Piperine enhances contractile force in slow- and fast-twitch muscle.

Piperine has been shown to bind to myosin and shift the distribution of conformational states of myosin molecules from the super-relaxed state to the disordered relaxed state. However, little is known about the implications for muscle force production and potential underlying mechanisms. Muscle contractility experiments were performed using isolated muscles and single fibres from rats and mice. The dose-response effect of piperine on muscle force was assessed at several stimulation frequencies. The potentiation of muscle force was also tested in muscles fatigued by eccentric contractions. Potential mechanisms of force potentiation were assessed by measuring Ca2+ levels during stimulation in enzymatically dissociated muscle fibres, while myofibrillar Ca2+ sensitivity was assessed in chemically skinned muscle fibres. Piperine caused a dose-dependent increase in low-frequency force with no effect on high-frequency force in both slow- and fast-twitch muscle, with similar relative increases in twitch force, rate of force development and relaxation rate. The potentiating effect of piperine on low-frequency force was reversible, and piperine partially recovered low-frequency force in fatigued muscle. Piperine had no effect on myoplasmic free [Ca2+] levels in mouse muscle fibres, whereas piperine substantially augmented the force response to submaximal levels of [Ca2+] in rat MyHCII fibres and MyHCI fibres along with a minor increase in maximum Ca2+-activated force. Piperine enhances low-frequency force production in both fast- and slow-twitch muscle. The effects are reversible and can counteract muscle fatigue. The primary underlying mechanism appears to be an increase in Ca2+ sensitivity. KEY POINTS: Piperine is a plant alkaloid derived from black pepper. It is known to bind to skeletal muscle myosin and enhance resting ATP turnover but its effects on contractility are not well known. We showed for the first time a piperine-induced force potentiation that was pronounced during low-frequency electrical stimulation of isolated muscles. The effect of piperine was observed in both slow and fast muscle types, was reversible, and could counteract the force decrements observed after fatiguing muscle contractions. Piperine treatment caused an increase in myofibrillar Ca2+ sensitivity in chemically skinned muscle fibres, while we observed no effect on intracellular Ca2+ concentrations during electrical stimulation in enzymatically dissociated muscle fibres.

Role of recovery of acetylcholine release in compromised neuromuscular junction function.

Everyday physical activities, such as walking, are enabled by repeated skeletal muscle contractions and require a well-functioning neuromuscular transmission. In myasthenic disorders, activities of daily living are debilitated by a compromised neuromuscular transmission leading to muscle weakness and fatiguability in patients. To enable physical activity, acetylcholine (ACh) is released repeatedly from the motor nerve, however, the role of the nerve terminals' capacity to sustain ACh release to support repetitive contractions under compromised neuromuscular transmission remains unclear. To explore this, we studied synaptic and contractile function during repeated contractions in healthy rat skeletal muscles under conditions of pharmacological induced compromised neuromuscular transmission. Using recordings of endplate potentials, compound muscle action potential (CMAP) and force production in isolated skeletal muscles and living, anesthetized animals, we found that force and CMAP were markedly reduced by even very light activity performed up to 5 s prior to contraction showing that recovery of ACh release was insufficient to maintain synaptic transmission strength. Our results suggest that the timing of depletion and restoration of ACh release may impact clinical signs of weakness and fatigability in patients with impaired neuromuscular transmission and affect the sensitivity of electromyographic recordings in the clinic.

The Effect of Alginate Encapsulated Plant-Based Carbohydrate and Protein Supplementation on Recovery and Subsequent Performance in Athletes.

The main purpose of this study was to investigate the effect of a novel alginate-encapsulated carbohydrate-protein (CHO-PRO ratio 2:1) supplement (ALG) on cycling performance. The ALG, designed to control the release of nutrients, was compared to an isocaloric carbohydrate-only control (CON). Alginate encapsulation of CHOs has the potential to reduce the risk of carious lesions. METHODS: In a randomised cross-over clinical trial, 14 men completed a preliminary test over 2 experimental days separated by ~6 days. An experimental day consisted of an exercise bout (EX1) of cycling until exhaustion at W~73%, followed by 5 h of recovery and a subsequent time-to-exhaustion (TTE) performance test at W~65%. Subjects ingested either ALG (0.8 g CHO/kg/hr + 0.4 g PRO/kg/hr) or CON (1.2 g CHO/kg/hr) during the first 2 h of recovery. RESULTS: Participants cycled on average 75.2 ± 5.9 min during EX1. Levels of plasma branched-chain amino acids decreased significantly after EX1, and increased significantly with the intake of ALG during the recovery period. During recovery, a significantly higher plasma insulin and glucose response was observed after intake of CON compared to ALG. Intake of ALG increased plasma glucagon, free fatty acids, and glycerol significantly. No differences were found in the TTE between the supplements (p = 0.13) nor in the pH of the subjects' saliva. CONCLUSIONS: During the ALG supplement, plasma amino acids remained elevated during the recovery. Despite the 1/3 less CHO intake with ALG compared to CON, the TTE performance was similar after intake of either supplement.

