Nox4 Is Dispensable for Exercise Induced Muscle Fibre Switch

Introduction

By producing H₂O₂, the NADPH oxidase Nox4 is involved in differentiation of mesenchymal cells. Exercise alters the composition of slow and fast twitch fibres in skeletal. Here we hypothesized that Nox4 contributes to exercise-induced adaptation such as changes in muscle metabolism or muscle fibre specification and studied this in wildtype and Nox4-/- mice.

Results

Exercise, as induced by voluntary running in a running wheel or forced running on a treadmill induced a switch from fast twitch to intermediate fibres. However the induced muscle fibre switch was similar between Nox4-/- and wildtype mice. The same held true for exercise-induced expression of PGC1α or AMPK activation. Both are increased in response to exercise, but with no difference was observed between wildtype and Nox4-/- mice.

Conclusion

Thus, exercise-induced muscle fibre switch is Nox4-independent.     

Effects of hypothyroidism on myosin heavy chain composition and fibre types of fast skeletal muscles in a small marsupial, Antechinus flavipes

Effects of drug-induced hypothyroidism on myosin heavy chain (MyHC) content and fibre types of fast skeletal muscles were studied in a small marsupial, Antechinus flavipes. SDS-PAGE of MyHCs from the tibialis anterior and gastrocnemius revealed four isoforms, 2B, 2X, 2A and slow, in that order of decreasing abundance. After 5 weeks treatment with methimazole, the functionally fastest 2B MyHC significantly decreased, while 2X, 2A and slow MyHCs increased. Immunohistochemistry using monospecific antibodies to each of the four MyHCs revealed decreased 2b and 2x fibres, and increased 2a and hybrid fibres co-expressing two or three MyHCs. In the normally homogeneously fast superficial regions of these muscles, evenly distributed slow-staining fibres appeared, resembling the distribution of slow primary myotubes in fast muscles during development. Hybrid fibres containing 2A and slow MyHCs were virtually absent. These results are more detailed but broadly similar to the earlier studies on eutherians. We hypothesize that hypothyroidism essentially reverses the effects of thyroid hormone on MyHC gene expression of muscle fibres during myogenesis, which differ according to the developmental origin of the fibre: it induces slow MyHC expression in 2b fibres derived from fast primary myotubes, and shifts fast MyHC expression in fibres of secondary origin towards 2A, but not slow, MyHC.     

Sexual dimorphism in histological characteristics and contractility of the iliofibularis muscle in the lizard Sceloporus torquatus

The iliofibularis is a hindlimb muscle used in lizard locomotion that is composed of at least three types of fibres: fast‐twitch‐glycolytic (FG), fast‐twitch‐oxidative‐glycolytic fibre (FOG) and slow‐twitch‐oxidative (SO). The striated skeletal muscle is a highly plastic tissue undergoing phenotypic change in response to activity. The lizard Sceloporus torquatus has sexual differences associated with microhabitat use, which can be reflected in the physiology and anatomy of the muscle, and thus, in our study, we analysed the morphological and contractile characteristics of the iliofibularis muscle (IF) of S. torquatus males and females. We found a larger prevalence of FOG compared with FG and SO fibres in the muscle of both sexes. We also found that males show larger areas of the three types of fibres, develop greater strength but also faster fatigue than females, suggesting that strength is a key functional feature that enables males to perform faster movements (but for shorter periods), associated with the demands of territoriality.     

Muscle fibre types and metabolism in post-larval and adult stages of notothenioid fish

