RESUMEN
Skeletal muscle atrophy is a morbidity and mortality risk factor that happens with disuse, chronic disease, and aging. The tissue remodeling that happens during recovery from atrophy or injury involves changes in different cell types such as muscle fibers, and satellite and immune cells. Here, we show that the previously uncharacterized gene and protein Zfp697 is a damage-induced regulator of muscle remodeling. Zfp697/ZNF697 expression is transiently elevated during recovery from muscle atrophy or injury in mice and humans. Sustained Zfp697 expression in mouse muscle leads to a gene expression signature of chemokine secretion, immune cell recruitment, and extracellular matrix remodeling. Notably, although Zfp697 is expressed in several cell types in skeletal muscle, myofiber-specific Zfp697 genetic ablation in mice is sufficient to hinder the inflammatory and regenerative response to muscle injury, compromising functional recovery. We show that Zfp697 is an essential mediator of the interferon gamma response in muscle cells and that it functions primarily as an RNA-interacting protein, with a very high number of miRNA targets. This work identifies Zfp697 as an integrator of cell-cell communication necessary for tissue remodeling and regeneration.
Asunto(s)
Músculo Esquelético , Proteínas de Unión al ARN , Animales , Ratones , Músculo Esquelético/metabolismo , Músculo Esquelético/patología , Proteínas de Unión al ARN/metabolismo , Proteínas de Unión al ARN/genética , Humanos , Inflamación/metabolismo , Inflamación/patología , Inflamación/genética , Ratones Noqueados , Atrofia Muscular/metabolismo , Atrofia Muscular/genética , Atrofia Muscular/patología , MicroARNs/genética , MicroARNs/metabolismo , Ratones Endogámicos C57BL , Interferón gamma/metabolismoRESUMEN
Skeletal muscle fibers need to have mechanisms to decrease energy consumption during intense physical exercise to avoid devastatingly low ATP levels, with the formation of rigor cross-bridges and defective ion pumping. These protective mechanisms inevitably lead to declining contractile function in response to intense exercise, characterizing fatigue. Through our work we have gained insights into cellular and molecular mechanisms underlying the decline in contractile function during acute fatigue. Key mechanistic insights have been gained from studies performed on intact and skinned single muscle fibers, and more recently from studies performed and single myosin molecules. Studies on intact single fibers revealed several mechanisms of impaired sarcoplasmic reticulum (SR) Ca2+ release and experiments on single myosin molecules provide direct evidence of how putative agents of fatigue impact myosin's ability to generate force and motion. We conclude that changes in metabolites due to an increased dependency on anaerobic metabolism (e.g. accumulation of inorganic phosphate ions and H+) act to directly and indirectly (i.e. via decreased Ca2+ activation) inhibit myosin's force and motion generating capacity. These insights into the acute mechanisms of fatigue may help improve endurance training strategies and reveal potential targets for therapies to attenuate fatigue in chronic diseases.
RESUMEN
The protein α-actinin-3 expressed in fast-twitch skeletal muscle fiber is absent in 1.5 billion people worldwide due to homozygosity for a nonsense polymorphism in ACTN3 (R577X). The prevalence of the 577X allele increased as modern humans moved to colder climates, suggesting a link between α-actinin-3 deficiency and improved cold tolerance. Here, we show that humans lacking α-actinin-3 (XX) are superior in maintaining core body temperature during cold-water immersion due to changes in skeletal muscle thermogenesis. Muscles of XX individuals displayed a shift toward more slow-twitch isoforms of myosin heavy chain (MyHC) and sarcoplasmic reticulum (SR) proteins, accompanied by altered neuronal muscle activation resulting in increased tone rather than overt shivering. Experiments on Actn3 knockout mice showed no alterations in brown adipose tissue (BAT) properties that could explain the improved cold tolerance in XX individuals. Thus, this study provides a mechanism for the positive selection of the ACTN3 X-allele in cold climates and supports a key thermogenic role of skeletal muscle during cold exposure in humans.
