Skeletal Muscle Of Control Research Articles

Running improves muscle mass by activating autophagic flux and inhibiting ubiquitination degradation in mdx mice

BackgroundExercise therapy can improve muscle mass, strengthen muscle and cardiorespiratory function, and may be an excellent adjunctive treatment option for Duchenne muscular dystrophy. MethodsThis article investigates the effects of 10 weeks of treadmill training on skeletal muscle in control and mdx mice. Hematoxylin and eosin (H&E) staining was used to detect the morphometry of skeletal muscle; the grip strength test, suspension test, and rotarod test were used to detect limb muscle strength of mice, and Aurora Scientific Instruments were used to detect in vivo Muscle Stimulation Measuring Maximum Force of pre-fatigue and post-fatigue. The expression levels of myogenic proteins, ubiquitination markers, autophagy pathway proteins, and the proportion of different muscle fiber types were detected. ResultsThe experimental results show that running exercise can significantly improve the muscle mass of mdx mice, promote muscle strength, endurance, and anti-fatigue ability, reverse the pathological state of skeletal muscle destruction in mdx mice, and promote muscle regeneration. WB experiments showed that running inhibited the ubiquitination and degradation of muscle protein in mdx mice, inhibited AKT activation, decreased phosphorylated FoxO1 and FoxO3a, and restored the suppressed autophagic flux. Running enhances muscle strength and endurance by comprehensively promoting the expression of Myh1/2/4/7 fast and slow muscle fibers in mdx mice. ConclusionsRunning can inhibit the degradation of muscle protein in mdx mice, and promote the reuse and accumulation of proteins, thereby slowing down muscle loss. Running improves skeletal muscle mass by activating autophagic flux and inhibiting ubiquitination degradation in mdx mice.

Mechanisms Underlying Absent Training-Induced Improvement in Insulin Action in Lean, Hyperandrogenic Women With Polycystic Ovary Syndrome.

Women with polycystic ovary syndrome (PCOS) have been shown to be less insulin sensitive compared with control (CON) women, independent of BMI. Training is associated with molecular adaptations in skeletal muscle, improving glucose uptake and metabolism in both healthy individuals and patients with type 2 diabetes. In the current study, lean hyperandrogenic women with PCOS (n = 9) and healthy CON women (n = 9) completed 14 weeks of controlled and supervised exercise training. In CON, the training intervention increased whole-body insulin action by 26% and insulin-stimulated leg glucose uptake by 53% together with increased insulin-stimulated leg blood flow and a more oxidative muscle fiber type distribution. In PCOS, no such changes were found, despite similar training intensity and improvements in VO2max In skeletal muscle of CON but not PCOS, training increased GLUT4 and HKII mRNA and protein expressions. These data suggest that the impaired increase in whole-body insulin action in women with PCOS with training is caused by an impaired ability to upregulate key glucose-handling proteins for insulin-stimulated glucose uptake in skeletal muscle and insulin-stimulated leg blood flow. Still, other important benefits of exercise training appeared in women with PCOS, including an improvement of the hyperandrogenic state.

Transduction Efficiency of Adeno-Associated Virus Serotypes After Local Injection in Mouse and Human Skeletal Muscle.

The adeno-associated virus (AAV) vector is an efficient tool for gene delivery in skeletal muscle. AAV-based therapies show promising results for treatment of various genetic disorders, including muscular dystrophy. These dystrophies represent a heterogeneous group of diseases affecting muscles and typically characterized by progressive skeletal muscle wasting and weakness and the development of fibrosis. The tropism of each AAV serotype has been extensively studied using systemic delivery routes, but very few studies have compared their transduction efficiency through direct intramuscular injection. Yet, in some muscular dystrophies, where only a few muscles are primarily affected, a local intramuscular injection to target these muscles would be the most appropriate route. A comprehensive comparison between different recombinant AAV (rAAV) serotypes is therefore needed. In this study, we investigated the transduction efficiency of rAAV serotypes 1-10 by local injection in skeletal muscle of control C57BL/6 mice. We used a CMV-nls-LacZ reporter cassette allowing nuclear expression of LacZ to easily localize targeted cells. Detection of β-galactosidase activity on muscle cryosections demonstrated that rAAV serotypes 1, 7, 8, 9, and 10 were more efficient than the others, with rAAV9 being the most efficient in mice. Furthermore, using a model of human muscle xenograft in immunodeficient mice, we observed that in human muscle, rAAV8 and rAAV9 had similar transduction efficiency. These findings demonstrate for the first time that the human muscle xenograft can be used to evaluate AAV-based therapeutical approaches in a human context.