Summary A histochemical study was carried out on muscle fibre types in the myotomes of post-larval and adult stages of seven species of notothenioid fish. There was little interspecific variation in the distribution of muscle fibre types in post-larvae. Slow fibres (diameter range 15–60 m) which stained darkly for succinic dehydrogenase activity (SDHase) formed a superficial layer 1–2 fibres thick around the entire lateral surface of the trunk. In all species a narrow band of very small diameter fibres (diameter range 5–62 m), with only weak staining activity, occurred between the skin and slow fibre layer. These have the characteristics of tonic fibres found in other teleosts. The remainder of the myotome was composed of fast muscle fibres (diameter range 9–75 m), which stain weakly for SDHase, -glycerophosphate dehydrogenase, glycogen and lipid. Slow muscle fibres were only a minor component of the trunk muscles of adult stages of the pelagic species Champsocephalus gunnari and Pseudochaenichthys georgianus, consistent with a reliance on pectoral fin swimming during sustained activity. Of the other species examined only Psilodraco breviceps and Notothenia gibberifrons had more than a few percent of slow muscle in the trunk (20%–30% in posterior myotomes), suggesting a greater involvement of sub-carangiform swimming at cruising speeds. The ultrastructure of slow fibres from the pectoral fin adductor and myotomal muscles of a haemoglobinless (P. georgianus) and red-blooded species (P. breviceps), both active swimmers, were compared. Fibres contained loosely packed, and regularly shaped myofibrils numerous mitochondria, glycogen granules and occasional lipid droplets. Mitochondria occupied >50% of fibre volume in the haemoglobinless species P. georgianus, each myofibril was surrounded by one or more mitochondria with densely packed cristae. No significant differences, however, were found in mean diameter between fibres from red-blooded and haemoglobinless species. The activities of key enzymes of energy metabolism were determined in the slow (pectoral) and fast (myotomal) muscles of N. gibberifrons. In contrast to other demersal Antarctic fish examined, much higher glycolytic activities were found in fast muscle fibres, probably reflecting greater endurance during burst swimming.     

Energy metabolism of fast- and slow-twitch skeletal muscle in the rat: Thyroid hormone induced changes

Summary Male Wistar rats were made hypothyroid or hyperthyroid over a period of six weeks, by administration of carbimazole or triiodothyronine (T₃). Serial frozen sections of soleus and extensor digitorum longus (EDL) muscle were stained histochemically for myosin ATPase, succinic dehydrogenase and phosphorylase. Muscle fibres were classified as either slow twitch oxidative (SO), fast twitch oxidative glycolytic (FOG) or fast twitch glycolytic (FG). In addition the activities of phosphorylase, phosphofructokinase (PFK), fructose-1,6-diphosphatase (FDP), lactate dehydrogenase (LDH), hexokinase, citrate synthetase, cytochrome oxidase, 3-hydroxyacyl-CoA dehydrogenase (HAD) and 5-AMP aminohydrolase were measured in both muscles.Increasing plasma levels of T₃ are associated with marked alterations in the fibre type populations in both muscles. In the soleus there is conversion of SO to FOG fibres while in the EDL, FG fibres are converted to FOG fibres. The quantitative changes in metabolic enzyme activity however, are in the main restricted to the soleus. Increased T₃ levels result in an increased capacity for the aerobic metabolism of both fat and carbohydrate and an increase in anaerobic glycolytic activity in the soleus muscle which parallels the change in fibre types. However, the extent of these increases cannot be explained solely on this basis and there is also an overall increase in aerobic activity in all fibres including slow oxidative ones. It is concluded that the effects of thyroid hormone on muscle phenotype and respiratory capacity involve both primary and secondary sites of action and the possible mechanisms are discussed.Abbreviations EDL extensor digitorum longus - FDP fructose-1,6-diphosphatase - FG fast twitch glycolytic - FOG fast twitch oxidative glycolytic - HAD 3-hydroxyacyl-CoA-dehydrogenase - LDH lactate dehydrogenase - PFK phosphofructokinase - SO slow twitch oxidative - T ₃ triiodothyronine - T ₄ thyroxine     

Expression of calcium channels in Xenopus oocyte injected with crab skeletal muscle fibre mRNA

Summary— Using the whole cell voltage-clamp technique and a Cl free and Na free Ba methane sulfonate solution, stage V and VI Xenopus oocytes demonstrated a Ba current (endogenous component) with a peak amplitude average of 6 nA (6 ± 2 nA). When oocytes were injected with crustacean skeletal muscle mRNA, an additional component of I_Ba could be detected (exogenous I_Ba). The latter current could be distinguished from the native one by several electrophysiological means: a peak amplitude average of 90 nA (90 ± 4 nA), activation potential threshold, steady state inactivation properties and sensitivity to Ca blockers. As shown by Jdaïâa and Guilbault in crustacean skeletal muscle fibres, exogenous I_Ba could be divided into two components: a “fast component” and a “slow component” probably passing through two types of Ca channels (fast and slow) since the peak Ba current voltage relationship was biphasic and the fast component of exogenous I_Ba was less sensitive than the slow to nifedipine. The features of the newly synthesized channels incorporated in the Xenopus oocyte membrane suggest that they may be associated with fast and slow channels, previously described in many preparations, particularly in crustacean skeletal muscle fibres.     