Asunto(s)
Actinina/genética , Termogénesis/genética , Tejido Adiposo Pardo/metabolismo , Animales , Temperatura Corporal/genética , Codón sin Sentido/genética , Evolución Molecular , Humanos , Masculino , Ratones , Ratones Noqueados , Músculo Esquelético/metabolismo , Selección Genética/genéticaRESUMEN
During the initial phase of fatigue induced by repeated contractions in fast-twitch muscle fibers, tetanic force decreases despite increasing tetanic free cytosolic [Ca2+ ] ([Ca2+ ]cyt ). Here, we hypothesized that the increase in tetanic [Ca2+ ]cyt nevertheless has positive effects on force in early fatigue. Experiments on enzymatically isolated mouse flexor digitorum brevis (FDB) fibers showed that an increase in tetanic [Ca2+ ]cyt during ten 350 ms contractions required trains of electrical pulses to be elicited at short intervals (≤2 s) and at high frequencies (≥70 Hz). Mechanically dissected mouse FDB fibers showed greater decrease in tetanic force when the stimulation frequency during contractions was gradually reduced to prevent the increase in tetanic [Ca2+ ]cyt . Novel analyses of data from previous studies revealed an increased rate of force development in the tenth fatiguing contraction in mouse FDB fibers, as well as in rat FDB and human intercostal fibers. Mouse FDB fibers deficient in creatine kinase showed no increase in tetanic [Ca2+ ]cyt and slowed force development in the tenth contraction; after injection of creatine kinase to enable phosphocreatine breakdown, these fibers showed an increase in tetanic [Ca2+ ]cyt and accelerated force development. Mouse FDB fibers exposed to ten short contractions (43 ms) produced at short intervals (142 ms) showed increased tetanic [Ca2+ ]cyt accompanied by a marked (~16%) increase in the developed force. In conclusion, the increase in tetanic [Ca2+ ]cyt in early fatigue is accompanied by accelerated force development, which under some circumstances can counteract the decline in physical performance caused by the concomitant decrease in maximum force.
Asunto(s)
Contracción Muscular , Fatiga Muscular , Humanos , Ratones , Ratas , Animales , Fatiga Muscular/fisiología , Contracción Muscular/fisiología , Calcio/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/metabolismo , Creatina Quinasa , Mamíferos/metabolismoRESUMEN
Interval training (IT) results in improved fatigue resistance in skeletal muscle mainly due to an increased aerobic capacity, which involves increased muscle mitochondrial content and/or improved mitochondrial function. We hypothesized that IT with high-intensity contractions is more effective in increasing mitochondrial function, and hence fatigue resistance, than low-intensity contractions. To study this hypothesis without interference from differences in muscle fiber recruitment obliged to occur during voluntary contractions, IT was performed with in situ supramaximal electrical stimulation where all muscle fibers are recruited. We compared the effect of IT with repeated low-intensity (20 Hz stimulation, IT20) and high-intensity (100 Hz stimulation, IT100) contractions on fatigue resistance and mitochondrial content and function in mouse plantar flexor muscles. Muscles were stimulated every other day for 4 weeks. The averaged peak torque during IT bouts was 4.2-fold higher with IT100 than with IT20. Both stimulation protocols markedly improved in situ fatigue resistance, although the improvement was larger with IT100. The citrate synthase activity, a biomarker of mitochondrial content, was similarly increased with IT20 and IT100. Conversely, increased expression of mitochondrial respiratory chain (MRC) complexes I, III, and IV was only observed with IT100 and this was accompanied by increases in MRC supercomplex formation and pyruvate-malate-driven state 3 respiration in isolated mitochondria. In conclusion, the IT-induced increase in fatigue resistance is larger with high-intensity than with low-intensity contractions and this is linked to improved mitochondrial function due to increased expression of MRC complexes and assembly of MRC supercomplexes.
Asunto(s)
Entrenamiento de Intervalos de Alta Intensidad/métodos , Mitocondrias/metabolismo , Contracción Muscular , Fatiga Muscular , Músculo Esquelético/metabolismo , Animales , Biomarcadores/metabolismo , Femenino , Ratones , Ratones Endogámicos BALB C , Músculo Esquelético/citologíaRESUMEN
PURPOSE: Unaccustomed eccentric contractions generally result in a long-lasting contractile impairment, referred to as prolonged low-frequency force depression (PLFFD), and delayed-onset muscle soreness (DOMS). We here used repeated drop jumps (DJs) as an eccentric contraction model and studied the effects of increasing the time between DJs from 20 s to 5 min. We hypothesized that both PLFFD and DOMS would be less marked at the longer DJ interval due to the longer time to restore structural elements between DJs. METHODS: Young men (n = 12) randomly performed 50 DJs with either 20-s (DJ-20 s) or 5-min (DJ-5 min) rest between DJs. Voluntary, 20 Hz and 100 Hz electrically stimulated isometric knee extension torques and muscle soreness were monitored before and for 7 days after DJs; serum CK activity was measured to assess muscle fibre protein leakage. In additional experiments, changes in mRNA levels were assessed in muscle biopsies collected before and 1 h after exercise. RESULTS: A marked PLFFD was observed with both protocols and the extent of 20 Hz torque depression was smaller immediately and 1 day after DJ-5 min than after DJ-20 s (p < 0.05), whereas the MVC and 100 Hz torques were similarly decreased with the two protocols. Markedly larger differences between the two protocols were observed for the muscle soreness score, which 1-4 days after exercise was about two times larger with DJ-20 s than with DJ-5 min (p < 0.01). CONCLUSIONS: The larger protective effect of the longer DJ interval against DOMS than against PLFFD indicates that their underlying mechanisms involve different structural elements.