Purification and characterization of a urea sensitive lactate dehydrogenase from skeletal muscle of the African clawed frog, Xenopus laevis.

The African clawed frog, Xenopus laevis endures whole body dehydration which can increase its reliance on anaerobic glycolysis for energy production. This makes the regulation of the terminal enzyme of glycolysis, lactate dehydrogenase (LDH), crucial to stress survival. We investigated the enzymatic properties and posttranslational modification state of purified LDH from the skeletal muscle of control and dehydrated (30% total body water loss) X. laevis. LDH from the muscle of dehydrated frogs showed a 93% reduction in phosphorylation on threonine residues and an 80% reduction of protein nitrosylation. LDH from dehydrated muscle also showed a 74% lower Vmax in the pyruvate oxidizing direction and a 78% decrease in Vmax in the lactate reducing direction along with a 33% lower Km for pyruvate and a 40% higher Km for lactate. In the presence of higher levels of urea and molecular crowding by polyethylene glycol, used to mimic conditions in the cells of dehydrated animals, the Km values of control and dehydrated LDH demonstrated opposite responses. In the pyruvate oxidizing direction, control muscle LDH was unaffected by these additives, whereas the affinity for pyruvate dropped further for LDH from dehydrated muscle. The opposite effect was more pronounced in the lactate reducing direction as control LDH showed an increased affinity for lactate, whereas LDH from dehydrated animals showed a further reduction in affinity. The physiological consequences of dehydration-induced LDH regulation appear to poise the enzyme towards lactate production when urea levels are high and lactate catabolism when urea levels are low, perhaps helping to maintain glycolysis under dehydrating conditions whilst providing for the ability to recycle lactate upon rehydration.

Purification and characterization of skeletal muscle pyruvate kinase from the hibernating ground squirrel, Urocitellus richardsonii: potential regulation by posttranslational modification during torpor.

Ground squirrel torpor during winter hibernation is characterized by numerous physiological and biochemical changes, including alterations to fuel metabolism. During torpor, many tissues switch from carbohydrate to lipid catabolism, often by regulating key enzymes within glycolytic and lipolytic pathways. This study investigates the potential regulation of pyruvate kinase (PK), a key member of the glycolytic pathway, within the skeletal muscle of hibernating ground squirrels. PK was purified from the skeletal muscle of control and torpid Richardson's ground squirrels, and PK kinetics, structural stability, and posttranslational modifications were subsequently assessed. Torpid PK displayed a nearly threefold increase in K m PEP as compared to control PK when assayed at 5°C. ProQ Diamond phosphoprotein staining as well as phospho-specific western blots indicated that torpid PK was significantly more phosphorylated than the euthermic control. PK from the torpid condition was also shown to possess nearly twofold acetyl content as compared to control PK. In conclusion, skeletal muscle PK from the Richardson's ground squirrel may be regulated posttranslationally between the euthermic and torpid states, and this may inhibit PK functioning during torpor in accordance with the decrease in glycolytic rate during dormancy.

Early energy metabolism-related molecular events in skeletal muscle of diabetic rats: The effects of l-arginine and SOD mimic

