Key points• Several biochemical measures of mitochondrial components are used as biomarkers of mitochondrial content and muscle oxidative capacity. However, no studies have validated these surrogates against a morphological measure of mitochondrial content in human subjects.• The most commonly used markers (citrate synthase activity, cardiolipin content, mitochondrial DNA content (mtDNA), complex I-V protein, and complex I-IV activity) were correlated with a measure of mitochondrial content (transmission electron microscopy) and muscle oxidative capacity (respiration in permeabilized fibres).• Cardiolipin content followed by citrate synthase activity and complex I activity were the biomarkers showing the strongest association with mitochondrial content.• mtDNA was found to be a poor biomarker of mitochondrial content.• Complex IV activity was closely associated with mitochondrial oxidative phosphorylation capacity.Abstract Skeletal muscle mitochondrial content varies extensively between human subjects. Biochemical measures of mitochondrial proteins, enzyme activities and lipids are often used as markers of mitochondrial content and muscle oxidative capacity (OXPHOS). The purpose of this study was to determine how closely associated these commonly used biochemical measures are to muscle mitochondrial content and OXPHOS. Sixteen young healthy male subjects were recruited for this study. Subjects completed a graded exercise test to determine maximal oxygen uptake (V O 2 peak ) and muscle biopsies were obtained from the vastus lateralis. Mitochondrial content was determined using transmission electron microscopy imaging and OXPHOS was determined as the maximal coupled respiration in permeabilized fibres. Biomarkers of interest were citrate synthase (CS) activity, cardiolipin content, mitochondrial DNA content (mtDNA), complex I-V protein content, and complex I-IV activity. Spearman correlation coefficient tests and Lin's concordance tests were applied to assess the absolute and relative association between the markers and mitochondrial content or OXPHOS. Subjects had a large range ofV O 2 peak (range 29.9-71.6 ml min −1 kg −1 ) and mitochondrial content (4-15% of cell volume). Cardiolipin content showed the strongest association with mitochondrial content followed by CS and complex I activities. mtDNA was not related to mitochondrial content. Complex IV activity showed the strongest association with muscle oxidative capacity followed by complex II activity. We conclude that cardiolipin content, and CS and complex I activities are the biomarkers that exhibit the strongest association with mitochondrial content, while complex IV activity is strongly associated with OXPHOS capacity in human skeletal muscle.

The crystal structure of a PNA duplex reveals both a right- and a left-handed helix in the unit cell. The helices are wide (28A), large pitched (18bp) with the base pairs perpendicular to the helix axis, thereby demonstrating that PNA besides adapting to oligonucleotide partners also has a unique structure by itself.

Non-technical summary Glucose is stored as glycogen in skeletal muscle. The importance of glycogen as a fuel during exercise has been recognized since the 1960s; however, little is known about the precise mechanism that relates skeletal muscle glycogen to muscle fatigue. We show that low muscle glycogen is associated with an impairment of muscle ability to release Ca 2+ , which is an important signal in the muscle activation. Thus, depletion of glycogen during prolonged, exhausting exercise may contribute to muscle fatigue by causing decreased Ca 2+ release inside the muscle. These data provide indications of a signal that links energy utilization, i.e. muscle contraction, with the energy content in the muscle, thereby inhibiting a detrimental depletion of the muscle energy store.Abstract Little is known about the precise mechanism that relates skeletal muscle glycogen to muscle fatigue. The aim of the present study was to examine the effect of glycogen on sarcoplasmic reticulum (SR) function in the arm and leg muscles of elite cross-country skiers (n = 10,V O 2 max 72 ± 2 ml kg −1 min −1 ) before, immediately after, and 4 h and 22 h after a fatiguing 1 h ski race. During the first 4 h recovery, skiers received either water or carbohydrate (CHO) and thereafter all received CHO-enriched food. Immediately after the race, arm glycogen was reduced to 31 ± 4% and SR Ca 2+ release rate decreased to 85 ± 2% of initial levels. Glycogen noticeably recovered after 4 h recovery with CHO (59 ± 5% initial) and the SR Ca 2+ release rate returned to pre-exercise levels. However, in the absence of CHO during the first 4 h recovery, glycogen and the SR Ca 2+ release rate remained unchanged (29 ± 2% and 77 ± 8%, respectively), with both parameters becoming normal after the remaining 18 h recovery with CHO. Leg muscle glycogen decreased to a lesser extent (71 ± 10% initial), with no effects on the SR Ca 2+ release rate. Interestingly, transmission electron microscopy (TEM) analysis revealed that the specific pool of intramyofibrillar glycogen, representing 10-15% of total glycogen, was highly significantly correlated with the SR Ca 2+ release rate. These observations strongly indicate that low glycogen and especially intramyofibrillar glycogen, as suggested by TEM, modulate the SR Ca 2+ release rate in highly trained subjects. Thus, low glycogen during exercise may contribute to fatigue by causing a decreased SR Ca 2+ release rate.