Comparative analysis of mouse skeletal muscle fibre type composition and contractile responses to calcium channel blocker

Background  

In this study, we examined the correlation between excitation-contraction coupling characteristics and skeletal muscle fibre type by (1) localizing the distribution of dihydropyridine receptor (DHPR) protein and (2) comparing the effect of DHPR blocker on muscles with different fibre type composition, in order to better understand the differences between contractile phenotypes of fibres and to explain the contradictory reports to date on the interaction of dihydropyridines with skeletal muscle isoform of DHPR.     

The localization of the different forms of troponin I in skeletal and cardiac muscle cells

1. 1. Antibodies raised against troponin I isolated from human cardiac and rabbit fast and slow skeletal muscles have been shown to be specific for the polymorphic forms of troponin I against which they were raised, i.e. they are tissue specific.

2. 2. These antibodies reacted with the polymorphic forms of troponin I, against which they were raised, that are present in tissues of other species such as the rhesus monkey, hamster and rat, i.e. they were species non-specific.

3. 3. Using the immunoperoxidase staining technique it has been shown that the fast and slow forms of troponin I are located in different cells in virtually all adult normal muscles examined.

4. 4. By comparison of the ATPase staining of skeletal muscle sections at pH 9.4 and 4.2 it is concluded that the fast form of troponin I is located in type II fibres and the slow form in type I fibres.

5. 5. It is suggested that immunoperoxidase staining with the antibodies to the fast and slow forms of troponin I provides an unambiguous new method of muscle fibre typing.

     

The influence of the <Emphasis Type="Italic">PRKAG3</Emphasis> mutation on glycogen,enzyme activities and fibre types in different skeletal muscles of exercise trained pigs

Background  

AMP-activated protein kinase (AMPK) plays an important role in the regulation of glucose and lipid metabolism in skeletal muscle. Many pigs of Hampshire origin have a naturally occurring dominant mutation in the AMPK γ3 subunit. Pigs carrying this PRKAG3 (R225Q) mutation have, compared to non-carriers, higher muscle glycogen levels and increased oxidative capacity in m. longissimus dorsi, containing mainly type II glycolytic fibres. These metabolic changes resemble those seen when muscles adapt to an increased physical activity level. The aim was to stimulate AMPK by exercise training and study the influence of the PRKAG3 mutation on metabolic and fibre characteristics not only in m. longissimus dorsi, but also in other muscles with different functions.     

Maged1, a new regulator of skeletal myogenic differentiation and muscle regeneration

Background  

In normal adult skeletal muscle, cell turnover is very slow. However, after an acute lesion or in chronic pathological conditions, such as primary myopathies, muscle stem cells, called satellite cells, are induced to proliferate, then withdraw definitively from the cell cycle and fuse to reconstitute functional myofibers.     

Fine structure and metabolism of multiply innervated fast muscle fibres in teleost fish

