Metabolomic responses to high-intensity interval exercise in equine skeletal muscle: effects of rest interval duration.

High-intensity interval training has attracted considerable attention as a time-efficient strategy for inducing physiological adaptations, but the underlying mechanisms have yet to be elucidated. By using metabolomics techniques, we investigated changes in the metabolic network responses in Thoroughbred horses to high-intensity interval exercise performed with two distinct (15 min or 2 min) rest intervals. The peak plasma lactate level was higher during high-intensity exercise with a 2 min rest duration than that with a 15 min rest duration (24.5±6.8 versus 13.3±2.7 mmol l-1). The arterial oxygen saturation was lower at the end of all exercise sessions with a 2 min rest duration than that with a 15 min rest duration. Metabolomic analysis of skeletal muscle revealed marked changes in metabolite concentrations in the first and third bouts of the 15 min rest interval conditions. In contrast, there were no metabolite concentrations or pathways that significantly changed during the third bout of exercise performed with a 2 min rest interval. Our findings suggest that the activity of each energy production system is not necessarily reflected by apparent changes in metabolite concentrations, potentially due in part to a better match between metabolite flux into and out of the pathway and cycle, as well as between metabolite production and disposal. This study provides evidence that changes in metabolite concentrations vary greatly depending on the number of repetitions and the length of rest periods between exercises, even if the exercises themselves are identical.

Identifying physiological determinants of 800 m running performance using post-exercise blood lactate kinetics.

PURPOSE: The aims of the present study were to investigate blood lactate kinetics following high intensity exercise and identify the physiological determinants of 800 m running performance. METHODS: Fourteen competitive 800 m runners performed two running tests. First, participants performed a multistage graded exercise test to determine physiological indicators related to endurance performance. Second, participants performed four to six 30-s high intensity running bouts to determine post-exercise blood lactate kinetics. Using a biexponential time function, lactate exchange ability (γ1), lactate removal ability (γ2), and the quantity of lactate accumulated (QLaA) were calculated from individual blood lactate recovery data. RESULTS: 800 m running performance was significantly correlated with peak oxygen consumption (r = -0.794), γ1 and γ2 at 800 m race pace (r = -0.604 and -0.845, respectively), and QLaA at maximal running speed (r = -0.657). V ˙ O2peak and γ2 at 800 m race pace explained 83% of the variance in 800 m running performance. CONCLUSION: Our results indicate that (1) a high capacity to exchange and remove lactate, (2) a high capacity for short-term lactate accumulation and, (3) peak oxygen consumption, are critical elements of 800 m running performance. Accordingly, while lactate has primarily been utilized as a performance indicator for long-distance running, post-exercise lactate kinetics may also prove valuable as a performance determinant in middle-distance running.

Branched-chain amino acid supplementation suppresses the detraining-induced reduction of mitochondrial content in mouse skeletal muscle.

Exercise training enhances oxidative capacity whereas detraining reduces mitochondrial content in skeletal muscle. The strategy to suppress the detraining-induced reduction of mitochondrial content has not been fully elucidated. As previous studies reported that branched-chain amino acid (BCAA) ingestion increased mitochondrial content in skeletal muscle, we evaluated whether BCAA supplementation could suppress the detraining-induced reduction of mitochondrial content. Six-week-old male Institute of Cancer Research (ICR) mice were randomly divided into four groups as follows: control (Con), endurance training (Tr), detraining (DeTr), and detraining with BCAA supplementation (DeTr + BCAA). Mice in Tr, DeTr, and DeTr + BCAA performed treadmill running exercises [20-30 m/min, 60 min, 5 times/week, 4 weeks]. Then, mice in DeTr and DeTr + BCAA were administered with water or BCAA [0.6 mg/g of body weight, twice daily] for 2 weeks of detraining. In whole skeletal muscle, mitochondrial enzyme activities and protein content were decreased after 2 weeks of detraining, but the reduction was suppressed by BCAA supplementation. Peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) protein content, a master regulator of mitochondrial biogenesis, was decreased by detraining irrespective of BCAA ingestion. Regarding mitochondrial degradation, BCL2/adenovirus E1B 19 kDa protein-interacting protein 3 (BNIP3), a mitophagy-related protein, was significantly higher in the Tr group than in the DeTr + BCAA group, but not different from in the DeTr group. With respect to mitochondrial quality, BCAA ingestion did not affect oxygen consumption rate (OCR) and reactive oxygen species (ROS) production in isolated mitochondria. Our findings suggest that BCAA ingestion suppresses the detraining-induced reduction of mitochondrial content partly through inhibiting mitophagy.