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Rodilla/fisiología , Contracción Muscular/fisiología , Mialgia/prevención & control , Descanso , Adulto , Biomarcadores/sangre , Biopsia con Aguja , Creatina Quinasa/sangre , Estimulación Eléctrica , Humanos , Masculino , Dimensión del Dolor , Factores de Tiempo , Torque , Adulto JovenRESUMEN
KEY POINTS: Changes in intramuscular Ca2+ handling contribute to development of fatigue and disease-related loss of muscle mass and function. To date, no data on human intact living muscle fibres have been described. We manually dissected intact single fibres from human intercostal muscle and simultaneously measured force and myoplasmic free [Ca2+ ] at physiological temperature. Based on their fatigue resistance, two distinct groups of fibres were distinguished: fatigue sensitive and fatigue resistant. Force depression in fatigue and during recovery was due to impaired sarcoplasmic reticulum Ca2+ release in both groups of fibres. Acidification did not affect force production in unfatigued fibres and did not affect fatigue development in fatigue-resistant fibres. The current study provides novel insight into the mechanisms of fatigue in human intercostal muscle. ABSTRACT: Changes in intracellular Ca2+ handling of individual skeletal muscle fibres cause a force depression following physical activity and are also implicated in disease-related loss of function. The relation of intracellular Ca2+ handling with muscle force production and fatigue tolerance is best studied in intact living single fibres that allow continuous measurements of force and myoplasmic free [Ca2+ ] during repeated contractions. To this end, manual dissections of human intercostal muscle biopsies were performed to isolate intact single fibres. Based on the ability to maintain tetanic force at >40% of the initial value during 500 fatiguing contractions, fibres were classified as either fatigue sensitive or fatigue resistant. Following fatigue all fibres demonstrated a marked reduction in sarcoplasmic reticulum Ca2+ release, while myofibrillar Ca2+ sensitivity was either unaltered or increased. In unfatigued fibres, acidosis caused a reduction in myofibrillar Ca2+ sensitivity that was offset by increased tetanic myoplasmic free [Ca2+ ] so that force remained unaffected. Acidification did not affect the fatigue tolerance of fatigue-resistant fibres, whereas uncertainties remain whether or not fatigue-sensitive fibres were affected. Following fatigue, a prolonged force depression at preferentially low-frequency stimulation was evident in fatigue-sensitive fibres and this was caused exclusively by an impaired sarcoplasmic reticulum Ca2+ release. We conclude that impaired sarcoplasmic reticulum Ca2+ release is the predominant mechanism of force depression both in the development of, and recovery from, fatigue in human intercostal muscle.
Asunto(s)
Señalización del Calcio , Músculos Intercostales/fisiopatología , Fatiga Muscular , Fibras Musculares Esqueléticas/patología , Retículo Sarcoplasmático/patología , Calcio/fisiología , Humanos , Técnicas In Vitro , Contracción MuscularRESUMEN
Prolonged low-frequency force depression (PLFFD) induced by fatiguing exercise is characterized by a persistent depression in submaximal contractile force during the recovery period. Muscle glycogen depletion is known to limit physical performance during prolonged low- and moderate-intensity exercise, and accelerating glycogen resynthesis with post-exercise carbohydrate intake can facilitate recovery and improve repeated bout exercise performance. Short-term, high-intensity exercise, however, can cause PLFFD without any marked decrease in glycogen. Here, we studied whether recovery from PLFFD was accelerated by carbohydrate ingestion after 60 minutes of moderate-intensity glycogen-depleting cycling exercise followed by six 30-seconds all-out cycling sprints. We used a randomized crossover study design where nine recreationally active males drank a beverage containing either carbohydrate or placebo after exercise. Blood glucose and muscle glycogen concentrations were determined at baseline, immediately post-exercise, and during the 3-hours recovery period. Transcutaneous electrical stimulation of the quadriceps muscle was performed to determine the extent of PLFFD by eliciting low-frequency (20 Hz) and high-frequency (100 Hz) stimulations. Muscle glycogen was severely depleted after exercise, with a significantly higher rate of muscle glycogen resynthesis during the 3-hours recovery period in the carbohydrate than in the placebo trials (13.7 and 5.4 mmol glucosyl units/kg wet weight/h, respectively). Torque at 20 Hz was significantly more depressed than 100 Hz torque during the recovery period in both conditions, and the extent of PLFFD (20/100 Hz ratio) was not different between the two trials. In conclusion, carbohydrate supplementation enhances glycogen resynthesis after glycogen-depleting exercise but does not improve force recovery when the exercise also involves all-out cycling sprints.