A century of exercise physiology: effects of muscle contraction and exercise on skeletal muscle Na+,K+-ATPase, Na+ and K+ ions, and on plasma K+ concentration-historical developments.

This historical review traces key discoveries regarding K+ and Na+ ions in skeletal muscle at rest and with exercise, including contents and concentrations, Na+,K+-ATPase (NKA) and exercise effects on plasma [K+] in humans. Following initial measures in 1896 of muscle contents in various species, including humans, electrical stimulation of animal muscle showed K+ loss and gains in Na+, Cl- and H20, then subsequently bidirectional muscle K+ and Na+ fluxes. After NKA discovery in 1957, methods were developed to quantify muscle NKA activity via rates of ATP hydrolysis, Na+/K+ radioisotope fluxes, [3H]-ouabain binding and phosphatase activity. Since then, it became clear that NKA plays a central role in Na+/K+ homeostasis and that NKA content and activity are regulated by muscle contractions and numerous hormones. During intense exercise in humans, muscle intracellular [K+] falls by 21 mM (range - 13 to - 39 mM), interstitial [K+] increases to 12-13 mM, and plasma [K+] rises to 6-8 mM, whilst post-exercise plasma [K+] falls rapidly, reflecting increased muscle NKA activity. Contractions were shown to increase NKA activity in proportion to activation frequency in animal intact muscle preparations. In human muscle, [3H]-ouabain-binding content fully quantifies NKA content, whilst the method mainly detects α2 isoforms in rats. Acute or chronic exercise affects human muscle K+, NKA content, activity, isoforms and phospholemman (FXYD1). Numerous hormones, pharmacological and dietary interventions, altered acid-base or redox states, exercise training and physical inactivity modulate plasma [K+] during exercise. Finally, historical research approaches largely excluded female participants and typically used very small sample sizes.

Potentiation of force by extracellular potassium is not dependent on muscle length in mouse EDL muscle.

Increases in myofiber extracellular potassium with prolonged contractile activity can potentiate twitch force. Activity-dependent potentiation, another mechanism of force increase in skeletal muscle, has a strong dependence on muscle or sarcomere length. Thus, potassium-mediated twitch potentiation could also be length-dependent. However, this has not been previously investigated. To this end, we used isolated C57BL/6 mouse extensor digitorum longus (EDL) muscles and elicited twitches at 0.9 Lo, Lo, and 1.1 Lo (Lo refers to optimal length) in normal (5 mM) and high (10 mM) potassium solutions. Potentiation magnitude was similar to previous observations and was not significantly different between lengths (0.9 Lo: 12.3 ± 4.4%, Lo: 12.2 ± 3.6%, 1.1 Lo: 11.8 ± 4.8%, values are means ± SD). Exposure to dantrolene sodium, a compound that attenuates calcium release, reduced twitch force across lengths by ∼70%. When dantrolene-affected muscles were subsequently exposed to high potassium, potentiation was similar to that observed in the absence of the former. In total, these findings provide novel information on potassium-mediated twitch potentiation.NEW & NOTEWORTHY Here, we investigated the length-dependence of twitch force potentiation by extracellular potassium in mouse extensor digitorum longus (EDL) in vitro, at 25°C. Potentiation magnitude did not display a statistically significant difference between the examined muscle lengths. These results describe, for the first time, the relationship of this form of potentiation with muscle length, thus furthering the understanding of how it is integrated in in vivo muscle function.

The effect of low-frequency fatigue on the torque-velocity relationship in human quadriceps.