Daily heat stress treatment rescues denervation-activated mitochondrial clearance and atrophy in skeletal muscle.

KEY POINTS: Traumatic nerve injury or nerve disease leads to denervation and severe muscle atrophy. Recent evidence shows that mitochondrial loss could be a key mediator of skeletal muscle atrophy. Here, we show that daily heat stress treatment rescues denervation-induced loss of mitochondria and concomitant muscle atrophy. We also found that denervation-activated autophagy-dependent mitochondrial clearance (mitophagy) was suppressed by daily heat stress treatment. The molecular basis of this observation is explained by our results showing that heat stress treatment attenuates the increase of key proteins that regulate the tagging step for mitochondrial clearance and the intermediate step of autophagosome formation in denervated muscle. These findings contribute to the better understanding of mitochondrial quality control in denervated muscle from a translational perspective and provide a mechanism behind the attenuation of muscle wasting by heat stress. ABSTRACT: Traumatic nerve injury or motor neuron disease leads to denervation and severe muscle atrophy. Recent evidence indicates that loss of mitochondria and the related reduction in oxidative capacity could be key mediators of skeletal muscle atrophy. As our previous study showed that heat stress increased the numbers of mitochondria in skeletal muscle, we evaluated whether heat stress treatment could have a beneficial impact on denervation-induced loss of mitochondria and subsequent muscle atrophy. Here, we report that daily heat stress treatment (mice placed in a chamber with a hot environment; 40°C, 30 min day(-1) , for 7 days) rescues the following parameters: (i) muscle atrophy (decreased gastrocnemius muscle mass); (ii) loss of mitochondrial content (decreased levels of ubiquinol-cytochrome c reductase core protein II, cytochrome c oxidase subunits I and IV and voltage-dependent anion channel protein); and (iii) reduction in oxidative capacity (reduced maximal activities of citrate synthase and 3-hydroxyacyl-CoA dehydrogenase) in denervated muscle (produced by unilateral sciatic nerve transection). In order to gain a better understanding of the above mitochondrial adaptations, we also examined the effects of heat stress on autophagy-dependent mitochondrial clearance (mitophagy). Daily heat stress normalized denervation-activated induction of mitophagy (increased mitochondrial microtubule-associated protein 1A/1B-light chain3-II (LC3-II) with and without blocker of autophagosome clearance). The molecular basis of this observation was explained by the results that heat stress attenuated the denervation-induced increase in key proteins that regulate the following steps: (i) the tagging step of mitochondrial clearance (increased mitochondrial Parkin, ubiquitin-conjugated, P62/sequestosome 1 (P62/SQSTM1)); and (ii) the elongation step of autophagosome formation (increased Atg5-Atg12 conjugate and Atg16L). Overall, our results contribute to the better understanding of mitochondrial quality control and the mechanisms behind the attenuation of muscle wasting by heat stress in denervated skeletal muscle.

Postexercise whole body heat stress additively enhances endurance training-induced mitochondrial adaptations in mouse skeletal muscle.

A recent study demonstrated that heat stress induces mitochondrial biogenesis in C2C12 myotubes, thereby implying that heat stress may be an effective treatment to enhance endurance training-induced mitochondrial adaptations in skeletal muscle. However, whether heat stress actually induces mitochondrial adaptations in skeletal muscle in vivo is unclear. In the present study, we report the novel findings that 1) whole body heat stress produced by exposure of ICR mice to a hot environment (40°C, 30 min/day, 5 days/wk, 3 wk) induced mitochondrial adaptations such as increased mitochondrial enzyme activity (citrate synthase and 3-hydroxyacyl CoA dehydrogenase) and respiratory chain protein content (complexes I-V) in skeletal muscle in vivo and 2) postexercise whole body heat stress additively enhanced endurance training-induced mitochondrial adaptations (treadmill running, 25 m/min, 30 min/day, 5 days/wk, 3 wk). Moreover, to determine the candidate mechanisms underlying mitochondrial adaptations, we investigated the acute effects of postexercise whole body heat stress on the phosphorylation status of cellular signaling cascades that subsequently induce mitochondrial gene transcription. We found that whole body heat stress boosted the endurance exercise-induced phosphorylation of p38 MAPK, increased the phosphorylation status of p70S6K, a biomarker of mammalian target of rapamycin complex 1 activity, and unexpectedly dephosphorylated AMP-activated protein kinase and its downstream target acetyl-CoA carboxylase in skeletal muscle. Our present observations suggest that heat stress can act as an effective postexercise treatment. Heat stress treatment appeared to be clinically beneficial for people who have difficulty participating in sufficient exercise training, such as the elderly, injured athletes, and patients.