Asunto(s)
Glucemia/metabolismo , Carbohidratos de la Dieta/administración & dosificación , Ejercicio Físico , Glucógeno/metabolismo , Contracción Muscular , Músculo Esquelético/metabolismo , Adolescente , Adulto , Bebidas , Estudios Cruzados , Humanos , Masculino , Músculo Cuádriceps , Adulto JovenRESUMEN
Measuring free Ca2+ concentration ([Ca2+]) in the cytosol or organelles is routine in many fields of research. The availability of membrane permeant forms of indicators coupled with the relative ease of transfecting cell lines with biological Ca2+ sensors have led to the situation where cellular and subcellular [Ca2+] is examined by many non-specialists. In this chapter, we evaluate the most used Ca2+ indicators and highlight what their major advantages and disadvantages are. We stress the potential pitfalls of non-ratiometric techniques for measuring Ca2+ and the clear advantages of ratiometric methods. Likely improvements and new directions for Ca2+ measurement are discussed.
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Calcio , Citosol , Orgánulos , Animales , Calcio/metabolismo , Técnicas Citológicas , Citosol/química , Citosol/metabolismo , Humanos , Orgánulos/química , Orgánulos/metabolismoRESUMEN
Background and Objectives: The all-out mode of sprint interval training (SIT) has been shown to be an efficient method for improving sports performance, exercise capacity, and aerobic fitness. Although the benefits of SIT are well described, the mechanisms underlying the different degrees of response remain largely unexplored. We aimed to assess the effects of exertion on the responsiveness to SIT. Materials and Methods: The participants were 28 young untrained men (mean ± SD age 25.7 ± 6.03 years) who exhibited either a large or small increase in Wingate test average power in response to nine SIT sessions performed over three weeks. Each training session comprised four-six bouts of 30 s all-out cycling interspaced with 4 min of rest. Individual responses were assessed using heart rate (HR) during exercise for all nine sessions, as well as blood lactate concentration up to 1 h, and the decrement in maximal voluntary knee extension torque (MVC) up to 24 h after the first and last training sessions. Peak oxygen uptake (VO2peak) and maximum HR were measured before and after training during an incremental cycling test to exhaustion. Results: Although all participants showed benefits of SIT such as increased VO2peak, the increase in anaerobic cycling power varied between participants. We identified 17 high responders and nine low responders, whose average power outputs were 0.80 ± 0.22 and 0.22 ± 0.19 W/kg, respectively. The HR achieved during any of the training sessions did not differ between high and low responders. The lactate kinetics did not differ between groups before and after the intervention. Training resulted in a more rapid recovery of MVC without any discernible differences between the high and low responders. Conclusion: The differences in the responses to SIT are not dependent on the exertion level during training.
Asunto(s)
Entrenamiento de Intervalos de Alta Intensidad/métodos , Esfuerzo Físico/fisiología , Adaptación Fisiológica/fisiología , Adulto , Rendimiento Atlético/fisiología , Humanos , Masculino , Consumo de Oxígeno/fisiología , Carrera/fisiologíaRESUMEN
The extracellular K+ concentration ([K+]o) increases during physical exercise. We here studied whether moderately elevated [K+]o may increase force and power output during contractions at in vivo-like subtetanic frequencies and whether such potentiation was associated with increased cytosolic free Ca2+ concentration ([Ca2+]i) during contractions. Isolated whole soleus and extensor digitorum longus (EDL) rat muscles were incubated at different levels of [K+]o, and isometric and dynamic contractility were tested at various stimulation frequencies. Furthermore, [Ca2+]i at rest and during contraction was measured along with isometric force in single mouse flexor digitorum brevis (FDB) fibers exposed to elevated [K+]o. Elevating [K+]o from 4 mM up to 8 mM (soleus) and 11 mM (EDL) increased isometric force at subtetanic frequencies, 2-15 Hz in soleus and up to 50 Hz in EDL, while inhibition was seen at tetanic frequency in both muscle types. Elevating [K+]o also increased peak power of dynamic subtetanic contractions, with potentiation being more pronounced in EDL than in soleus muscles. The force-potentiating effect of elevated [K+]o was transient in FDB single fibers, reaching peak after ~4 and 2.5 min in 9 and 11 mM [K+]o, respectively. At the time of peak potentiation, force and [Ca2+]i during 15-Hz contractions were significantly increased, whereas force was slightly decreased and [Ca2+]i unchanged during 50-Hz contractions. Moderate elevation of [K+]o can transiently potentiate force and power during contractions at subtetanic frequencies, which can be explained by a higher [Ca2+]i during contractions.
