at the Z-line in longitudinal section. Both neostigmine methyl sulfate and tenotomy treatments induced muscle atrophy associated with Z-line degradation, streaming, and irregular distribution and accumulation of Z-line material and Z-rod formation. Tenotomized muscle also was characterized by the presence of N-line-like bands and I-Z-I brushes. CAF digestion removed the electron-dense covering material from Z-rods and revealed a backbone of actin filaments. The origin of Z-rods, their structural similarity to Z-lines in longitudinal and cross section, and their susceptibility to CAF indicate that Z-rods are directly related to native Z-lines and are probably lateral polymers of a basic Z-line unit. The regular square net alignment (22 nm) of I-filaments (actin) in cross sections of I-Z-I brushes which contain no N-lines suggests that the I-square net arrangement near the Z-line is determined by Z-filament-actin filament interaction rather than by the N-line or other factors. The results suggest that I-filaments (actin) penetrate the mammalian Z-line and are Z-line constituents and that the width of Z-lines and the length of Z-rods are determined by the amount of overlap of actin filaments. The perpendicular periodicity of Z-rods and the zigzag-oblique arrowheadlike appearance seen in longitudinal sections of Z-lines are attributed to alpha-actinin.     

Ultrastructural Changes and Glutathione Depletion in the Skeletal Muscle Induced by Protein Malnutrition

Ultrastructural changes and glutathione level were investigated in the pectoralis muscle of rats fed a low-protein diet. Electron microscopy demonstrated the ultrastructural changes of occasional myofibrils affected with protein deficiency that were characterized with the streaming and/ or disruption of the Z-line and disintegration of sarcomeric striation. In the affected sarcomeres, sarcomere length was often elongated and fragmented thick filaments were present together with dense amorphous materials flowing from the damaged Z-line. Glutathione level of muscle in the low-protein diet group (5%casein) was reduced to about one-third of that in the control diet group (20% casein). The study suggests that depletion of glutathione by protein malnutrition is responsible for inducing myofibrillar damage through the excess leaking of Ca 2+ into the cytosol.     

Skeletal muscle HSP72 response to mechanical unloading: influence of endurance training

AIMS: It has been shown that increased contractile activity results in heat shock protein 72 (HSP72) accumulation in various skeletal muscles. By contrast, there is no consensus for muscle HSP72 response to muscle disuse for short duration (5-8 days). On the basis of a greater constitutive HSP72 expression in slow-twitch muscles we tested the hypothesis that mechanical unloading for a longer period (2 weeks) would affect this phenotype to a greater extent. Secondly, we evaluated the effects of a physiological muscle heat shock protein (HSP) enhancer (endurance training) on HSP response to unloading and muscle remodelling. METHODS: Adult male Wistar rats were assigned randomly to four groups: (1) sedentary weight-bearing; (2) hindlimb-unloaded (HU) via tail suspension for 2 week; (3) trained on a treadmill (6 week) and (4) trained 6 week and then HU for 2 week. RESULTS: Unloading resulted in a preferential atrophy of slow muscles [soleus (SOL), adductor longus (AL)] and a slow-to-fast fibre transition with no change in HSP72 level. HSP72 levels were significantly lower in fast muscles [extensor digitorum longus (EDL) and plantaris (PLA)], and did not change with mechanical unloading. Endurance training was accompanied by a small (SOL) or a large (EDL, PLA) increase in HSP72 level with no change in AL. Training-induced accumulation of HSP72 disappeared with subsequent unloading in the SOL and PLA whereas HSP72 content remained elevated in EDL. CONCLUSION: The results of this study indicate that (1) after 2 weeks of unloading no change occurred in HSP72 protein levels of slow-twitch muscles despite a slow-to-fast fibre transition; and (2) the training-induced increase of HSP72 content in skeletal muscles did not attenuate fibre transition.     

Ultrastructural Changes and Glutathione Depletion in the Skeletal Muscle Induced by Protein Malnutrition

Ultrastructural changes and glutathione level were investigated in the pectoralis muscle of rats fed a low-protein diet. Electron microscopy demonstrated the ultrastructural changes of occasional myofibrils affected with protein deficiency that were characterized with the streaming and/ or disruption of the Z-line and disintegration of sarcomeric striation. In the affected sarcomeres, sarcomere length was often elongated and fragmented thick filaments were present together with dense amorphous materials flowing from the damaged Z-line. Glutathione level of muscle in the low-protein diet group (5%casein) was reduced to about one-third of that in the control diet group (20% casein). The study suggests that depletion of glutathione by protein malnutrition is responsible for inducing myofibrillar damage through the excess leaking of Ca 2+ into the cytosol.     