Better maintenance of enzymatic capacity and higher levels of substrate transporter proteins in skeletal muscle of aging female mice.

This study investigated sex-specific differences in high-energy phosphate, glycolytic, and mitochondrial enzyme activities and also metabolite transporter protein levels in the skeletal muscles of adult (5 months old), middle-aged (12 months old), and advanced-aged (24 months old) mice. While gastrocnemius glycogen content increased with age regardless of sex, gastrocnemius triglyceride levels increased only in advanced-aged female mice. Aging decreased creatine kinase and adenylate kinase activities in the plantaris muscle of both sexes and in the soleus muscle of male mice but not in female mice. Irrespective of sex, phosphofructokinase and lactate dehydrogenase (LDH) activities decreased in the plantaris and soleus muscles. Additionally, hexokinase activity in the plantaris muscle and LDH activity in the soleus muscle decreased to a greater extent in aged male mice compared with those in aged female mice. Mitochondrial enzyme activities increased in the plantaris muscle of aged female mice but did not change in male mice. The protein content of the glucose transporter 4 in the aged plantaris muscle and fatty acid translocase/cluster of differentiation 36 increased in the aged plantaris and soleus muscles of both sexes, with a significantly higher content in female mice. These findings suggest that females possess a better ability to maintain metabolic enzyme activity and higher levels of metabolite transport proteins in skeletal muscle during aging, despite alterations in lipid metabolism. Our data provide a basis for studying muscle metabolism in the context of age-dependent metabolic perturbations and diseases that affect females and males differently.

Effects of endurance training under calorie restriction on energy substrate metabolism in mouse skeletal muscle and liver.

We investigated whether calorie restriction (CR) enhances metabolic adaptations to endurance training (ET). Ten-week-old male Institute of Cancer Research (ICR) mice were fed ad libitum or subjected to 30% CR. The mice were subdivided into sedentary and ET groups. The ET group performed treadmill running (20-25 m/min, 30 min, 5 days/week) for 5 weeks. We found that CR decreased glycolytic enzyme activity and monocarboxylate transporter (MCT) 4 protein content, while enhancing glucose transporter 4 protein content in the plantaris and soleus muscles. Although ET and CR individually increased citrate synthase activity in the plantaris muscle, the ET-induced increase in respiratory chain complex I protein content was counteracted by CR. In the soleus muscle, mitochondrial enzyme activity and protein levels were increased by ET, but decreased by CR. It has been suggested that CR partially interferes with skeletal muscle adaptation to ET.

Inter-individual variability in peripheral oxygen saturation and repeated sprint performance in hypoxia: an observational study of highly-trained subjects.

Individual variations in peripheral oxygen saturation (SpO2) during repeated sprints in hypoxia and their impact on exercise performance remain unclear despite fixed external hypoxic stimuli (inspired oxygen fraction: FiO2). This study examined SpO2 individual variations during repeated sprints in hypoxia and their impact on exercise performance. Thirteen highly-trained sprint runners performed 10 × 10-s cycle sprints with 30-s passive recoveries in normobaric hypoxia (FiO2: 0.150). Mean power output (MPO), post-sprint SpO2, and heart rate for each sprint were assessed. Sprint decrement score (Sdec), evaluating fatigue development, was calculated using MPO variables. Participants were categorized into a high saturation group (HiSat, n = 7) or a low saturation group (LowSat, n = 6) based on their mean post-sprint SpO2 (measured 10-15 s after each sprint). Individual mean post-sprint SpO2 ranged from 91.6% to 82.2%. Mean post-sprint SpO2 was significantly higher (P < 0.001, d = 1.54) in HiSat (89.1% ± 1.5%) than LowSat (84.7% ± 1.6%). A significantly larger decrease in Sdec (P = 0.008, d = 1.68) occurred in LowSat (-22.3% ± 2.3%) compared to HiSat (-17.9% ± 2.5%). MPO (P = 0.342 d = 0.55) and heart rate (P = 0.225 d = 0.67) did not differ between groups. There was a significant correlation (r = 0.61; P = 0.028) between SpO2 and Sdec. In highly-trained sprint runners, individual responses to hypoxia varied widely and significantly affected repeated sprint ability, with greater decreases in SpO2 associated with larger performance alterations (i.e., larger decrease in Sdec).