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Calcio/metabolismo , Líquido Extracelular/metabolismo , Contracción Muscular/fisiología , Fatiga Muscular/fisiología , Músculo Esquelético/metabolismo , Potasio/metabolismo , Animales , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Técnicas de Cultivo de Órganos , Ratas , Ratas WistarRESUMEN
Skeletal muscle weakness is associated with oxidative stress and oxidative posttranslational modifications on contractile proteins. There is indirect evidence that reactive oxygen/nitrogen species (ROS/RNS) affect skeletal muscle myofibrillar function, although the details of the acute effects of ROS/RNS on myosin-actin interactions are not known. In this study, we examined the effects of peroxynitrite (ONOO-) on the contractile properties of individual skeletal muscle myofibrils by monitoring myofibril-induced displacements of an atomic force cantilever upon activation and relaxation. The isometric force decreased by ~50% in myofibrils treated with the ONOO- donor (SIN-1) or directly with ONOO-, which was independent of the cross-bridge abundancy condition (i.e., rigor or relaxing condition) during SIN-1 or ONOO- treatment. The force decrease was attributed to an increase in the cross-bridge detachment rate (gapp) in combination with a conservation of the force redevelopment rate (kTr) and hence, an increase in the population of cross-bridges transitioning from force-generating to non-force-generating cross-bridges during steady-state. Taken together, the results of this study provide important information on how ROS/RNS affect myofibrillar force production which may be of importance for conditions where increased oxidative stress is part of the pathophysiology.
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Contracción Isométrica/efectos de los fármacos , Molsidomina/análogos & derivados , Miofibrillas/efectos de los fármacos , Miosinas/antagonistas & inhibidores , Donantes de Óxido Nítrico/farmacología , Oxidantes/farmacología , Ácido Peroxinitroso/farmacología , Actinas/antagonistas & inhibidores , Actinas/química , Actinas/fisiología , Animales , Contracción Isométrica/fisiología , Molsidomina/química , Molsidomina/farmacología , Miofibrillas/fisiología , Miofibrillas/ultraestructura , Miosinas/química , Miosinas/fisiología , Donantes de Óxido Nítrico/química , Estrés Oxidativo , Músculos Psoas/efectos de los fármacos , Músculos Psoas/fisiología , Músculos Psoas/ultraestructura , Conejos , Técnicas de Cultivo de TejidosRESUMEN
Discrepant results have been reported regarding an intramuscular mechanism underlying the ergogenic effect of caffeine on neuromuscular function in humans. Here, we reevaluated the effect of caffeine on muscular force production in humans and combined this with measurements of the caffeine dose-response relationship on force and cytosolic free [Ca2+] ([Ca2+]i) in isolated mouse muscle fibers. Twenty-one healthy and physically active men (29 ± 9 yr, 178 ± 6 cm, 73 ± 10 kg, mean ± SD) took part in the present study. Nine participants were involved in two experimental sessions during which supramaximal single and paired electrical stimulations (at 10 and 100 Hz) were applied to the femoral nerve to record evoked forces. Evoked forces were recorded before and 1 h after ingestion of 1) 6 mg caffeine/kg body mass or 2) placebo. Caffeine plasma concentration was measured in 12 participants. In addition, submaximal tetanic force and [Ca2+]i were measured in single mouse flexor digitorum brevis (FDB) muscle fibers exposed to 100 nM up to 5 mM caffeine. Six milligrams of caffeine per kilogram body mass (plasma concentration ~40 µM) did not increase electrically evoked forces in humans. In superfused FDB single fibers, millimolar caffeine concentrations (i.e., 15- to 35-fold above usual concentrations observed in humans) were required to increase tetanic force and [Ca2+]i. Our results suggest that toxic doses of caffeine are required to increase muscle contractility, questioning the purported intramuscular ergogenic effect of caffeine in humans.