Low-frequency fatigue (LFF) is usually defined as the decline in low:high-frequency force of electrically evoked isometric muscle contractions. The influence of LFF on dynamic muscle function is not well studied. Our aim was to assess the effect of LFF on the electrically evoked torque-velocity relationship in humans. Sixteen participants underwent a series of electrically evoked knee extensions in an isokinetic dynamometer to establish torque-velocity relationships at 15 and 50 Hz using isokinetic contractions. Hereafter, fatigue was induced by five sets of 10 repetitions of maximal voluntary dynamic knee extensions. After 30 min of rest, torque-velocity tests were repeated. Maximal torque (Fmax) was measured, whereas maximal contraction velocity (Vmax) and maximal power (Pmax) were estimated using Hill's force-velocity equation, 15:50 Hz ratios were calculated for Fmax, Vmax, and Pmax as markers of LFF. Fmax decreased by 40% at 15 Hz (P = 0.001) and by 15% at 50 Hz (P = 0.001) in the fatigued state. No significant change was detected for Vmax at 15 Hz [-2%, (P = 0.349)] or 50 Hz [+3% (P = 0.763)], whereas 15 and 50 Hz Pmax decreased by 30% (P = 0.004) and 10% (P = 0.008), respectively. Following the fatigue protocol, the 15:50 Hz Fmax ratio decreased by 31% (P < 0.001), indicating LFF. The 15:50 Hz Pmax ratio also decreased by 23% (P = 0.002), whereas the 15:50 Hz Vmax ratio was unchanged (P = 0.313). In conclusion, fatiguing contractions decreased Fmax and Pmax at both high and low stimulation frequencies, whereas Vmax appeared unaffected. Nevertheless, LFF influences power production during human dynamic contractions at a range of submaximal velocities.NEW & NOTEWORTHY Force-velocity relationships were established using either low- or high-frequency electrical stimulation before and after fatiguing voluntary eccentric/concentric contractions of the knee extensors. Low-frequency fatigue was assessed by the relative decrease in low- and high-frequency maximal torque, maximal shortening velocity, and maximal power estimated by the force-velocity relationship. Low-frequency fatigue manifests itself as a large decrease in low-frequency maximal force and power with a modest decrease in high-frequency maximal force and power. Contraction velocity does not seem to decrease in the same manner.

No net utilization of intramuscular lipid droplets during repeated high-intensity intermittent exercise.

Intramuscular lipids are stored as subsarcolemmal or intramyofibrillar droplets with potential diverse roles in energy metabolism. We examined intramuscular lipid utilization through transmission electron microscopy during repeated high-intensity intermittent exercise, an aspect that is hitherto unexplored. Seventeen moderately to well-trained males underwent three periods (EX1-EX3) of 10 × 45-s high-intensity cycling [∼100%-120% Wattmax (Wmax)] combined with maximal repeated sprints (∼250%-300% Wmax). M. vastus lateralis biopsies were obtained at baseline, after EX1, and EX3. During the complete exercise session, no net decline in either subsarcolemmal or intermyofibrillar lipid volume density occurred. However, a temporal relationship emerged for subsarcolemmal lipids with an ∼11% increase in droplet size after EX1 (P = 0.024), which reverted to baseline levels after EX3 accompanied by an ∼30% reduction in the numerical density of subsarcolemmal lipid droplets compared with both baseline (P = 0.019) and after EX1 (P = 0.018). Baseline distinctions were demonstrated with an approximately twofold higher intermyofibrillar lipid volume in type 1 versus type 2 fibers (P = 0.008), mediated solely by a higher number rather than the size of lipid droplets (P < 0.001). No fiber-type-specific differences were observed in subsarcolemmal lipid volume although type 2 fibers exhibited ∼17% larger droplets (P = 0.034) but a lower numerical density (main effect; P = 0.010) including 3% less droplets at baseline. Collectively, these findings suggest that intramuscular lipids do not serve as an important substrate during high-intensity intermittent exercise; however, the repeated exercise pattern mediated a temporal remodeling of the subsarcolemmal lipid pool. Furthermore, fiber-type- and compartment-specific differences were found at baseline underscoring the heterogeneity in lipid droplet deposition.NEW & NOTEWORTHY Undertaking a severe repeated high-intensity intermittent exercise protocol led to no net decline in neither subsarcolemmal nor intermyofibrillar lipid content in the thigh muscle of young moderately to well-trained participants. However, a temporal remodeling of the subsarcolemmal pool of lipid droplets did occur indicative of potential transient lipid accumulation. Moreover, baseline fiber-type distinctions in subcellular lipid droplet deposition were present underscoring the diversity in lipid droplet storage among fiber types and subcellular regions.

The recovery of muscle function and glycogen levels following game-play in young elite male ice hockey players.