The structure of the sarcotubular system in avian muscle

Most avian skeletal muscles have their triads at the A-I junction, and there is a complex sarcoplasmic reticulum in the Z-line region because of the architectural arrangement of two triads in one sarcomere. The sarcotubular system in the pigeon serratus superficialis metapatagialis muscle was examined with the electron microscope. In these muscles the t-tubules open to the extra-cellular space, and there is an indication that mitochondria may connect to the sarcotubular system. The elements of the sarcoplasmic reticulum between two triads of a single sarcomere is similar to that in the frog, and consists of the triads, intermediate cisternae, longitudinal tubules, and fenestrated collar. In the portion of the sarcoplasmic reticulum between triads of adjacent sarcomeres, the triads are coupled. This is brought about by triads extending from one sarcomere to the next and by the connection of the reticulum between triads of adjacent sarcomeres. Some of the tubules making up the Z-line reticulum have a fenestrated appearance, similar to the fenestrated collar.     

The ultrastructure of skeletal and smooth muscle in experimental protein malnutrition in rats fed a low protein diet

Light microscopy of the pectoralis muscle of rats on a low protein diet did not show such morphological alterations as atrophy, degeneration, or sarcoplasmic edema, but electron microscopy occasionally demonstrated ultrastructural changes only in the sarcomeres of myofibrils. In the affected sarcomeres, the Z-line was disrupted and often showed a jagged structure. The Z-substance with electron opacity was frequently present flowing along the long axis of myofibrils, here referred to as the streaming of Z-lines. In addition, regular striations formed by the reciprocal arrangement of thick and thin filaments disappeared from the affected sarcomeres, though these filaments were still discernible. Two or more consecutive sarcomeres in a single myofibril were occasionally involved in these changes. A further two or more neighboring sarcomeres at the same level of myofibrils were affected transversely by these structural alterations. On the other hand, the ultrastructure of the intestinal smooth muscle was not affected by protein deficiency. The study suggests that the ultrastructural damage induced by a low protein diet is attributed to the activation of endogenous protease by the excess leaking of Ca2+ into the cytosol as a result of lipid peroxidation of cell membrane by raised free radicals, owing to the depletion of glutathione production by protein deficiency. It also suggests that the smooth muscle cells differ in their susceptibility to protein deficiency from the skeletal muscle cells.     

Injury and repair of the soleus muscle after electrical stimulation of the sciatic nerve in the rat.

To study injury and subsequent changes in skeletal muscles, the rat sciatic nerve was electrically stimulated at 50 Hz and muscle contraction was induced for 30 min. Muscle damage was classified into five types (hypercontraction, hyperstretching, Z band disorders, misalignment of myofilament and regions of scarce myofilaments) by electron microscopy and quantified by ultrastructural assessment. After electrical nerve stimulation, the percentages of the injured areas of the soleus muscle were 18.8 +/- 15.8% (mean +/- SD) at 0 h, 9.7 +/- 1.0% at 6 h, 22.0 +/- 23.6% at 12 h, 13.1 +/- 3.2% at 24 h, 4.9 +/- 6.0% at 3 days and 0.5 +/- 0.4% at 7 days. At 0 h, the vast majority of ultrastructural alterations were sarcomere hypercontraction. At 6 h, hypercontraction was not recognizable and sarcomere hyperstretching and Z band disarrangement constituted the major findings. At 12 h, when the injury reached its maximum, myofilament disorganization and hyperstretching were predominant. At 24 h or afterwards, the injury began to decrease and recovered to almost normal conditions by 7 days. There were very few necrotic muscle fibers in all specimens. It is considered that the muscle lesions in the present study were reversible, and recovered through changes in various types of sarcomere alterations. Z band streaming and free ribosomes were frequently found at 12 and 24 h, which may indicate repair processes rather than newly formed lesions.     

Muscle-specific effects of hindlimb suspension and clenbuterol in mature male rats