Ørtenblad N. Increased subsarcolemmal lipids in type 2 diabetes: effect of training on localization of lipids, mitochondria, and glycogen in sedentary human skeletal muscle. Am J Physiol Endocrinol Metab 298: E706 -E713, 2010. First published December 22, 2009; doi:10.1152/ajpendo.00692.2009.-The purpose of the study was to investigate the effect of aerobic training and type 2 diabetes on intramyocellular localization of lipids, mitochondria, and glycogen. Obese type 2 diabetic patients (n ϭ 12) and matched obese controls (n ϭ 12) participated in aerobic cycling training for 10 wk. Endurance-trained athletes (n ϭ 15) were included for comparison. Insulin action was determined by euglycemic-hyperinsulinemic clamp. Intramyocellular contents of lipids, mitochondria, and glycogen at different subcellular compartments were assessed by transmission electron microscopy in biopsies obtained from vastus lateralis muscle. Type 2 diabetic patients were more insulin resistant than obese controls and had threefold higher volume of subsarcolemmal (SS) lipids compared with obese controls and endurance-trained subjects. No difference was found in intermyofibrillar lipids. Importantly, following aerobic training, this excess SS lipid volume was lowered by ϳ50%, approaching the levels observed in the nondiabetic subjects. A strong inverse association between insulin sensitivity and SS lipid volume was found (r 2 ϭ0.62, P ϭ 0.002). The volume density and localization of mitochondria and glycogen were the same in type 2 diabetic patients and control subjects, and showed in parallel with improved insulin sensitivity a similar increase in response to training, however, with a more pronounced increase in SS mitochondria and SS glycogen than in other localizations. In conclusion, this study, estimating intramyocellular localization of lipids, mitochondria, and glycogen, indicates that type 2 diabetic patients may be exposed to increased levels of SS lipids. Thus consideration of cell compartmentation may advance the understanding of the role of lipids in muscle function and type 2 diabetes. cell compartmentation; transmission electron microscopy; insulin sensitivity INTRAMYOCELLULAR LIPID (IMCL) accumulation in skeletal muscle of humans has been related to impaired insulin sensitivity (20,30). The causality has been challenged by reports of increased IMCL levels in endurance-trained athletes compared with untrained (8) and higher IMCL levels in women than in men without concomitant differences in insulin sensitivity (11,17). Thus many have suggested that high IMCL levels per se do not influence insulin sensitivity but represent a marker of increased fatty acid metabolites such as diacylglycerol (DAG), ceramide, and long-chain acyl-CoAs, which in turn could be detrimental for insulin sensitivity (17,28,31).However, evaluation of the role of IMCL in subcellular fractions has not been considered in previous studies (8,11,20,30). The muscle cell consists mainly of contractile filaments arranged in myofibrils with mitochondria, lipids, gly...

Studies performed at the beginning of the last century revealed the importance of carbohydrate as a fuel during exercise, and the importance of muscle glycogen on performance has subsequently been confirmed in numerous studies. However, the link between glycogen depletion and impaired muscle function during fatigue is not well understood and a direct cause-and-effect relationship between glycogen and muscle function remains to be established. The use of electron microscopy has revealed that glycogen is not homogeneously distributed in skeletal muscle fibres, but rather localized in distinct pools. Furthermore, each glycogen granule has its own metabolic machinery with glycolytic enzymes and regulating proteins. One pool of such glycogenolytic complexes is localized within the myofibrils in close contact with key proteins involved in the excitation-contraction coupling and Ca 2+ release from the sarcoplasmic reticulum (SR). We and others have provided experimental evidence in favour of a direct role of decreased glycogen, localized within the myofibrils, for the reduction in SR Ca 2+ release during fatigue. This is consistent with compartmentalized energy turnover and distinctly localized glycogen pools being of key importance for SR Ca 2+ release and thereby affecting muscle contractility and fatigability.