Considering the vital role of skeletal muscle in control of whole-body metabolism and the severity of long-term diabetic complications, we aimed to reveal the molecular pattern of early diabetes-related skeletal muscle phenotype in terms of energy metabolism, focusing on regulatory mechanisms, and the possibility to improve it using two redox modulators, l-arginine and superoxide dismutase (SOD) mimic. Alloxan-induced diabetic rats (120 mg/kg) were treated with l-arginine or the highly specific SOD mimic, M40403, for 7 days. As appropriate controls, non-diabetic rats received the same treatments. We found that l-arginine and M40403 restored diabetes-induced impairment of phospho-5′-AMP-activated protein kinase α (AMPKα) signaling by upregulating AMPKα protein itself and its downstream effectors, peroxisome proliferator-activated receptor-γ coactivator-1α and nuclear respiratory factor 1. Also, there was a restitution of the protein levels of oxidative phosphorylation components (complex I, complex II and complex IV) and mitofusin 2. Furthermore, l-arginine and M40403 induced translocation of glucose transporter 4 to the membrane and upregulation of protein of phosphofructokinase and acyl coenzyme A dehydrogenase, diminishing negative diabetic effects on limiting factors of glucose and lipid metabolism. Both treatments abolished diabetes-induced downregulation of sarcoplasmic reticulum calcium-ATPase proteins (SERCA 1 and 2). Similar effects of l-arginine and SOD mimic treatments suggest that disturbances in the superoxide/nitric oxide ratio may be responsible for skeletal muscle mitochondrial and metabolic impairment in early diabetes. Our results provide evidence that l-arginine and SOD mimics have potential in preventing and treating metabolic disturbances accompanying this widespread metabolic disease.

The hibernating South American marsupial, Dromiciops gliroides, displays torpor-sensitive microRNA expression patterns.

When faced with adverse environmental conditions, the marsupial Dromiciops gliroides uses either daily or seasonal torpor to support survival and is the only known hibernating mammal in South America. As the sole living representative of the ancient Order Microbiotheria, this species can provide crucial information about the evolutionary origins and biochemical mechanisms of hibernation. Hibernation is a complex energy-saving strategy that involves changes in gene expression that are elicited in part by microRNAs. To better elucidate the role of microRNAs in orchestrating hypometabolism, a modified stem-loop technique and quantitative PCR were used to characterize the relative expression levels of 85 microRNAs in liver and skeletal muscle of control and torpid D. gliroides. Thirty-nine microRNAs were differentially regulated during torpor; of these, 35 were downregulated in liver and 11 were differentially expressed in skeletal muscle. Bioinformatic analysis predicted that the downregulated liver microRNAs were associated with activation of MAPK, PI3K-Akt and mTOR pathways, suggesting their importance in facilitating marsupial torpor. In skeletal muscle, hibernation-responsive microRNAs were predicted to regulate focal adhesion, ErbB, and mTOR pathways, indicating a promotion of muscle maintenance mechanisms. These tissue-specific responses suggest that microRNAs regulate key molecular pathways that facilitate hibernation, thermoregulation, and prevention of muscle disuse atrophy.

A hydrogen peroxide safety valve: The reversible phosphorylation of catalase from the freeze-tolerant North American wood frog, Rana sylvatica

BackgroundThe North American wood frog, Rana sylvatica, endures whole body freezing while wintering on land and has developed multiple biochemical adaptations to elude cell/tissue damage and optimize its freeze tolerance. Blood flow is halted in the frozen state, imparting both ischemic and oxidative stress on cells. A potential build-up of H2O2 may occur due to increased superoxide dismutase activity previously discovered. The effect of freezing on catalase (CAT), which catalyzes the breakdown of H2O2 into molecular oxygen and water, was investigated as a result. MethodsThe present study investigated the purification and kinetic profile of CAT in relation to the phosphorylation state of CAT from the skeletal muscle of control and frozen R. sylvatica. ResultsCatalase from skeletal muscle of frozen wood frogs showed a significantly higher Vmax (1.48 fold) and significantly lower Km for H2O2 (0.64 fold) in comparison to CAT from control frogs (5°C acclimated). CAT from frozen frogs also showed higher overall phosphorylation (1.73 fold) and significantly higher levels of phosphoserine (1.60 fold) and phosphotyrosine (1.27 fold) compared to control animals. Phosphorylation via protein kinase A or the AMP-activated protein kinase significantly decreased the Km for H2O2 of CAT, whereas protein phosphatase 2B or 2C action significantly increased the Km. ConclusionThe physiological consequence of freeze-induced CAT phosphorylation appears to improve CAT function to alleviate H2O2 build-up in freezing frogs. General significanceAugmented CAT activity via reversible phosphorylation may increase the ability of R. sylvatica to overcome oxidative stress associated with ischemia.