Anabolic agents are useful tools for probing the mechanisms by which muscle fibers perceive and respond to disuse. beta(2)-Adrenergic agonists exert protective, and/or reparative, effects on atrophying muscle tissue. The effects of one such agent, clenbuterol (Cb), were examined on muscle mass, total protein content, and myofibrillar protein content in selected hindlimb muscles [adductor longus (ADL), extensor digitorum longus (EDL), plantaris (PLAN), soleus (SOL)] of mature male rats, under different loading conditions. Pair-fed rats were divided into four experimental groups: vehicle- and Cb-treated nonsuspended, vehicle- and Cb-treated hindlimb suspended (HLS). Experiments lasted 14 days, during which the rats received subcutaneous injections of 1 mg/kg Cb or 1 ml/kg vehicle. HLS induced significant atrophy in all muscles, except the EDL, in a generally fiber type-related pattern. However, myofibrillar protein content was affected in a more regional pattern. Cb treatment of nonsuspended rats induced hypertrophy in all muscles, in a generally uniform pattern. However, myofibrillar protein content was affected in a more fiber type-related pattern. Cb treatment of HLS rats reduced or eliminated HLS-induced atrophy in all muscles, in a muscle-specific pattern. Overall, the ADL and SOL were most susceptible to HLS-induced atrophy. The PLAN had the greatest magnitude of Cb-induced sparing of atrophy. The results show that, in mature male rats, Cb exerts anabolic effects that are load-dependent and muscle-specific. Responses to this drug cannot be reliably predicted by fiber-type composition alone.     

Effect of thyroid hormones on acetylcholinesterase mRNA levels in the slow soleus and fast extensor digitorum longus muscles of the rat

In the rat, the level of acetylcholinesterase messenger RNA in the typical slow soleus muscles is only about 20-30% of that in the fast extensor digitorum longus muscles. The expression of contractile proteins in muscles is influenced by thyroid hormones and hyperthyroidism makes the slow soleus muscle faster. The influence of thyroid hormones on the levels of acetylcholinesterase messenger RNA level in the slow soleus and fast extensor digitorum longus muscle of the rat was studied in order to examine the effect of thyroid hormones on muscle acetylcholinesterase expression. Hyperthyroidism was induced in rats by daily thyroid hormone injection or thyroid hormone releasing tablet implantation. Hind-limb suspension was applied to produce muscle unloading. Muscle denervation or reinnervation was achieved by sciatic nerve transection or crush. Acetylcholinesterase messenger RNA levels were analyzed by Northern blots and evaluated densitometrically. Hyperthyroidism increased the levels of acetylcholinesterase messenger RNA in the slow soleus muscles close to the levels in the fast extensor digitorum longus. The effect was the same in the unloaded soleus muscles. Acetylcholinesterase expression increased also in the absence of innervation (denervation), in the presence of changed nerve activation pattern (reinnervation), and under enhanced tonic neural activation of the soleus muscle (electrical stimulation). However, the changes were substantially smaller than those observed in the control soleus muscles. Enhancement of acetylcholinesterase expression in the soleus muscles by the thyroid hormones is, therefore, at last in part due to hormonal effect on the muscle itself. On the contrary, increased level of the thyroid hormones had no influence on acetylcholinesterase expression in the normal fast extensor digitorum longus muscles. However, some enhancing influence was apparent whenever the total number of nerve-induced muscle activations per day in the extensor digitorum longus muscle was increased. Thyroid hormones seem to be an independent extrinsic factor of acetylcholinesterase regulation in the slow soleus muscle.     

Influences of sarcomere length and selective elimination of myosin filaments on the localization and orientation of triads in rat muscle fibres

Summary Ultrastructural features of internal membrane systems directly concerned with the excitation-contraction coupling were observed in chemically skinned muscle bundles prepared from Wistar rat extensor digitorum longus muscle to clarify two questions: (1) whether triads localization and orientation are influenced by the sarcomere length and (2) whether triads localization and orientation are influenced by the selective elimination of myosin filaments. The distance between triads and Z-lines depends on the sarcomere length: it increase with sarcomere length. There is a highly significant (p<0.01) positive correlation between sarcomere length and the distance between triads and Z-line. The distance between Z-line and triads is dependent on sarcomere length, but the width of junctional gap remains constant when the sarcomere length was changed. Incubation in a concentration of KCl, which dissolves the myosin filaments. The localization and orientation of triads was not altered by the elimination of myosin filaments, however, the distance between the Z-line and triads becomes shorter when the myosin filaments was completely eliminated. There were significant differences (p<0.01) between control and myosin filament eliminated fibres in the distances between Z-lines and triads (over 2 m). These results indicate that the distance between triads and Z-lines depend on the sarcomere length and that there may be some connection(s) between triads and the myofibrils. There is that the elastic component responsible for tethering the triads in their normal position is interrupted either because it is normally attached to the myosin filaments, or because it is extracted by the conditions that dissociate myosin filaments.     