Summary Both the fast and slow muscle fibres of advanced teleost fish are multiply innervated. The fraction of slow-fibre volume occupied by mitochondria is 31.3%, 25.5% and 24.6%, respectively, for the myotomal muscles of brook trout (Salvelinus fontinalis), crucian carp (Carassius carassius), and plaice (Pleuronectes platessa), respectively. The corresponding figures for the fast muscles of these species are 9.3%, 4.6% and 2.0%, respectively. Cytochrome-oxidase and citrate-synthetase activities in the fast muscles of 9 species of teleost range from 0.20–0.93 moles substrate utilised, g wet weight muscle^-1 min^-1 (at 15° C) or around 4–17% of that of the corresponding slow fibres. Ultrastructural analyses reveal a marked heterogeneity within the fast-fibre population. For example, the fraction of fibres with <1% or >10% mitochondria is 0,4,42% and 36, 12 and 0%, respectively, for trout, carp and plaice. In general, small fibres (<500 m²) have the highest and large fibres (>1,500 m²) the lowest mitochondrial densities. The complexity of mitochondrial cristae is reduced in fast compared to slow fibres.Hexokinase activities range from 0.4–2.5 in slow and from 0.08–0.7 moles, g wet weight^-1 min^-1 in fast muscles, indicating a wide variation in their capacity for aerobic glucose utilisation. Phosphofructokinase activities are 1.2 to 3.6 times higher in fast than slow muscles indicating a greater glycolytic potential. Lactate dehydrogenase activities are not correlated with either the predicted anaerobic scopes for activity or the anoxic tolerances of the species studied. The results indicate a considerable variation in the aerobic capacities and principal fuels supporting activity among the fast muscles of different species. Brook trout and crucian carp are known to recruit fast fibres at low swimming speeds. For these species the aerobic potential of the fast muscle is probably sufficient to meet the energy requirements of slow swimming.     

Relationship between myoglobin and succinate dehydrogenase in mouse soleus and plantaris muscle fibres

Summary This report describes a quantitative histochemical study of myoglobin in skeletal muscle fibres. The muscle fibres were classified as fast or slow on the basis of their quantitative myofibrillar ATPase histochemistry. A large range of myoglobin absorbance values was found among fast skeletal muscle fibres. This range was relatively small among slow fibres. The concentrations of myoglobin and the activities of succinate dehydrogenase in individual muscle fibres in serial sections are weakly correlated in both the mouse soleus and plantaris muscle. The myoglobin concentration is higher in fast and slow oxidative soleus muscle fibres and the succinate dehydrogenase activity in these fibres is lower than in oxidative plantaris muscle fibres in the same range of cross-sectional area.     

The production of fluorescent transgenic trout to study <Emphasis Type="Italic">in vitro</Emphasis> myogenic cell differentiation

Background  

Fish skeletal muscle growth involves the activation of a resident myogenic stem cell population, referred to as satellite cells, that can fuse with pre-existing muscle fibers or among themselves to generate a new fiber. In order to monitor the regulation of myogenic cell differentiation and fusion by various extrinsic factors, we generated transgenic trout (Oncorhynchus mykiss) carrying a construct containing the green fluorescent protein reporter gene driven by a fast myosin light chain 2 (MlC2f) promoter, and cultivated genetically modified myogenic cells derived from these fish.     

Effects of Cannabinoids on Caffeine Contractures in Slow and Fast Skeletal Muscle Fibers of the Frog

The effect of cannabinoids on caffeine contractures was investigated in slow and fast skeletal muscle fibers using isometric tension recording. In slow muscle fibers, WIN 55,212-2 (10 and 5 μM) caused a decrease in tension. These doses reduced maximum tension to 67.43 ± 8.07% (P = 0.02, n = 5) and 79.4 ± 14.11% (P = 0.007, n = 5) compared to control, respectively. Tension-time integral was reduced to 58.37 ± 7.17% and 75.10 ± 3.60% (P = 0.002, n = 5), respectively. Using the CB₁ cannabinoid receptor agonist ACPA (1 μM) reduced the maximum tension of caffeine contractures by 68.70 ± 11.63% (P = 0.01, n = 5); tension-time integral was reduced by 66.82 ± 6.89% (P = 0.02, n = 5) compared to controls. When the CB₁ receptor antagonist AM281 was coapplied with ACPA, it reversed the effect of ACPA on caffeine-evoked tension. In slow and fast muscle fibers incubated with the pertussis toxin, ACPA had no effect on tension evoked by caffeine. In fast muscle fibers, ACPA (1 μM) also decreased tension; the maximum tension was reduced by 56.48 ± 3.4% (P = 0.001, n = 4), and tension-time integral was reduced by 57.81 ± 2.6% (P = 0.006, n = 4). This ACPA effect was not statistically significant with respect to the reduction in tension in slow muscle fibers. Moreover, we detected the presence of mRNA for the cannabinoid CB₁ receptor on fast and slow skeletal muscle fibers, which was significantly higher in fast compared to slow muscle fiber expression. In conclusion, our results suggest that in the slow and fast muscle fibers of the frog cannabinoids diminish caffeine-evoked tension through a receptor-mediated mechanism.     