Despite the frequent occurrence of congested game fixtures in elite ice hockey, the postgame recovery pattern has not previously been investigated. The purpose of the present study was therefore to evaluate the acute decrements and subsequent recovery of skeletal muscle glycogen levels, muscle function and repeated-sprint ability following ice hockey game-play. Sixteen male players from the Danish U20 national team completed a training game with muscle biopsies obtained before, postgame and following ~38 h of recovery (day 2). On-ice repeated-sprint ability and muscle function (maximal voluntary isometric [MVIC] and electrically induced low- (20 Hz) and high-frequency (50 Hz) knee-extensor contractions) were assessed at the same time points, as well as ~20 h into recovery (day 1). Muscle glycogen decreased 31% (p < 0.001) postgame and had returned to pregame levels on day 2. MVIC dropped 11%, whereas 50 and 20 Hz torque dropped 21% and 29% postgame, respectively, inducing a 10% reduction in the 20/50 Hz torque ratio indicative of low-frequency force depression (all p < 0.001). While MVIC torque returned to baseline on day 1, 20 and 50 Hz torque remained depressed by 9%-11% (p = 0.010-0.040), hence restoring the pre-exercise 20/50 Hz ratio. Repeated-sprint ability was only marginally reduced by 1% postgame (p = 0.041) and fully recovered on day 1. In conclusion, an elite youth ice hockey game induces substantial reductions in muscle glycogen content and muscle function, but only minor reductions in repeated-sprint ability and with complete recovery of all parameters within 1-2 days postgame.

Exercise and fatigue: integrating the role of K+, Na+ and Cl- in the regulation of sarcolemmal excitability of skeletal muscle.

Perturbations in K+ have long been considered a key factor in skeletal muscle fatigue. However, the exercise-induced changes in K+ intra-to-extracellular gradient is by itself insufficiently large to be a major cause for the force decrease during fatigue unless combined to other ion gradient changes such as for Na+. Whilst several studies described K+-induced force depression at high extracellular [K+] ([K+]e), others reported that small increases in [K+]e induced potentiation during submaximal activation frequencies, a finding that has mostly been ignored. There is evidence for decreased Cl- ClC-1 channel activity at muscle activity onset, which may limit K+-induced force depression, and large increases in ClC-1 channel activity during metabolic stress that may enhance K+ induced force depression. The ATP-sensitive K+ channel (KATP channel) is also activated during metabolic stress to lower sarcolemmal excitability. Taking into account all these findings, we propose a revised concept in which K+ has two physiological roles: (1) K+-induced potentiation and (2) K+-induced force depression. During low-moderate intensity muscle contractions, the K+-induced force depression associated with increased [K+]e is prevented by concomitant decreased ClC-1 channel activity, allowing K+-induced potentiation of sub-maximal tetanic contractions to dominate, thereby optimizing muscle performance. When ATP demand exceeds supply, creating metabolic stress, both KATP and ClC-1 channels are activated. KATP channels contribute to force reductions by lowering sarcolemmal generation of action potentials, whilst ClC-1 channel enhances the force-depressing effects of K+, thereby triggering fatigue. The ultimate function of these changes is to preserve the remaining ATP to prevent damaging ATP depletion.

Reduced 3-year risk of hospital admission and mortality after 12-week resistance training of cirrhosis patients: A follow-up of a randomized clinical trial.

BACKGROUND AND AIM: Physical activity confers health benefits in many diseases but remains almost unstudied for cirrhosis. We investigated whether a period of resistance training affects the subsequent long-term risk of hospitalization or mortality among patients with cirrhosis. METHODS: The study includes 39 participants with cirrhosis Child-Pugh class A/B who participated in a prior clinical trial randomized to either resistance training three times per week for 12 weeks or a control group. We gathered data through medical records from trial entry and the following 3 years. The outcomes were time to first hospitalization and all-cause mortality. We used regression models to examine the associations between trial groups and outcomes, adjusting for Child-Pugh class, age, gender, and comorbidity. RESULTS: Nine patients who trained and 15 controls were hospitalized, resulting in a lower risk of first hospitalization in the training group (adjusted subdistribution hazard ratio of 0.40, 95% confidence interval [CI] [0.17, 0.92]; P = 0.03). One patient who trained and six controls died, resulting in a lower all-cause mortality in the training group (adjusted hazard ratio of 0.06, 95% CI [0.01, 0.66]; P = 0.02). CONCLUSION: Twelve weeks of resistance training was associated with a reduced risk of first hospitalization and mortality among patients with cirrhosis Child-Pugh class A/B 3 years after trial entry. The mechanisms of this effect are not identified, and larger studies are warranted.

Force potentiation during eccentric contractions in rat skeletal muscle.