Non-technical summary During prolonged high-intensity exercise the main fuel for muscular work is glycogen, the storage form of glucose in skeletal muscle. The role of muscle glycogen in muscle function is best demonstrated by the inability to sustain prolonged high-intensity exercise when the glycogen stores are depleted. Despite this knowledge, the reason why muscle function is depressed when glycogen levels are low is still not known. We show that after prolonged exhaustive exercise the depletion of glycogen stores is dependent on its localization within the muscle cells. These results show that consideration of distinct localizations within the muscle cells may advance understanding of how and why low muscle glycogen content impairs muscle function.Abstract Although glycogen is known to be heterogeneously distributed within skeletal muscle cells, there is presently little information available about the role of fibre types, utilization and resynthesis during and after exercise with respect to glycogen localization. Here, we tested the hypothesis that utilization of glycogen with different subcellular localizations during exhaustive arm and leg exercise differs and examined the influence of fibre type and carbohydrate availability on its subsequent resynthesis. When 10 elite endurance athletes (22 ± 1 years, V O 2 max = 68 ± 5 ml kg −1 min −1 , mean ± SD) performed one hour of exhaustive arm and leg exercise, transmission electron microscopy revealed more pronounced depletion of intramyofibrillar than of intermyofibrillar and subsarcolemmal glycogen. This phenomenon was the same for type I and II fibres, although at rest prior to exercise, the former contained more intramyofibrillar and subsarcolemmal glycogen than the latter. In highly glycogen-depleted fibres, the remaining small intermyofibrillar and subsarcolemmal glycogen particles were often found to cluster in groupings. In the recovery period, when the athletes received either a carbohydrate-rich meal or only water the impaired resynthesis of glycogen with water alone was associated primarily with intramyofibrillar glycogen. In conclusion, after prolonged high-intensity exercise the depletion of glycogen is dependent on subcellular localization. In addition, the localization of glycogen appears to be influenced by fibre type prior to exercise, as well as carbohydrate availability during the subsequent period of recovery. These findings provide insight into the significance of fibre type-specific compartmentalization of glycogen metabolism in skeletal muscle during exercise and subsequent recovery.

The analysis of gene polymorphism allows delineation of a group of patients (30%) with a response rate to a single drug of approximately 50%. This information should be used in the design of tailored treatment.

In vitro experiments indicate a non-metabolic role of muscle glycogen in contracting skeletal muscles. Since the sequence of events in excitation-contraction (E-C) coupling is known to be located close to glycogen granules, at specific sites on the fibre, we hypothesized that the distinct compartments of glycogen have specific effects on muscle fibre contractility and fatigability. Single skeletal muscle fibres (n = 19) from fed and fasted rats were mechanically skinned and divided into two segments. In one segment glycogen localization and volume fraction were estimated by transmission electron microscopy. The other segment was mechanically skinned and, in the presence of high and constant myoplasmic ATP and PCr, electrically stimulated (10 Hz, 0.8 s every 3 s) eliciting repeated tetanic contractions until the force response was decreased by 50% (mean ± s.e.m., 81 ± 16, range 22-252 contractions). Initially the total myofibrillar glycogen volume percentage was 0.46 ± 0.07%, with 72 ± 3% in the intermyofibrillar space and 28 ± 3% in the intramyofibrillar space. The intramyofibrillar glycogen content was positively correlated with the fatigue resistance capacity (r 2 = 0.32, P = 0.02). Intermyofibrillar glycogen was inversely correlated with the half-relaxation time in the unfatigued tetanus (r 2 = 0.25, P = 0.03). These results demonstrate for the first time that two distinct subcellular populations of glycogen have different roles in contracting single muscle fibres under conditions of high myoplasmic ATP. The consistent observations that muscle glycogen stores at the beginning of exercise are closely related to endurance capacity (Bergström et al. 1967) and that the time point of exhaustion after prolonged exercise coincides with low muscle glycogen levels , clearly suggest a role for muscle glycogen in fatigue. However, the link between glycogen and impaired muscle function during fatigue is not well understood and a direct cause-and-effect relationship between glycogen and muscle function remains to be established. In in vitro intermittent stimulation protocols of whole muscles and single fibres fatigue is related to excitation-contraction (E-C) coupling failure (Westerblad & Allen, 1991) and accumulating evidence indicates that glycogen depletion impairs E-C coupling leading to fatigue (Chin & Allen, 1997;Kabbara et al. 2000;Helander et al. 2002). This is evident from experiments where isolated small bundles of mouse fast-twitch muscle fibres were fatigued by repeated tetani, which markedly reduced glycogen content and tetanic intracellular free [Ca 2+ ] ([Ca 2+ ] i ). Following 1 h recovery maximum force (F max ) and [Ca 2+ ] i were normalized. However, when fibres recovered in the absence of glucose the low glycogen levels were maintained and there was a sustained reduction in [Ca 2+ ] i , depressed reduction in F max and muscles fatigued faster in a second fatigue run (Chin & Allen, 1997). These observations were later confirmed in cane toad fibres and mouse extensor digitorum longus (EDL) fibres (...