MicroRNA regulation in heart and skeletal muscle over the freeze-thaw cycle in the freeze tolerant wood frog.

The North American wood frog, Rana sylvatica, is one of just a few anuran species that tolerates whole body freezing during the winter and has been intensely studied to identify the biochemical adaptations that support freeze tolerance. Among these adaptations is the altered expression of many genes, making freeze-responsive changes to gene regulatory mechanisms a topic of interest. The present study focuses on the potential involvement of microRNAs as one such regulatory mechanism and aims to better understand freeze/thaw stress-induced microRNA responses in the freeze-tolerant wood frog. Using quantitative PCR, relative levels of 53 microRNAs were measured in heart and skeletal muscle of control, 24 h frozen, and 8 h thawed frogs. MicroRNAs showed tissue specific expression patterns: 21 microRNAs decreased in the heart during thawing, whereas 16 microRNAs increased during freezing stress in skeletal muscle. These findings suggest that select genes may be activated and suppressed in heart and skeletal muscle, respectively, in response to freezing. Bioinformatics analysis using the DIANA miRPath program (v.2.0) predicted that the differentially expressed microRNAs may collectively regulate tissue-specific cellular pathways to promote survival of wood frogs undergoing freezing and thawing.

Overload-induced skeletal muscle hypertrophy is not impaired in STZ-diabetic rats.

The aim of this study was to evaluate the effect of overload-induced hypertrophy on extensor digitorum longus (EDL) and soleus muscles of streptozotocin-induced diabetic rats. The overload-induced hypertrophy and absolute tetanic and twitch forces increases in EDL and soleus muscles were not different between diabetic and control rats. Phospho-Akt and rpS6 contents were increased in EDL muscle after 7 days of overload and returned to the pre-overload values after 30 days. In the soleus muscle, the contents of total and phospho-Akt and total rpS6 were increased in both groups after 7 days. The contents of total Akt in controls and total rpS6 and phospho-Akt in the diabetic rats remained increased after 30 days. mRNA expression after 7 days of overload in the EDL muscle of control and diabetic animals showed an increase in MGF and follistatin and a decrease in myostatin and Axin2. The expression of FAK was increased and of MuRF-1 and atrogin-1 decreased only in the control group, whereas Ankrd2 expression was enhanced only in diabetic rats. In the soleus muscle caused similar changes in both groups: increase in FAK and MGF and decrease in Wnt7a, MuRF-1, atrogin-1, and myostatin. Differences between groups were observed only in the increased expression of follistatin in diabetic animals and decreased Ankrd2 expression in the control group. So, insulin deficiency does not impair the overload-induced hypertrophic response in soleus and EDL muscles. However, different mechanisms seem to be involved in the comparable hypertrophic responses of skeletal muscle in control and diabetic animals.

Fine-structural distribution of MMP-2 and MMP-9 activities in the rat skeletal muscle upon training: a study by high-resolution in situ zymography

Matrix metalloproteinases (MMPs) are key regulators of extracellular matrix remodeling, but have also important intracellular targets. The purpose of this study was to examine the activity and subcellular localization of the gelatinases MMP-2 and MMP-9 in skeletal muscle of control and physically trained rats. In control hind limb muscle, the activity of the gelatinases was barely detectable. In contrast, after 5 days of intense exercise, in Soleus (Sol), but not Extensor digitorum longus (EDL) muscle, significant upregulation of gelatinolytic activity in myofibers was observed mainly in the nuclei, as assessed by high resolution in situ zymography. The nuclei of quiescent satellite cells did not contain the activity. Within the myonuclei, the gelatinolytic activity colocalized with an activated RNA Polymerase II. Also in Sol, but not in EDL, there were few foci of mononuclear cells with strongly positive cytoplasm, associated with apparent necrotic myofibers. These cells were identified as activated satellite cells/myoblasts. No extracellular gelatinase activity was observed. Gel zymography combined with subcellular fractionation revealed training-related upregulation of active MMP-2 in the nuclear fraction, and increase of active MMP-9 in the cytoplasmic fraction of Sol. Using RT-PCR, selective increase in MMP-9 mRNA was observed. We conclude that training activates nuclear MMP-2, and increases expression and activity of cytoplasmic MMP-9 in Sol, but not in EDL. Our results suggest that the gelatinases are involved in muscle adaptation to training, and that MMP-2 may play a novel role in myonuclear functions.Electronic supplementary materialThe online version of this article (doi:10.1007/s00418-012-0940-5) contains supplementary material, which is available to authorized users.

