Intramuscular triacylglycerol utilization in human skeletal muscle during exercise: is there a controversy?

Intramuscular triacylglyerols (IMTGs) represent a potentially important energy source for contracting human skeletal muscle. Although the majority of evidence from isotope tracer and (1)H-magnetic resonance spectroscopy (MRS) studies demonstrate IMTG utilization during exercise, controversy regarding the importance of IMTG as a metabolic substrate persists. The controversy stems from studies that measure IMTG in skeletal muscle biopsy samples and report no significant net IMTG degradation during prolonged moderate-intensity (55-70% maximal O(2) consumption) exercise lasting 90-120 min. Although postexercise decrements in IMTG levels are often reported from direct muscle measurements, the marked between-biopsy variability (approximately 23%) that has been reported with this technique in untrained subjects is larger than the expected decrease in IMTG content, effectively precluding significant findings. In contrast, recent data obtained in endurance-trained subjects demonstrated reduced variability between duplicate biopsies (approximately 12%), and significant changes in IMTG were detected after 120 min of moderate-intensity exercise. Therefore, it is our contention that the muscle biopsy, isotope tracer, and (1)H-MRS techniques report significant and energetically important oxidation of free fatty acids derived from IMTGs during prolonged moderate exercise.     

Is a guanine nucleotide-binding protein involved in excitation-contraction coupling in skeletal muscle?

Plasma membrane depolarization causes skeletal muscle contraction by triggering Ca2+ release from an intracellular membrane network, the sarcoplasmic reticulum. A specialized portion of the sarcoplasmic reticulum, the terminal cisternae, is junctionally associated with sarcolemmal invaginations called the transverse tubules, but the mechanism by which the action potential at the level of the transverse tubules is coupled to Ca2+ release from the terminal cisternae is still mysterious. Here we show that: (i) GTP gamma S, a non-hydrolyzable analog of GTP, elicits isometric force development in skinned muscle fibre; (ii) GTP gamma S is unable to release CA2+ from isolated sarcoplasmic reticulum fractions; (iii) the threshold for tension development is shifted to higher GTP gamma S concentrations by pre-incubation with pertussis toxin. These results suggest that a GTP-binding protein is involved in coupling the action potential of transverse tubules to Ca2+ release from the terminal cisternae.     

Does impaired mitochondrial function affect insulin signaling and action in cultured human skeletal muscle cells?

Insulin-resistant type 2 diabetic patients have been reported to have impaired skeletal muscle mitochondrial respiratory function. A key question is whether decreased mitochondrial respiration contributes directly to the decreased insulin action. To address this, a model of impaired cellular respiratory function was established by incubating human skeletal muscle cell cultures with the mitochondrial inhibitor sodium azide and examining the effects on insulin action. Incubation of human skeletal muscle cells with 50 and 75 microM azide resulted in 48 +/- 3% and 56 +/- 1% decreases, respectively, in respiration compared with untreated cells mimicking the level of impairment seen in type 2 diabetes. Under conditions of decreased respiratory chain function, insulin-independent (basal) glucose uptake was significantly increased. Basal glucose uptake was 325 +/- 39 pmol/min/mg (mean +/- SE) in untreated cells. This increased to 669 +/- 69 and 823 +/- 83 pmol/min/mg in cells treated with 50 and 75 microM azide, respectively (vs. untreated, both P < 0.0001). Azide treatment was also accompanied by an increase in basal glycogen synthesis and phosphorylation of AMP-activated protein kinase. However, there was no decrease in glucose uptake following insulin exposure, and insulin-stimulated phosphorylation of Akt was normal under these conditions. GLUT1 mRNA expression remained unchanged, whereas GLUT4 mRNA expression increased following azide treatment. In conclusion, under conditions of impaired mitochondrial respiration there was no evidence of impaired insulin signaling or glucose uptake following insulin exposure in this model system.     

Does exercise-induced muscle damage play a role in skeletal muscle hypertrophy?