Postactivation potentiation refers to an acute enhancement of contractile properties following muscle activity. Previously, the effects of prior muscle activation on eccentric force at tetanic activation frequencies have only been sparsely reported. This paper aimed to study acute activity-induced effects on eccentric force of slow and fast-twitch muscles and characterize them in relation to postactivation potentiation. We elicited eccentric contractions in isolated rat extensor digitorum longus and soleus muscles by actively lengthening muscles at a constant velocity. We assessed contractile properties by measuring force over shortly interspaced, identical eccentric, and isometric contractions. We then analyzed stretch force, isometric peak force, rate of force development, and relaxation times. Finally, we compared the time courses for the development and cessation of changes in stretch force to known features of postactivation potentiation. In extensor digitorum longus, muscles stretch force consistently increased in a contraction-to-contraction manner by up to 49% [95% confidence interval (CI): 35-64%] whereas isometric peak force simultaneously showed minor declines (8%, 95% CI: 5-10%). The development and cessation of eccentric force potentiation coincided with the development of twitch potentiation and increases in rate of force development. In soleus muscles we found no consistent eccentric potentiation. Characterization of the increase in eccentric force revealed that force only increased in the very beginning of an active stretch. Eccentric force at tetanic activation frequencies potentiates substantially in extensor digitorum longus muscles over consecutive contractions with a time course coinciding with postactivation potentiation. Such eccentric potentiation may be important in sport performance.NEW & NOTEWORTHY Force during eccentric contractions can increase to a magnitude that may have profound consequences for our understanding of skeletal muscle locomotion. This increase in eccentric force occurs over consecutive, shortly interspaced, tetanic contractions in rat extensor digitorum longus muscles-not in rat soleus muscles-and coincides with well-known traits of postactivation potentiation. Eccentric force potentiation may significantly enhance muscle performance in activities involving stretch-shortening cycles.

Torque and Discomfort During Electrically Evoked Muscle Contractions in Healthy Young Adults: Influence of Stimulation Current and Pulse Frequency.

OBJECTIVE: To investigate (1) how current and pulse frequency of electrical stimulation (ES) as well as contraction mode (isometric, concentric, and eccentric) influence torque output and discomfort and (2) how familiarization by repeated ES sessions influences ratings of perceived discomfort. DESIGN: An experimental study, 3 sessions. SETTING: A university laboratory. PARTICIPANTS: Eight healthy participants (5 men, 3 women; mean age 25.2 years; N=8). INTERVENTIONS: Participants completed 3 trial days, each including 17 electrically evoked thigh muscle contractions. On each trial day, the first 6 contractions consisted of 2 isometric, 2 concentric, and 2 eccentric muscle contractions randomly ordered with a fixed stimulation current and pulse frequency (200 mA, 20 Hz), while the remaining 11 muscle contractions were all isometric with randomly ordered combinations of current (100-250 mA) and pulse frequency (20-100 Hz). MAIN OUTCOME MEASURES: Torque and perceived discomfort were measured for each ES-evoked contraction. RESULTS: Overall, the findings revealed that a higher stimulation frequency was associated with an increased torque without increased discomfort, while higher currents were associated with increases of both torque and discomfort. Contraction type did not influence level of discomfort, despite eccentric contractions eliciting higher torque compared with concentric and isometric contractions (P<.001). Finally, a significant familiarization to ES (P<.001) was observed after just 1 of 3 identical stimulation sessions. CONCLUSIONS: The outlined data suggest that to elicit high torque levels while minimizing levels of discomfort in young subjects, eccentric muscle contractions evoked with a low stimulation current, and a high pulse frequency are preferable. Furthermore, a single familiarization session significantly lowers rating of perceived discomfort during ES.

Prolonged loss of force and power following fatiguing contractions in rat soleus muscles. Is low-frequency fatigue an issue during dynamic contractions?

Low-frequency fatigue (LFF) is defined by a relatively larger deficit in isometric force elicited by low-frequency electrical stimulation compared with high-frequency stimulation. However, the effects of LFF on power during dynamic contractions elicited at low and high frequencies have not been thoroughly characterized. In the current study, rat soleus muscles underwent fatiguing either concentric, eccentric, or isometric contractions. Before and 1 h after the fatiguing contractions, a series of brief isometric and dynamic contractions elicited at 20 and 80 Hz stimulation to establish force-velocity relationships. Maximal force (Fmax), velocity (Vmax), and power (Pmax) were assessed for each frequency. Sarcoplasmic reticulum (SR) Ca2+ release and reuptake rates were assessed pre- and postfatigue. Prolonged fatigue was observed as a loss of Fmax and Pmax in muscles fatigued by concentric or eccentric, but not by isometric contractions. When quantified as a decrease in the ratio between 20 Hz and 80 Hz contractile output, LFF was more pronounced for isometric force than for power (-21% vs. -16% for concentrically fatigued muscles, P = 0.003; 29 vs. 13% for eccentrically fatigued muscles, P < 0.001). No changes in SR Ca2+ release or reuptake rates were observed. We conclude that LFF is less pronounced when expressed in terms of power deficits than when expressed in terms of force deficits, and that LFF, therefore, likely affects performance to a lesser degree during fast concentric contractions than during static or slow contractions.