Cardiac and skeletal muscle troponin I isoforms are encoded by a dispersed gene family on mouse Chromosomes 1 and 7

We mapped the locations of the genes encoding the slow skeletal muscle, fast skeletal muscle, and cardiac isoforms of troponin I (Tnni) in the mouse genome by interspecific hybrid backcross analysis of species-specific (C57BL/6 vs Mus spretus) restriction fragment length polymorphisms (RFLPs). The slow skeletal muscle troponin I locus (Tnni1) mapped to Chromosome (Chr) 1. The fast skeletal muscle troponin I locus (Tnni2), mapped to Chr 7, approximately 70 cM from the centromere. The cardiac troponin I locus (Tnni3) also mapped to Chr 7, approximately 5–10 cM from the centromere and unlinked to the fast skeletal muscle troponin I locus. Thus, the troponin I gene family is dispersed in the mouse genome. Received: 10 May 1995 / Accepted: 1 September 1995     

Variance in multiplex suspension array assays: microsphere size variation impact

Background

Skeletal muscle fibres contain transverse tubular (t-tubule) networks that allow electrical signals to rapidly propagate into the fibre. These electrical signals are generated by the transport of ions across the t-tubule membranes and this can result in significant changes in ion concentrations within the t-tubules during muscle excitation. During periods of repeated high-frequency activation of skeletal muscle the t-tubule K⁺ concentration is believed to increase significantly and diffusive K⁺ transport from the t-tubules into the interstitial space provides a mechanism for alleviating muscle membrane depolarization. However, the tortuous nature of the highly branched space-filling t-tubule network impedes the diffusion of material through the network. The effective diffusion coefficient for ions in the t-tubules has been measured to be approximately five times lower than in free solution, which is significantly different from existing theoretical values of the effective diffusion coefficient that range from 2–3 times lower than in free solution. To resolve this discrepancy, in this paper we study the process of diffusion within electron microscope scanned sections of the skeletal muscle t-tubule network using mathematical modelling and computer simulation techniques. Our model includes t-tubule geometry, tautness, hydrodynamic and non-planar network factors.

Results

Using our model we found that the t-tubule network geometry reduced the K⁺ diffusion coefficient to 19–27% of its value in free solution, which is consistent with the experimentally observed value of 21% and is significantly smaller than existing theoretical values that range from 32–50%. We also found that diffusion in the t-tubules is anomalous for skeletal muscle fibres with a diameter of less than approximately 10–20 μm as a result of obstructed diffusion. We also observed that the [K⁺] within the interior of the t-tubule network during high-frequency activation is greater for fibres with a larger diameter. Smaller skeletal muscle fibres are therefore more resistant to membrane depolarization. Because the t-tubule network is anisotropic and inhomogeneous, we also found that the [K⁺] distribution generated within the network was irregular for fibres of small diameter.

Conclusion

Our model explains the measured effective diffusion coefficient for ions in skeletal muscle t-tubules.     

Polymorphic forms of troponin T and troponin C and their localization in striated muscle cell types

1. 1. Immunochemical studies have shown that the major forms of troponin T present in fast skeletal, slow skeletal and cardiac muscles are different proteins.

2. 2. Similar studies indicate that the major form of troponin C present in fast skeletal muscles differs from troponin C present in slow skeletal and cardiac muscle cells. The forms of troponin C present in slow skeletal and cardiac muscles are immunochemically very similar.

3. 3. The antibodies to the polymorphic forms of troponin T and troponin C are specific for the muscle type, except in the case of the slow skeletal and cardiac muscle forms of troponin C.

4. 4. By the immunoperoxidase technique, it has been shown that the fast skeletal muscle troponin T is localized in type II cells and slow skeletal muscle troponin T in type I cells.

5. 5. Fast skeletal muscle troponin C is present in type II cells and a different troponin C, identified by its reaction with the antibody against cardiac troponin C, is present in type I cells.

6. 6. It is concluded that in normal adult skeletal muscle, fast muscle forms of troponin I, troponin T and troponin C are present together as a homocomplex in type II cells and the slow muscle forms exist as an analagous homocomplex in type I cells.