Morphofunctional responses to anaemia in rat skeletal muscle

Adult male Sprague-Dawley rats were randomly assigned to two groups: control and anaemic. Anaemia was induced by periodical blood withdrawal. Extensor digitorum longus and soleus muscles were excised under pentobarbital sodium total anaesthesia and processed for transmission electron microscopy, histochemical and biochemical analyses. Mitochondrial volume was determined by transmission electron microscopy in three different regions of each muscle fibre: pericapillary, sarcolemmal and sarcoplasmatic. Muscle samples sections were also stained with histochemical methods (SDH and m-ATPase) to reveal the oxidative capacity and shortening velocity of each muscle fibre. Determinations of fibre and capillary densities and fibre type composition were made from micrographs of different fixed fields selected in the equatorial region of each rat muscle. Determination of metabolites (ATP, inorganic phosphate, creatine, creatine phosphate and lactate) was done using established enzymatic methods and spectrophotometric detection. Significant differences in mitochondrial volumes were found between pericapillary, sarcolemmal and sarcoplasmic regions when data from animal groups were tested independently. Moreover, it was verified that anaemic rats had significantly lower values than control animals in all the sampled regions of both muscles. These changes were associated with a significantly higher proportion of fast fibres in anaemic rat soleus muscles (slow oxidative group = 63.8%; fast glycolytic group = 8.2%; fast oxidative glycolytic group = 27.4%) than in the controls (slow oxidative group = 79.0%; fast glycolytic group = 3.9%; fast oxidative glycolytic group = 17.1%). No significant changes were detected in the extensor digitorum longus muscle. A significant increase was found in metabolite concentration in both the extensor digitorum longus and soleus muscles of the anaemic animals as compared to the control group. In conclusion, hypoxaemic hypoxia causes a reduction in mitochondrial volumes of pericapillary, sarcolemmal, and sarcoplasmic regions. However, a common proportional pattern of the zonal distribution of mitochondria was maintained within the fibres. A significant increment was found in the concentration of some metabolites and in the proportion of fast fibres in the more oxidative soleus muscle in contrast to the predominantly anaerobic extensor digitorum longus.     

Macrophages promote muscle membrane repair and muscle fibre growth and regeneration during modified muscle loading in mice in vivo

Muscle injury or modified muscle use can stimulate muscle invasion by leucocytes that have the potential to increase tissue damage or promote tissue growth and repair. In the present investigation, we examined the role of macrophages in muscle injury, repair and regeneration during modified muscle loading. Weight-bearing was removed from the hindlimbs of mice for 10 days followed by reloading through normal ambulation. During the unloading period, soleus muscle fibre cross-section decreased by 38%. Prior to the onset of reloading, mice received a series of intraperitoneal injections of anti-F4/80, which binds a mouse macrophage surface antigen. Although anti-F4/80 injections did not affect macrophage numbers in soleus muscles at 2 days of reloading, macrophages were reduced by 86% at 4 days of reloading. Muscle membrane lysis during the reloading period did not differ at 2 days of reloading between anti-F4/80-treated mice and mice that received isotype control antibody. However, control animals showed large decreases in the number of fibres with membrane lesions at 4 days of reloading, but this membrane repair did not occur in macrophage-depleted mice. Macrophage-depletion also reduced muscle regeneration (indicated by central nucleation) and satellite cell differentiation (indicated by reductions in MyoD-expressing satellite cells) and prevented growth of muscle fibres that normally occurred in control animals between days 2 and 4 of reloading. These findings collectively show that macrophages play a significant role in muscle fibre membrane repair, regeneration and growth during increased muscle use after a period of atrophy.     

Reconstruction of the contractile apparatus of striated muscle. I. Muscle maintained in extension

Summary The ultrastructure of the contractile apparatus was observed in muscles maintained in excessive extension, i.e. in conditions in which an increase takes place in the number of sarcomeres. Rat leg muscles (soleus, extensor digitorum longus and gastrocnemius) were studied, at variable time intervals in the range 3–7 days.Several irregularities were found in the contractile structure. The most frequent were the variability of sarcomere length, the appearance of extra sarcomeres, irregularities of the Z-line (including Z-band streaming) and A-bands of abnormal length. The character of these irregularities depended on the muscle fibre type. Variations of the Z-line were seen mostly within continuously working fibres, especially slow ones, while anomalies in the size of the A-band and variability of the sarcomere length were more pronounced in fast fibres. All these irregularities appearing in the muscles maintained in excessive extension were also occasionally found in control muscles.The reasons for these contractile structure irregularities, and their possible significance for contractile structure reorganization, are discussed.     