Fibre type- and localisation-specific muscle glycogen utilisation during repeated high-intensity intermittent exercise.

Glycogen particles are situated in key areas of the muscle cell in the vicinity of the main energy-consumption sites and may be utilised heterogeneously dependent on the nature of the metabolic demands. The present study aimed to investigate the time course of fibre type-specific utilisation of muscle glycogen in three distinct subcellular fractions (intermyofibrillar, IMF; intramyofibrillar, Intra; and subsarcolemmal, SS) during repeated high-intensity intermittent exercise. Eighteen moderately to well-trained male participants performed three periods of 10 × 45 s cycling at ∼105% watt max (EX1-EX3) coupled with 5 × 6 s maximal sprints at baseline and after each period. Muscle biopsies were sampled at baseline and after EX1 and EX3. A higher glycogen breakdown rate in type 2 compared to type 1 fibres was found during EX1 for the Intra (-72 vs. -45%) and IMF (-59 vs. -35%) glycogen fractions (P < 0.001) but with no differences for SS glycogen (-52 vs. -40%). In contrast, no fibre type differences were observed during EX2-EX3, where the utilisation of Intra and IMF glycogen in type 2 fibres was reduced, resulting in depletion of all three subcellular fractions to very low levels post-exercise within both fibre types. Importantly, large heterogeneity in single-fibre glycogen utilisation was present with an early depletion of especially Intra glycogen in individual type 2 fibres. In conclusion, there is a clear fibre type- and localisation-specific glycogen utilisation during high-intensity intermittent exercise, which varies with time course of exercise and is characterised by exacerbated pool-specific glycogen depletion at the single-fibre level. KEY POINTS: Muscle glycogen is the major fuel during high-intensity exercise and is stored in distinct subcellular areas of the muscle cell in close vicinity to the main energy consumption sites. In the present study quantitative electron microscopy imaging was used to investigate the utilisation pattern of three distinct subcellular muscle glycogen fractions during repeated high-intensity intermittent exercise. It is shown that the utilisation differs dependent on fibre type, subcellular localisation and time course of exercise and with large single-fibre heterogeneity. These findings expand on our understanding of subcellular muscle glycogen metabolism during exercise and may help us explain how reductions in muscle glycogen can attenuate muscle function even at only moderately lowered whole-muscle glycogen concentrations.

The Role of Muscle Glycogen Content and Localization in High-Intensity Exercise Performance: A Placebo-Controlled Trial.

PURPOSE: We investigated the coupling between muscle glycogen content and localization and high-intensity exercise performance using a randomized, placebo-controlled, parallel-group design with emphasis on single-fiber subcellular glycogen concentrations and sarcoplasmic reticulum Ca 2+ kinetics. METHODS: Eighteen well-trained participants performed high-intensity intermittent glycogen-depleting exercise, followed by randomization to a high- (CHO; ~1 g CHO·kg -1 ·h -1 ; n = 9) or low-carbohydrate placebo diet (PLA, <0.1 g CHO·kg -1 ·h -1 ; n = 9) for a 5-h recovery period. At baseline, after exercise, and after the carbohydrate manipulation assessments of repeated sprint ability (5 × 6-s maximal cycling sprints with 24 s of rest), neuromuscular function and ratings of perceived exertion during standardized high-intensity cycling (~90% Wmax ) were performed, while muscle and blood samples were collected. RESULTS: The exercise and carbohydrate manipulations led to distinct muscle glycogen concentrations in CHO and PLA at the whole-muscle (291 ± 78 vs 175 ± 100 mmol·kg -1 dry weight (dw), P = 0.020) and subcellular level in each of three local regions ( P = 0.001-0.046). This was coupled with near-depleted glycogen concentrations in single fibers of both main fiber types in PLA, especially in the intramyofibrillar region (within the myofibrils). Furthermore, increased ratings of perceived exertion and impaired repeated sprint ability (~8% loss, P < 0.001) were present in PLA, with the latter correlating moderately to very strongly ( r = 0.47-0.71, P = 0.001-0.049) with whole-muscle glycogen and subcellular glycogen fractions. Finally, sarcoplasmic reticulum Ca 2+ uptake, but not release, was superior in CHO, whereas neuromuscular function, including prolonged low-frequency force depression, was unaffected by dietary manipulation. CONCLUSIONS: Together, these results support an important role of muscle glycogen availability for high-intensity exercise performance, which may be mediated by reductions in single-fiber levels, particularly in distinct subcellular regions, despite only moderately lowered whole-muscle glycogen concentrations.