The effect of long-term DHEA treatment on glucose metabolism, hydrogen peroxide and thioredoxin levels in the skeletal muscle of diabetic rats

Dehydroepiandrosterone (DHEA) is an endogenous steroid hormone involved in a number of biological actions. This study shows the effects of DHEA on glucose metabolism, hydrogen peroxide and thioredoxin levels in the skeletal muscle of control and diabetic rats. Control and diabetic rats were chronically treated with DHEA (10 mg/kg) diluted in oil. Plasma concentration of DHEA and glucose, glucose uptake and oxidation, hydrogen peroxide, GLUT4, Akt and thioredoxin (Trx) was measured in the muscle. Results showed that there was a decrease in blood glucose in diabetic rats, probably linked to an increase in the glucose oxidation by the muscle or glucose uptake by some tissues. Despite the increase in the expression of GLUT4 in DHEA-treated rats, the glucose uptake was only higher in the control rats, showing that the glucose transporter may be present but not functional in the diabetic rats. The low expression of Trx due to diabetes became even lower with DHEA treatment. Although the reduction in blood glucose may be favorable, the decrease in Akt and Trx displays an environment conducive to redox imbalance. Thus, further studies are needed to ascertain the effects of DHEA treatment in diabetic rats.

Enhanced Leptin-Stimulated Pi3k Activation in the CNS Promotes White Adipose Tissue Transdifferentiation

The contribution of different leptin-induced signaling pathways in control of energy homeostasis is only partly understood. Here we show that selective Pten ablation in leptin-sensitive neurons (Pten(DeltaObRb)) results in enhanced Pi3k activation in these cells and reduces adiposity by increasing energy expenditure. White adipose tissue (WAT) of Pten(DeltaObRb) mice shows characteristics of brown adipose tissue (BAT), reflected by increased mitochondrial content and Ucp1 expression resulting from enhanced leptin-stimulated sympathetic nerve activity (SNA) in WAT. In contrast, leptin-deficient ob/ob-Pten(DeltaObRb) mice exhibit unaltered body weight and WAT morphology compared to ob/ob mice, pointing to a pivotal role of endogenous leptin in control of WAT transdifferentiation. Leanness of Pten(DeltaObRb) mice is accompanied by enhanced sensitivity to insulin in skeletal muscle. These data provide direct genetic evidence that leptin-stimulated Pi3k signaling in the CNS regulates energy expenditure via activation of SNA to perigonadal WAT leading to BAT-like differentiation of WAT.

Local and systemic alterations in cyclic 3′,5′ AMP phosphodiesterase activity in relation to tail regeneration under hypothyroidism and T4 replacement in the lizard,Mabuya carinata

To establish the relationship between thyroid hormone and cyclic Adenosine monophosphate (cAMP) during lacertilian tail regeneration, cAMP phosphodiesterase, the hydrolytic enzyme of cAMP, was assayed in the tail regenerate, liver, and skeletal muscle of control (group A), chemically thyroidectomized (group B), and thyroidectomized and T4-replaced (group C) animals during various periods of tail regeneration. Enzyme activity was elevated in all three tissues of group B animals. Animals of group C showed an intermediate level of enzyme activity between controls (group A) and experimental animals (group B). These observations suggest a possible regulatory role of thyroxine in maintaining optimum levels of phosphodiesterase. The retardation in regeneration observable in the hypothyroid group of animals may be correlated with low levels of tissue cAMP. However, the operation of other influencing factors on phosphodiesterase during regeneration can be surmised from the observed tendency to exhibit similar patterns of phase-specific modulations in enzyme activity. Our observations are discussed in terms of phase-specific involvement of cAMP in regeneration, as well as its role in other metabolic aspects and the possible mode of indirect control exerted by thyroxine on lacertilian tail regeneration.