Transformation of contraction speed in muscle following cross-reinnervation; dependence on muscle size

Summary The characteristics of isometric contractions and the force-velocity relation were studied in flexor digitorum longus, flexor hallucis longus and soleus muscles of the cat,in situ, at 37° C and with nerve stimulation. The two flexors were identified as typical fast twitch muscles and the soleus as a typical slow twitch muscle. Following self-reinnervation, both fast and slow muscles retained, to a large extent, their basic contraction characteristics. The soleus muscle, when cross-reinnervated with the nerve of either flexor hallucis longus muscle or extensor digitorum longus muscle exhibited a more complete slow-to-fast transformation than when cross-reinnervated with the nerve of flexor digitorum longus muscle. The flexor digitorum longus muscle underwent a greater degree of fast-to-slow transformation than the flexor hallucis longus muscle, when each was cross-reinnervated with the soleus nerve. The data previously reported for sarcomere shortening velocities of the cross-reinnervated muscles in the rat, the rabbit and the cat are reviewed in the light of present findings. It is found that the discrepancies obtained between species and between different muscles in the same species, with respect to the degree of muscle-speed transformation following cross-reinnervation, are correlated with the differences in the size-ratio of the muscles used in the cross-reinnervation procedure.     

Morphological changes in rat hindlimb muscle fibres during recovery from disuse atrophy

AIM: The present study attempted to use HE staining to clarify morphological changes in muscle fibres during recovery from disuse muscle atrophy. METHODS: Disuse muscle atrophy was induced by suspending 7-week-old male Wistar rats by their tails for 5 weeks (hindlimb unloading or HU group). Histological changes in the soleus muscle (SOL) during the recovery process were examined and compared with those in control rats who were raised freely without unloading (C group). RESULTS: Wet muscle mass and muscle cross-sectional area per fibre of SOL in the HU group were 52 +/- 5 and 22 +/- 5% of those in the C group, respectively. Muscle atrophy was largely attributable to decreases in the size of muscle fibres, rather than to muscle fibre damage or loss. Muscle mass in the HU group increased quickly after reloading, but recovery of cross-sectional area per fibre was slow, with mean area in the HU group measuring 69 +/- 10% of that in the C group even after 5 weeks of reloading. After 1, 2 and 5 weeks of reloading, incidences of muscle fibres displaying central nuclei (regenerated muscle fibres) were 7.4 +/- 2.4, 7.2 +/- 6.3 and 19.2 +/- 14.5%, respectively. CONCLUSION: These findings suggest that recovery of muscle fibres atrophied by disuse involves not only growth of atrophied muscle fibres, but also regeneration of muscle fibres. Cross-sectional areas recovery of atrophied muscle fibres thus continues after increases of muscle mass.     

Electromyographic analysis of hip adductor muscles during incremental fatiguing pedaling exercise

The purpose of this study was to investigate activity of hip adductor muscles over time and during a representative crank cycle in fatiguing pedaling. Sixteen healthy men performed incremental pedaling exercise until exhaustion. During the exercise, surface electromyogram (EMG) was detected from adductor magnus (AM), adductor longus (AL), and selected thigh muscles. Temporal changes to normalized EMG in AM muscle resembled those in vastus lateralis (VL) muscle, whereas those in AL muscle showed later onset of increase from baseline compared with AM and VL muscles. During a representative crank cycle, the same level of normalized EMG was found between propulsive and pulling phases for AM muscle, whereas muscle activation of AL muscle during the pulling phase was statistically significant higher than that during the propulsive phase. We concluded that AM and AL muscles were gradually recruited over time during fatiguing pedaling exercise, but their temporal change and activation phases were not completely the same.     

Macrophage invasion does not contribute to muscle membrane injury during inflammation

Previous observations have shown that neutrophil invasion precedes macrophage invasion during muscle inflammation and that peak muscle injury is observed at the peak of ED1+ macrophage invasion. We tested the hypothesis that neutrophil invasion causes subsequent invasion by ED1+ macrophages and that ED1+ macrophages then contribute significantly to muscle membrane injury during modified muscle use. Rat hindlimbs were unloaded for 10 days followed by reloading by normal ambulation to induce inflammation. Membrane injury was measured by assaying Evans blue-bound serum protein influx through membrane lesions. Muscle neutrophil populations increased significantly during the first 2 h of reloading but ED1+ macrophages did not increase until 24 h. Neutrophil invasion was uncoupled from subsequent macrophage invasion by reloading rat hindlimbs for 2 h to cause neutrophil invasion, followed by resuspension for hours 2-24. This produced similar increases in neutrophil concentration as measured in muscles continuously reloaded for 24 h without causing an increase in macrophages. However, resuspension did not reduce the extent of muscle damage compared with that occurring in muscles that were reloaded continuously for 24 h. Thus, muscle invasion by neutrophils is not sufficient to cause invasion by ED1+ macrophages. In addition, muscle membrane injury that occurs during reloading is independent of invasion by ED1+ macrophages.