Short communication: Leucine, but not muscle contractions, stimulates protein synthesis in isolated EDL muscles from golden geckos.

Resistance exercise and protein ingestion stimulate muscle protein synthesis in mammals and the combination of both stimuli exert an additive effect. However, mechanisms regulating muscle mass may be different in ectothermic vertebrates because these animals are adapted to low energy consumption, short bouts of physical activity, and prolonged periods of inactivity. Here, we investigated the effects of administration of leucine and simulated resistance exercise induced by electrical stimulation (ES) on protein synthesis rate in isolated extensor digitorum longus muscle from golden geckos (Gekko badenii). Muscles were placed in Krebs-Ringer buffer equilibrated with O2 (97%) and CO2 (3%) at 30 °C. One muscle from each animal was subjected to one of three interventions: 1) administration of leucine (0.5 mM) at rest, 2) isometric contractions evoked by ES, or 3) a combination of contractions and leucine, while the contralateral muscle served as untreated control. The rate of protein synthesis was measured through pyromycin-labeling. Administration of leucine led to a 2.75 (±1.88)-fold rise in protein synthesis rate in inactive muscles, whereas isometric contractions had no effect (0.67 ± 0.37-fold). The combination of isometric contractions and leucine did not affect protein synthesis rate (1.02 ± 0.34-fold), suggesting that muscle contractions attenuated the positive influence of leucine. Our study identifies leucine as a potent positive regulator of muscle protein synthesis in golden geckos, but also demonstrates that muscle contraction is not. More studies should be conducted in other taxonomic groups of ectothermic vertebrates to identify whether this is a general pattern.

Potentiation of force by extracellular potassium and posttetanic potentiation are additive in mouse fast-twitch muscle in vitro.

The influence of moderately elevated extracellular potassium concentration ([K+]) on muscle force has marked similarities to that of posttetanic potentiation (PTP) in that twitch force may be enhanced whilst high-frequency force is depressed. The purpose of this work was to test whether K+-induced potentiation is mechanistically related to PTP via skeletal myosin light-chain kinase (skMLCK)-catalyzed phosphorylation of the myosin regulatory light chains (RLC). To do this, we assessed the influence of elevated [K+] on the force response at various frequencies in extensor digitorum longus (EDL) muscles isolated from wild-type and skeletal myosin light-chain kinase (skMLCK-/-) absent mice. Changing [K+] of the incubation medium from 5 to 10 mmol increased isometric twitch force by a similar amount in wild-type and skMLCK-/- muscles (~ 13% in both genotypes) (all data n = 7-8, P < 0.05). In contrast, 100- and 200-Hz forces were depressed in both genotypes (by 5-7 and 15-18%, respectively). The isometric twitch potentiation caused by a tetanic stimulus series was similar at both [K+] levels for each genotype but was much greater for wild-type than for skMLCK-/- muscles (i.e., 23-25 and 8-9%, respectively). Thus, we conclude that [K+]- and stimulation-induced potentiation are additive and that [K+]-induced potentiation is independent of RLC phosphorylation.

Effects of progressive resistance training in individuals with type 2 diabetic polyneuropathy: a randomised assessor-blinded controlled trial.