Local and systemic alterations in cyclic 3′,5′ AMP phosphodiesterase activity in relation to tail regeneration under hypothyroidism and T4 replacement in the lizard, Mabuya carinata

To establish the relationship between thyroid hormone and cyclic Adenosine monophosphate (cAMP) during lacertilian tail regeneration, cAMP phosphodiesterase, the hydrolytic enzyme of cAMP, was assayed in the tail regenerate, liver, and skeletal muscle of control (group A), chemically thyroidectomized (group B), and thyroidectomized and T4-replaced (group C) animals during various periods of tail regeneration. Enzyme activity was elevated in all three tissues of group B animals. Animals of group C showed an intermediate level of enzyme activity between controls (group A) and experimental animals (group B). These observations suggest a possible regulatory role of thyroxine in maintaining optimum levels of phosphodiesterase. The retardation in regeneration observable in the hypothyroid group of animals may be correlated with low levels of tissue cAMP. However, the operation of other influencing factors on phosphodiesterase during regeneration can be surmised from the observed tendency to exhibit similar patterns of phase-specific modulations in enzyme activity. Our observations are discussed in terms of phase-specific involvement of cAMP in regeneration, as well as its role in other metabolic aspects and the possible mode of indirect control exerted by thyroxine on lacertilian tail regeneration. © 1996 Wiley-Liss, Inc.

A patch-clamp study of delayed rectifier currents in skeletal muscle of control and mdx mice.

1. Potassium currents were measured in the extensor digitorum longus muscle of normal and mdx mice, which lack the protein dystrophin, using the cell-attached and inside-out patch clamp techniques, in the presence of asymmetrical K+ concentrations (3 mM in the pipette, 160 mM in the bath). 2. In cell-attached patches, the delayed rectifier was the most commonly found potassium channel, with a density of roughly 8 channels microns-2. Outward macroscopic currents were activated in macropatches depolarized to potentials positive to -60 mV. The probability of opening reached half-maximal values around -40 mV for control patches and -31 mV for patches from mdx mice. 3. Tail currents were linear in the range between -60 and +20 mV, reversing close to -100 mV. The single channel current at 0 mV, estimated from non-stationary analysis of variance, was used in conjunction with the slope of the linear part of the tail current to calculate the single channel conductance, yielding a value of 19 +/- 1 pS. 4. At 0 mV, the delayed rectifier inactivated with two time constants, of 70 +/- 20 ms and 600 +/- 200 ms. Prepulses of 500 ms duration to different potentials produced incomplete inactivation with inactivation reaching 50% of its maximum at -50 mV. 5. Single channel activity was recorded using small pipettes. Both single channel conductance and kinetic behaviour were in agreement with the macroscopic current data. 6. In excised patches, the delayed rectifier current ran down, unmasking other K+ channels. A Ca(2+)-dependent K+ channel of 186 pS (BK-like channel) was found frequently in patches bathed in solutions containing appropriate concentrations of calcium, especially at stronger depolarizations. A K+ channel of 63 pS was unmasked in control excised patches bathed in solutions devoid of ATP. This channel was not observed in patches excised from mdx fibers.

Effect of hypothyroidism on the expression of cytochrome c and cytochrome c oxidase in heart and muscle during development.