Differential patterns of sweat and blood lactate concentration response during incremental exercise in varied ambient temperatures: A pilot study.

Blood lactate concentration during exercise is a reliable indicator of energy metabolism and endurance performance. Lactate is also present in sweat, and sweating plays an important role in thermoregulation, especially in hot conditions. Recently, wearable sensors have enabled the real-time and noninvasive measurement of sweat lactate concentration, potentially serving as an alternative indicator of blood lactate response. However, the evidence regarding the relationship between sweat and blood lactate responses during incremental exercise in hot conditions is lacking. In a randomized cross-over design, six highly trained male runners completed two incremental treadmill tests under normal (20°C/50%RH) or hot (30°C/50%RH) conditions. The tests include 3-min running stages and 1-min recovery, starting at 12 km/h and increasing by 1 km/h at each stage. Blood and sweat lactate concentrations were measured at each stage to determine blood and sweat lactate thresholds (LT). Blood lactate concentrations were higher under hot conditions (p < 0.01), but there was no difference in the response pattern or velocity at blood LT between conditions. Significant early increase (p < 0.01) in sweat lactate and low velocity at sweat LT (p < 0.05) were observed under hot conditions. A significant correlation between blood and sweat lactate concentrations was found under normal conditions (p < 0.001) but not under hot conditions, and no significant correlations were observed between the velocity at blood and sweat LT. In conclusion, sweat lactate concentration does not consistently reflect blood lactate concentration during incremental exercise.

Effects of pacing strategy on metabolic responses to 2-min intense exercise in Thoroughbred horses.

Evidence suggests that positive pacing strategy improves exercise performance and fatigue tolerance in athletic events lasting 1-5 min. This study investigated muscle metabolic responses to positive and negative pacing strategies in Thoroughbred horses. Eight Thoroughbred horses performed 2 min treadmill running using positive (1 min at 110% maximal O2 uptake [V̇O2max], followed by 1 min at 90% V̇O2max) and negative (1 min at 90% V̇O2max, followed by 1 min at 110% V̇O2max) pacing strategies. The arterial-mixed venous O2 difference did not significantly differ between the two strategies. Plasma lactate levels increased toward 2 min, with significantly higher concentrations during positive pacing than during negative pacing. Muscle glycogen level was significantly lower at 1 and 2 min of positive pacing than those of negative pacing. Metabolomic analysis showed that the sum of glycolytic intermediates increased during the first half of positive pacing and the second half of negative pacing. Regardless of pacing strategy, the sum of tricarboxylic acid cycle metabolites increased during the first half but remained unchanged thereafter. Our data suggest that positive pacing strategy is likely to activate glycolytic metabolism to a greater extent compared to negative pacing, even though the total workload is identical.

Sex-specific differences in the metabolic enzyme activity and transporter levels in mouse skeletal muscle during postnatal development.

Although sex-associated differences in energy metabolism in adults are well-characterized, developmental sex-specific changes in skeletal muscle metabolism are largely unknown. This study investigated sex differences in high-energy phosphate, glycolytic, and mitochondrial enzyme activities and metabolite transporter protein levels in mouse skeletal muscles during the early postnatal period (day 10), post-weaning (day 28), sexual maturity (day 56), and adult life (day 140). No significant sex-specific differences were observed on days 10 and 28, except for glucose transporter (GLUT) 4 level. The hexokinase, phosphofructokinase, and lactate dehydrogenase activities of skeletal muscle were higher and the citrate synthase, cytochrome c oxidase, and ß-hydroxyacyl-CoA dehydrogenase activities were lower in female mice than those in male mice on days 56 and 140. The GLUT4 and FAT/CD36 protein levels were higher and the monocarboxylate transporter 4 level was lower in the skeletal muscles of female mice than those of male mice, particularly on days 56 and 140. At 140 days of age, the respiratory exchange ratio during treadmill running (15 m/min, 60 min) was lower in females than that in males, despite no sex differences at rest. In summary, sex differences were not evident in the early postnatal and post-weaning periods but became apparent after the mice reached sexual maturity. These findings indicate that sexually mature animals are a better model for investigating sex differences, particularly in the context of studying energy metabolism in mice.

Effects of Endurance Training on Metabolic Enzyme Activity and Transporter Proteins in Skeletal Muscle of Ovariectomized Mice.