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Cafeína/toxicidad , Electromiografía/efectos de los fármacos , Contracción Muscular/efectos de los fármacos , Músculo Esquelético/efectos de los fármacos , Adulto , Animales , Cafeína/administración & dosificación , Cafeína/sangre , Relación Dosis-Respuesta a Droga , Electromiografía/métodos , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Contracción Muscular/fisiología , Músculo Esquelético/fisiología , Técnicas de Cultivo de Órganos , Adulto JovenRESUMEN
KEY POINTS: Skeletal muscle fatigue limits performance in various physical activities, with exercise intolerance being a key symptom in a broad spectrum of diseases. We investigated whether a small molecule fast skeletal troponin activator (FSTA), CK-2066260, can mitigate muscle fatigue by reducing the cytosolic free [Ca2+ ] required to produce a given submaximal force and hence decreasing the energy requirement. Isolated intact single mouse muscle fibres and rat muscles in-situ treated with CK-2066260 showed improved muscle endurance., which was accompanied by decreased ATP demand and reduced glycogen usage. CK-2066260 treatment improved in-vivo exercise capacity in healthy rats and in a rat model of peripheral artery insufficiency. In conclusion, we show that the FSTA CK-2066260 effectively counteracts muscle fatigue in rodent skeletal muscle in vitro, in situ, and in vivo. This may translate to humans and provide a promising pharmacological treatment to patients suffering from severe muscle weakness and exercise intolerance. ABSTRACT: Skeletal muscle fatigue limits performance during physical exercise and exacerbated muscle fatigue is a prominent symptom among a broad spectrum of diseases. The present study investigated whether skeletal muscle fatigue is affected by the fast skeletal muscle troponin activator (FSTA) CK-2066260, which increases myofibrillar Ca2+ sensitivity and amplifies the submaximal force response. Because more force is produced for a given Ca2+ , we hypothesized that CK-2066260 could mitigate muscle fatigue by reducing the energetic cost of muscle activation. Isolated single mouse muscle fibres were fatigued by 100 repeated 350 ms contractions while measuring force and the cytosolic free [Ca2+ ] or [Mg2+ ] ([Mg2+ ]i ). When starting fatiguing stimulation at matching forces (i.e. lower stimulation frequency with CK-2066260): force was decreased by â¼50% with and by â¼75% without CK-2066260; [Mg2+ ]i was increased by â¼10% with and â¼32% without CK-2066260, reflecting a larger decrease in [ATP] in the latter. The glycogen content in in situ stimulated rat muscles fatigued by repeated contractions at matching forces was about two times higher with than without CK-2066260. Voluntary exercise capacity, assessed by rats performing rotarod exercise and treadmill running, was improved in the presence of CK-2066260. CK-2066260 treatment also increased skeletal muscle fatigue resistance and exercise performance in a rat model of peripheral artery insufficiency. In conclusion, we demonstrate that the FSTA CK-2066260 mitigates skeletal muscle fatigue by reducing the metabolic cost of force generation.
Asunto(s)
Contracción Muscular/fisiología , Fatiga Muscular/fisiología , Fibras Musculares de Contracción Rápida/metabolismo , Troponina/metabolismo , Animales , Calcio/metabolismo , Femenino , Glucógeno/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Miofibrillas/metabolismo , Condicionamiento Físico Animal/fisiología , Ratas , Ratas Sprague-DawleyRESUMEN
PURPOSE: Mechanisms underlying the efficacy of sprint interval training (SIT) remain to be understood. We previously reported that an acute bout of SIT disrupts the integrity of the sarcoplasmic reticulum (SR) Ca2+ release channel, the ryanodine receptor 1 (RyR1), in recreationally active human subjects. We here hypothesize that in addition to improving the exercise performance of recreationally active humans, a period of repeated SIT sessions would make the RyR1 protein less vulnerable and accelerate recovery of contractile function after a SIT session. METHODS: Eight recreationally active males participated in a 3-week SIT program consisting of nine sessions of four-six 30-s all-out cycling bouts with 4 min of rest between bouts. RESULTS: Total work performed during a SIT session and maximal power (Wmax) reached during an incremental cycling test were both increased by ~ 7.5% at the end of the training period (P < 0.05). Western blots performed on vastus lateralis muscle biopsies taken before, 1 h, 24 h and 72 h after SIT sessions in the untrained and trained state showed some protection against SIT-induced reduction of full-length RyR1 protein expression in the trained state. SIT-induced knee extensor force deficits were similar in the untrained and trained states, with a major reduction in voluntary and electrically evoked forces immediately and 1 h after SIT (P < 0.05), and recovery after 24 h. CONCLUSIONS: Three weeks of SIT improves exercise performance and provides some protection against RyR1 modification, whereas it does not accelerate recovery of contractile function.