The effect of thyroid hormone on the expression of mitochondrial proteins was evaluated during development by measuring cytochrome c oxidase (CYTOX) activity and cytochrome c protein and mRNA levels in heart and skeletal muscle of control and hypothyroid rats. Animals were killed at the late fetal, early, and late postnatal stages up to 56 days of age. In heart, CYTOX activity increased 2.3-fold above the fetal level throughout development, most of which occurred prior to 2 days of age. No increase was observed in muscle. CYTOX activity was reduced in hypothyroid animals throughout development in heart compared to controls (by 50% at 56 days), but in muscle no effect of hypothyroidism was observed. In muscle and heart 4- and 1.5-fold increases in cytochrome c above the fetal level were evident by 1 day of age, with further increases to 8.5- and 2.7-fold by 56 days, respectively. The increase in cytochrome c differed from the increase in CYTOX, indicating changes in mitochondrial composition. Hypothyroidism reduced cytochrome c in muscle by 30-35% at 56 days, but had no effect in heart, indicating a muscle type-specific effect of thyroid hormone on cytochrome c protein expression. Cytochrome c mRNA increased rapidly to 4-5 fold above the fetal level in both heart and muscle by 6 h post-partum. Between 7 and 56 days of age, further increases to 6- and 25-fold were observed in muscle and heart, respectively. In muscle, the 6-fold developmental increase in mRNA paralleled that of the protein, suggesting transcriptional regulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Spatially resolved changes in diabetic rat skeletal muscle metabolism in vivo studied by 31P-n.m.r. spectroscopy

Phase-modulated rotating-frame imaging (p.m.r.f.i.), a localization technique for 31P-n.m.r. spectroscopy, has been applied to obtain information on the heterogeneity of phosphorus-containing metabolites and pH in the skeletal muscle of control and streptozotocin-diabetic rats. Using this method, the metabolic changes in four spatially resolved longitudinal slices (where slice I is superficial and slice IV is deep muscle) through the ankle flexor muscles have been investigated at rest and during steady-state isometric twitch-contraction at 2 Hz. At rest, intracellular pH was lower, and phosphocreatine (PCr)/ATP was higher, throughout the muscle mass in diabetic compared with control animals. The change in PCr/ATP in diabetic muscle correlated with a decrease in the chemically determined ATP concentration. During the muscle stimulation period, the decrease in pH observed in diabetic muscle at rest was maintained, but not exacerbated, by the contractile stimulus. Stimulation of muscle contraction caused more marked changes in PCr/(PCr + Pi), PCr/ATP and Pi/ATP in the diabetic group. These changes were most evident in slice III, which contains the greatest proportion of fast glycolytic-oxidative (type IIa) fibres, in which statistically significant differences were observed for all metabolite ratios. The results presented suggest that some degree of heterogeneity occurs in diabetic skeletal muscle in vivo with respect to the extent of metabolic dysfunction caused by the diabetic insult and that regions of the muscle containing high proportions of type IIa fibres appear to be most severely affected.

Severe mitochondrial anomaly in dystrophic mouse skeletal muscle

Mitochondrial fractions were isolated from skeletal muscle of control (C57 BL 6J dy/+) and dystrophic (C57 BL 6J dy/dy) mice, and enzymatic activities (cytochrome c oxidase, rotenone-insensitive NADH cytochrome c reductase) were determined. After electrophoretic separation, calcium-binding proteins were identified. An important anomaly was observed in the mitochondria of dystrophic muscle, i.e., a considerable reduction of a specific calcium-binding protein (61,000 Da mol. wt.).

Effect of iron deficiency on energy conservation in rat liver and skeletal muscle submitochondrial particles

Submitochondrial particles prepared from liver and skeletal muscle of control and iron-deficient rats were examined for cytochrome content and for both energy-independent and energy-conserving functions. Liver submitochondrial particles appear quite resistant to iron deficiency with cytochrome content and electron-transferring or energy-conserving functions maintained at a level of 85% or better of normal. Iron-deficient skeletal muscle submitochondrial particles, in contrast, have decreased cytochrome content and only 15–20% of the normal capacity for oxidation through either complex I (NADH dehydrogenase) or complex II (succinate dehydrogenase). Energy-linked reactions which involve substrate oxidation/reduction (succinate → NAD + reversed electron flow and succinate-driven energy-dependent transhydrogenation) are likewise markedly decreased, while ATP-driven energy-dependent transhydrogenation and mitochondrial ATPase are normal. Our data support the concept that iron deficiency leads to decreased electroncarrying capacity of iron-containing mitochondrial enzymes, with skeletal muscle being much more susceptible than liver, but that the mitochondria are otherwise normal with regard to energy conservation.