AIMS/HYPOTHESIS: The aim of this study was to evaluate the effects of progressive resistance training (PRT) on muscle strength, intraepidermal nerve fibre density (IENFD) and motor function in individuals with type 2 diabetic polyneuropathy (DPN) and to compare potential adaptations to those of individuals with type 2 diabetes without DPN and healthy controls. METHODS: This was an assessor-blinded trial conducted at the Neurology department, Aarhus University Hospital. Adults with type 2 diabetes, with and without DPN and healthy control participants were randomised to either supervised PRT or non-PRT for 12 weeks. Allocation was concealed by a central office unrelated to the study. The co-primary outcomes were muscle strength in terms of the peak torque of the knee and ankle extensors and flexors, and IENFD. Secondary outcome measures included the 6 min walk test (6MWT), five-time sit-to-stand test (FTSST) and postural stability index obtained by static posturography. RESULTS: A total of 109 individuals were enrolled in three groups (type 2 diabetes with DPN [n = 42], type 2 diabetes without DPN [n = 32] and healthy control [n = 35]). PRT resulted in muscle strength gains of the knee extensors and flexors in all three groups using comparative analysis (DPN group, PRT 10.3 ± 9.6 Nm vs non-PRT -0.4 ± 8.2 Nm; non-DPN group, PRT 7.5 ± 5.8 Nm vs non-PRT 0.6 ± 8.8 Nm; healthy control group, PRT 6.3 ± 9.0 Nm vs non-PRT -0.4 ± 8.4 Nm; p<0.05, respectively). Following PRT the DPN group improved the 6MWT (PRT 34.6 ± 40.9 m vs non-PRT 2.7 ± 19.6 m; p=0.001) and the FTSST (PRT -1.5 ± 2.2 s vs non-PRT 1.5 ± 4.6 s; p=0.02). There was no change in IENFD following PRT in any of the groups. CONCLUSIONS/INTERPRETATION: PRT improved muscle strength of the knee extensors and flexors and motor function in individuals with type 2 diabetic polyneuropathy at levels comparable with those seen in individuals with diabetes without DPN and healthy control individuals, while no effects were observed in IENFD. TRIAL REGISTRATION: ClinicalTrials.gov NCT03252132 FUNDING: Research reported in this paper is part of the International Diabetic Neuropathy Consortium (IDNC) research programme, supported by a Novo Nordisk Foundation Challenge Program grant (grant no. NNF14OC0011633) and Aarhus University.

Potassium-induced potentiation of subtetanic force in rat skeletal muscles: influences of ß2-activation, lactic acid, and temperature.

Moderate elevations of extracellular K+ concentration ([K+]o) occur during exercise and have been shown to potentiate force during contractions elicited with subtetanic frequencies. Here, we investigated whether lactic acid (reduced chloride conductance), ß2-adrenoceptor activation, and increased temperature would influence the potentiating effect of potassium in slow- and fast-twitch muscles. Isometric contractions were elicited by electrical stimulation at various frequencies in isolated rat soleus and extensor digitorum longus (EDL) muscles incubated at normal (4 mM) or elevated K+, in combination with salbutamol (5 µM), lactic acid (18.1 mM), 9-anthracene-carboxylic acid (9-AC; 25 µM), or increased temperature (30-35°C). Elevating [K+]o from 4 mM to 7 mM (soleus) and 10 mM (EDL) potentiated isometric twitch and subtetanic force while slightly reducing tetanic force. In EDL, salbutamol further augmented twitch force (+27 ± 3%, P < 0.001) and subtetanic force (+22 ± 4%, P < 0.001). In contrast, salbutamol reduced subtetanic force (-28 ± 6%, P < 0.001) in soleus muscles. Lactic acid and 9-AC had no significant effects on isometric force of muscles already exposed to moderate elevations of [K+]o. The potentiating effect of elevated [K+]o was still well maintained at 35°C. Addition of salbutamol exerts a further force-potentiating effect in fast-twitch but not in slow-twitch muscles already potentiated by moderately elevated [K+]o, whereas lactic acid, 9-AC, or increased temperature does not exert any further augmentation. However, the potentiating effect of elevated [K+]o was still maintained in the presence of these, thus emphasizing the positive influence of moderately elevated [K+]o for contractile performance during exercise.

Free diving-inspired breathing techniques for COPD patients: A pilot study.

Objectives: Pulmonary rehabilitation (PR) is a key factor in enhancing self-management and exercise capacity in patients with chronic obstructive pulmonary disease (COPD). The content and length of PR varies between countries and authorities responsible for rehabilitation. After completion of rehabilitation, it is often difficult for patients to stay motivated and perform regular exercise. Methods: In this pilot study, nine patients with moderate to severe COPD completed a 6-week training programme consisting of free diving-inspired breathing techniques, designed to be incorporated into daily activities. Results: Participants significantly increased the distance walked in 6 min by 48 m (p < 0.05) and a significant reduction was seen on the COPD self-efficacy scale (p < 0.05). Furthermore, adherence to the programme sessions was very high at 96.3% and no adverse events occurred. Discussion: This pilot study tested the feasibility of introducing breathing techniques used by COPD patients to enhance their walking capacity. The techniques were well tolerated and participant's adherence to the weekly group sessions was high.