Asunto(s)
Ejercicio Físico/fisiología , Resistencia Física/fisiología , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , Adaptación Fisiológica/fisiología , Adulto , Prueba de Esfuerzo/métodos , Entrenamiento de Intervalos de Alta Intensidad/métodos , Humanos , Masculino , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiología , Consumo de Oxígeno/fisiología , Adulto JovenRESUMEN
KEY POINTS: We examined the mechanisms underlying the positive effect of preconditioning contractions (PCs) on the recovery of muscle force after damaging eccentric contractions (ECCs). The mechanisms underlying the immediate force decrease after damaging ECCs differ from those causing depressed force with a few days' delay, where reactive oxygen species (ROS) produced by invading immune cells play an important causative role. PCs counteracted the delayed onset force depression and this could be explained by prevention of immune cell invasion, which resulted in decreased myeloperoxidase-mediated ROS production, hence avoiding cell membrane disruption, calpain activation and degenerative changes in myosin and actin molecules. ABSTRACT: Preconditioning contractions (PCs) have been shown to result in markedly improved contractile function during the recovery periods after muscle damage from eccentric contractions (ECCs). Here, we examined the mechanisms underlying the beneficial effect of PCs with a special focus on the myofibrillar function. Rat medial gastrocnemius muscles were exposed to 100 repeated damaging ECCs in situ and excised immediately (recovery 0, REC0) or after 4 days (REC4). PCs with 10 repeated non-damaging ECCs were applied 2 days before the damaging ECCs. PCs improved in situ maximal isometric torque at REC4. Skinned muscle fibres were used to directly assess changes in myofibrillar function. PCs prevented the damaging ECC-induced depression in maximum Ca2+ -activated force at REC4. PCs also prevented the following damaging ECC-induced effects at REC4: (i) the reduction in myosin heavy chain and actin content; (ii) calpain activation; (iii) changes in redox homeostasis manifested as increased expression levels of malondialdehyde-protein adducts, NADPH oxidase 2, superoxide dismutase 2 and catalase, and activation of myeloperoxidase (MPO); (iv) infiltration of immune cells and loss of cell membrane integrity. Additionally, at REC0, PCs enhanced the expression levels of heat shock protein (HSP) 70, HSP25, and αB-crystallin in the myofibrils and prevented the increased mRNA levels of granulocyte-macrophage colony-stimulating factor and interleukin-6. In conclusion, PCs prevent the delayed force depression after damaging ECCs by an HSP-dependent inhibition of degenerative changes in myosin and actin molecules caused by myeloperoxidase-induced membrane lysis and subsequent calpain activation, which were triggered by an inflammatory reaction with immune cells invading damaged muscles.
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Contracción Isométrica , Miofibrillas/fisiología , Estrés Oxidativo , Actinas/metabolismo , Animales , Calcio/metabolismo , Calpaína/metabolismo , Células Cultivadas , Proteínas de Choque Térmico/metabolismo , Interleucina-6/metabolismo , Macrófagos/fisiología , Masculino , Miofibrillas/metabolismo , Miofibrillas/patología , Cadenas Pesadas de Miosina/metabolismo , NADPH Oxidasas/metabolismo , Neutrófilos/fisiología , Peroxidasa/metabolismo , Ratas , Ratas Wistar , Especies Reactivas de Oxígeno/metabolismo , Superóxido Dismutasa/metabolismoRESUMEN
Increased production of reactive oxygen/nitrogen species (ROS) and impaired cellular Ca2+ handling are implicated in the prolonged low-frequency force depression (PLFFD) observed in skeletal muscle after both metabolically and mechanically demanding exercise. Metabolically demanding high-intensity exercise can induce PLFFD accompanied by ROS-dependent fragmentation of the sarcoplasmic reticulum Ca2+ release channels, the ryanodine receptor 1s (RyR1s). We tested whether similar changes occur after mechanically demanding eccentric contractions. Human subjects performed 100 repeated drop jumps, which require eccentric knee extensor contractions upon landing. This exercise caused a major PLFFD, such that maximum voluntary and electrically evoked forces did not recover within 24 h. Drop jumps induced only minor signs of increased ROS, and RyR1 fragmentation was observed in only 3 of 7 elderly subjects. Also, isolated mouse muscle preparations exposed to drop-jump-mimicking eccentric contractions showed neither signs of increased ROS nor RyR1 fragmentation. Still, the free cytosolic [Ca2+] during tetanic contractions was decreased by â¼15% 1 h after contractions, which can explain the exaggerated force decrease at low-stimulation frequencies but not the major frequency-independent force depression. In conclusion, PLFFD caused by mechanically demanding eccentric contractions does not involve any major increase in ROS or RyR1 fragmentation.-Kamandulis, S., de Souza Leite, F., Hernandez, A., Katz, A., Brazaitis, M., Bruton, J. D., Venckunas, T., Masiulis, N., Mickeviciene, D., Eimantas, N., Subocius, A., Rassier, D. E., Skurvydas, A., Ivarsson, N., Westerblad, H. Prolonged force depression after mechanically demanding contractions is largely independent of Ca2+ and reactive oxygen species.