Exercise-induced muscle damage (EIMD) occurs primarily from the performance of unaccustomed exercise, and its severity is modulated by the type, intensity, and duration of training. Although concentric and isometric actions contribute to EIMD, the greatest damage to muscle tissue is seen with eccentric exercise, where muscles are forcibly lengthened. Damage can be specific to just a few macromolecules of tissue or result in large tears in the sarcolemma, basal lamina, and supportive connective tissue, and inducing injury to contractile elements and the cytoskeleton. Although EIMD can have detrimental short-term effects on markers of performance and pain, it has been hypothesized that the associated skeletal muscle inflammation and increased protein turnover are necessary for long-term hypertrophic adaptations. A theoretical basis for this belief has been proposed, whereby the structural changes associated with EIMD influence gene expression, resulting in a strengthening of the tissue and thus protection of the muscle against further injury. Other researchers, however, have questioned this hypothesis, noting that hypertrophy can occur in the relative absence of muscle damage. Therefore, the purpose of this article will be twofold: (a) to extensively review the literature and attempt to determine what, if any, role EIMD plays in promoting skeletal muscle hypertrophy and (b) to make applicable recommendations for resistance training program design.     

Homocysteine induces energy imbalance in rat skeletal muscle: Is creatine a protector?

Homocystinuria is a neurometabolic disease caused by a severe deficiency of cystathionine beta‐synthase activity, resulting in severe hyperhomocysteinemia. Affected patients present several symptoms including a variable degree of motor dysfunction. In this study, we investigated the effect of chronic hyperhomocysteinemia on the cell viability of the mitochondrion, as well as on some parameters of energy metabolism, such as glucose oxidation and activities of pyruvate kinase, citrate synthase, isocitrate dehydrogenase, malate dehydrogenase, respiratory chain complexes and creatine kinase in gastrocnemius rat skeletal muscle. We also evaluated the effect of creatine on biochemical alterations elicited by hyperhomocysteinemia. Wistar rats received daily subcutaneous injections of homocysteine (0.3–0.6 µmol/g body weight) and/or creatine (50 mg/kg body weight) from the 6th to the 28th days of age. The animals were decapitated 12 h after the last injection. Homocysteine decreased the cell viability of the mitochondrion and the activities of pyruvate kinase and creatine kinase. Succinate dehydrogenase was increased other evaluated parameters were not changed by this amino acid. Creatine, when combined with homocysteine, prevented or caused a synergistic effect on some changes provoked by this amino acid. Creatine per se or creatine plus homocysteine altered glucose oxidation. These findings provide insights into the mechanisms by which homocysteine exerts its effects on skeletal muscle function, more studies are needed to elucidate them. Although creatine prevents some alterations caused by homocysteine, it should be used with caution, mainly in healthy individuals because it could change the homeostasis of normal physiological functions. Copyright © 2012 John Wiley & Sons, Ltd.     

Is rat skeletal muscle hexokinase an allosteric enzyme?

A recent report by M. Gregoriou, I. P. Trayer, and A. Cornish-Bowden (1986, Eur. J. Biochem. 161, 171-176) showed that the mechanism for rat skeletal muscle hexokinase contains two allosteric sites: one for ATP and one for glucose 6-phosphate. In this report, we show that the allosteric mechanism is at variance with a large amount of kinetic data for the skeletal muscle hexokinase reaction in the literature. In addition, the allosteric mechanism conflicts with isotope exchange at chemical equilibrium data reported by M. Gregoriou, I. P. Trayer, and A. Cornish-Bowden (1983, Eur. J. Biochem. 134, 283-288).     

Is regulation of proteolysis associated with redox-state changes in rat skeletal muscle?

In isolated rat diaphragms, only those substrates that increased the tissue NADH/NAD+ ratio lowered the rate of proteolysis. However, direct inhibition of proteinase activity by leupeptin promoted oxidation of the NAD couple of the muscles. These results suggest that changes in muscle reduction-oxidation state may be important in the regulation of proteolysis.     

Do P2X purinergic receptors regulate skeletal muscle blood flow during exercise?

Although there is evidence that sympathetic nerves release ATP as a neurotransmitter to produce vasoconstriction via P2X purinergic receptors, the role of these receptors in the regulation of blood flow to exercising skeletal muscle has yet to be determined. We hypothesized that there is tonic P2X receptor-mediated vasoconstriction in exercising skeletal muscle. To test this hypothesis, the effect of P2X receptor blockade on skeletal muscle blood flow was examined in six exercising mongrel dogs. P2X receptor antagonism was accomplished with pyridoxal-phosphate-6-azophenyl-2'4'-disulfonic acid (PPADs). Animals were instrumented chronically with flow probes on the external iliac arteries of both hindlimbs and a catheter in one femoral artery. PPADs (40 mg) was infused as a bolus into the femoral artery catheter during steady-state exercise at 6 miles/h. Intra-arterial infusion of PPADs increased iliac blood flow from 542 +/- 55 to 677 +/- 69 ml/min (P < 0.05) and iliac vascular conductance from 5.17 +/- 0.62 to 6.53 +/- 0.80 ml.min(-1).mmHg(-1). The PPADs infusion did not affect blood flow in the contralateral iliac artery. These data support the hypothesis that P2X purinergic receptors produce vasoconstriction in exercising skeletal muscle.     

Identification and characterization of PDGFRα+ mesenchymal progenitors in human skeletal muscle

Fatty and fibrous connective tissue formation is a hallmark of diseased skeletal muscle and deteriorates muscle function. We previously identified non-myogenic mesenchymal progenitors that contribute to adipogenesis and fibrogenesis in mouse skeletal muscle. In this study, we report the identification and characterization of a human counterpart to these progenitors. By using PDGFRα as a specific marker, mesenchymal progenitors can be identified in the interstitium and isolated from human skeletal muscle. PDGFRα⁺ cells represent a cell population distinct from CD56⁺ myogenic cells, and adipogenic and fibrogenic potentials were highly enriched in the PDGFRα⁺ population. Activation of PDGFRα stimulates proliferation of PDGFRα⁺ cells through PI3K-Akt and MEK2-MAPK signaling pathways, and aberrant accumulation of PDGFRα⁺ cells was conspicuous in muscles of patients with both genetic and non-genetic muscle diseases. Our results revealed the pathological relevance of PDGFRα⁺ mesenchymal progenitors to human muscle diseases and provide a basis for developing therapeutic strategy to treat muscle diseases.     

Calcium signaling in isolated skeletal muscle fibers investigated under “silicone voltage-clamp” conditions

In skeletal muscle, release of calcium from the sarcoplasmic reticulum (SR) represents the major source of cytoplasmic Ca²⁺ elevation. SR calcium release is under the strict command of the membrane potential, which drives the interaction between the voltage sensors in the t-tubule membrane and the calcium-release channels. Either detection or control of the membrane voltage is thus essential when studying intracellular calcium signaling in an intact muscle fiber preparation. The silicone-clamp technique used in combination with intracellular calcium measurements represents an efficient tool for such studies. This article reviews some properties of the plasma membrane and intracellular signals measured with this methodology in mouse skeletal muscle fibers. Focus is given to the potency of this approach to investigate both fundamental aspects of excitation-contraction coupling and potential alterations of intracellular calcium handling in some muscle diseases.     

Deubiquitinating enzyme A20 negatively regulates NF-κB signaling in skeletal muscle in mdx mice

Duchenne muscular dystrophy (DMD) is caused by the lack of a functional dystrophin protein that results in muscle fiber membrane disruption and, ultimately, degeneration. Regeneration of muscle fibers fails progressively, and muscle tissue is replaced with connective tissue. As a result, DMD causes progressive limb muscle weakness and cardiac and respiratory failure. The absence of dystrophin from muscle fibers triggers the chronic activation of the nuclear factor of kappa B (NF-κB). Chronic activation of NF-κB in muscle leads to infiltration of macrophages, up-regulation of the ubiquitin-proteosome system, and down-regulation of the helix-loop-helix muscle regulatory factor, MyoD. These processes, triggered by NF-κB activation, promote muscle degeneration and failure of muscle regeneration. A20 (TNFAIP3) is a critical negative regulator of NF-κB. In this study, we characterize the role of A20 in regulating NF-κB activation in skeletal muscle, identifying a novel role in muscle regeneration. A20 is highly expressed in regenerating muscle fibers, and knockdown of A20 impairs muscle differentiation in vitro, which suggests that A20 expression is critically important for regeneration of dystrophic muscle tissue. Furthermore, down-regulation of the classic pathway of NF-κB activation is associated with up-regulation of the alternate pathway in regenerating muscle fibers, suggesting a mechanism by which A20 promotes muscle regeneration. These results demonstrate the important role of A20 in muscle fiber repair and suggest the potential of A20 as a therapeutic target to ameliorate the pathology and clinical symptoms of DMD.