Asunto(s)
Calcio/metabolismo , Contracción Muscular/fisiología , Fuerza Muscular/fisiología , Músculo Esquelético/fisiología , Especies Reactivas de Oxígeno/metabolismo , Adulto , Animales , Humanos , Masculino , Ratones , Canal Liberador de Calcio Receptor de Rianodina/metabolismoRESUMEN
High-intensity interval training (HIIT) is a time-efficient way of improving physical performance in healthy subjects and in patients with common chronic diseases, but less so in elite endurance athletes. The mechanisms underlying the effectiveness of HIIT are uncertain. Here, recreationally active human subjects performed highly demanding HIIT consisting of 30-s bouts of all-out cycling with 4-min rest in between bouts (≤3 min total exercise time). Skeletal muscle biopsies taken 24 h after the HIIT exercise showed an extensive fragmentation of the sarcoplasmic reticulum (SR) Ca(2+) release channel, the ryanodine receptor type 1 (RyR1). The HIIT exercise also caused a prolonged force depression and triggered major changes in the expression of genes related to endurance exercise. Subsequent experiments on elite endurance athletes performing the same HIIT exercise showed no RyR1 fragmentation or prolonged changes in the expression of endurance-related genes. Finally, mechanistic experiments performed on isolated mouse muscles exposed to HIIT-mimicking stimulation showed reactive oxygen/nitrogen species (ROS)-dependent RyR1 fragmentation, calpain activation, increased SR Ca(2+) leak at rest, and depressed force production due to impaired SR Ca(2+) release upon stimulation. In conclusion, HIIT exercise induces a ROS-dependent RyR1 fragmentation in muscles of recreationally active subjects, and the resulting changes in muscle fiber Ca(2+)-handling trigger muscular adaptations. However, the same HIIT exercise does not cause RyR1 fragmentation in muscles of elite endurance athletes, which may explain why HIIT is less effective in this group.
Asunto(s)
Calcio/metabolismo , Ejercicio Físico/fisiología , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , Retículo Sarcoplasmático/metabolismo , Adulto , Animales , Atletas , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Fibras Musculares Esqueléticas/fisiología , Resistencia Física , Especies Reactivas de Oxígeno/metabolismo , RecreaciónRESUMEN
KEY POINTS: We report that the small molecule CK-2066260 selectively slows the off-rate of Ca2+ from fast skeletal muscle troponin, leading to increased myofibrillar Ca2+ sensitivity in fast skeletal muscle. Rodents dosed with CK-2066260 show increased hindlimb muscle force and power in response to submaximal rates of nerve stimulation in situ. CK-2066260 has no effect on free cytosolic [Ca2+ ] during contractions of isolated muscle fibres. We conclude that fast skeletal muscle troponin sensitizers constitute a potential therapy to address an unmet need of improving muscle function in conditions of weakness and premature muscle fatigue. ABSTRACT: Skeletal muscle dysfunction occurs in many diseases and can lead to muscle weakness and premature muscle fatigue. Here we show that the fast skeletal troponin activator, CK-2066260, counteracts muscle weakness by increasing troponin Ca2+ affinity, thereby increasing myofibrillar Ca2+ sensitivity. Exposure to CK-2066260 resulted in a concentration-dependent increase in the Ca2+ sensitivity of ATPase activity in isolated myofibrils and reconstituted hybrid sarcomeres containing fast skeletal muscle troponin C. Stopped-flow experiments revealed a â¼2.7-fold decrease in the Ca2+ off-rate of isolated troponin complexes in the presence of CK-2066260 (6 vs. 17 s-1 under control conditions). Isolated mouse flexor digitorum brevis fibres showed a rapidly developing, reversible and concentration-dependent force increase at submaximal stimulation frequencies. This force increase was not accompanied by any changes in the free cytosolic [Ca2+ ] or its kinetics. CK-2066260 induced a slowing of relaxation, which was markedly larger at 26°C than at 31°C and could be linked to the decreased Ca2+ off-rate of troponin C. Rats dosed with CK-2066260 showed increased hindlimb isometric and isokinetic force in response to submaximal rates of nerve stimulation in situ producing significantly higher absolute forces at low isokinetic velocities, whereas there was no difference in force at the highest velocities. Overall muscle power was increased and the findings are consistent with a lack of effect on crossbridge kinetics. In conclusion, CK-2066260 acts as a fast skeletal troponin activator that may be used to increase muscle force and power in conditions of muscle weakness.