PURPOSE: Estrogen deficiency or insufficiency can occur under several conditions, leading to negative health outcomes. To establish an effective countermeasure against estrogen loss, we investigated the effects of endurance training on ovariectomy (OVX)-induced metabolic disturbances. METHODS: Female Institute of Cancer Research mice underwent OVX or sham operations. On day 7 of recovery, the mice were randomized to remain either sedentary or undergo 5 wk of treadmill running (15-20 m·min -1 , 60 min, 5 d·wk -1 ). During week 5 of the training, all animals performed a treadmill running test (15 m·min -1 , 60 min). RESULTS: After the experimental period, OVX resulted in greater gains in body mass, fat mass, and triglyceride content in the gastrocnemius muscle. OVX enhanced phosphofructokinase activity in the plantaris muscle and decreased lactate dehydrogenase activity in the plantaris and soleus muscles. OVX decreased the protein content of NDUFB8, a mitochondrial respiratory chain subunit, but did not decrease other mitochondrial proteins or enzyme activities. Endurance training significantly enhanced mitochondrial enzyme activity and protein content in the skeletal muscles. Although OVX increased the respiratory exchange ratio during the treadmill running test, and postexercise blood lactate levels, endurance training normalized these parameters. CONCLUSIONS: The present findings suggest that endurance training is a viable strategy to counteract the negative metabolic consequences in hypoestrogenism.

Stride-to-stride variability and fluctuations at intensities around lactate threshold in distance runners.

Stride-to-stride variability and fluctuations in running have been widely investigated in relation to fatigue, injury, and other factors. However, no studies have examined the relationship of stride-to-stride variability and fluctuations with lactate threshold (LT), a well-known performance indicator for distance runners that represents the threshold at which fast-twitch muscle fibers are activated and the glycolytic system is hyperactivated. In this study, we examined a relationship between LT and stride-to-stride variability and fluctuations in trained middle- and long-distance runners (n = 33). All runners were asked to perform multistage graded exercise tests while wearing accelerometers on the upper surface of their shoes. The LT was determined by measuring blood lactate concentrations after each stage. Three gait parameters for each step were calculated based on the acceleration data: stride time (ST), ground contact time (CT), and peak acceleration (PA). The coefficient of variation (CV) and the long-range correlations (α) for each parameter were also calculated. The effects of the runner's group and the relative intensity for CV and α on gait parameters were evaluated using a two-way repeated measures analysis of variance. Although no significant effect was observed in the CV and α of ST, significant intensity main effects were observed for the CV and α of CT and PA. The lack of significant changes in ST might be the result of runners' adequate control of ST to minimize energy cost. All the parameters showing significant changes with increasing intensity decreased dramatically when they were close to LT. This might have been caused by an increase in physiological load near LT and be interpreted as a variation in motor control because of alternations in the mobilized muscle fibers and physiological changes around the LT. The α should be useful for non-invasive LT detection.

Clenbuterol, a ß2-adrenergic agonist, reciprocally alters PGC-1 alpha and RIP140 and reduces fatty acid and pyruvate oxidation in rat skeletal muscle.

Clenbuterol, a ß2-adrenergic agonist, reduces mitochondrial content and enzyme activities in skeletal muscle, but the mechanism involved has yet to be identified. We examined whether clenbuterol-induced changes in the muscles' metabolic profile and the intrinsic capacity of mitochondria to oxidize substrates are associated with reductions in the nuclear receptor coactivator PGC-1 alpha and/or an increase in the nuclear corepressor RIP140. In rats, clenbuterol was provided in the drinking water (30 mg/l). In 3 wk, this increased body (8%) and muscle weights (12-17%). In red (R) and white (W) muscles, clenbuterol induced reductions in mitochondrial content (citrate synthase: R, 27%; W, 52%; cytochrome-c oxidase: R, 24%; W, 34%), proteins involved in fatty acid transport (fatty acid translocase/CD36: R, 36%; W, 35%) and oxidation [ß-hydroxyacyl CoA dehydrogenase (ß-HAD): R, 33%; W, 62%], glucose transport (GLUT4: R, 8%; W, 13%), lactate transport monocarboxylate transporter (MCT1: R, 61%; W, 37%), and pyruvate oxidation (PDHE1α, R, 18%; W, 12%). Concurrently, only red muscle lactate dehydrogenase activity (25%) and MCT4 (31%) were increased. Palmitate oxidation was reduced in subsarcolemmal (SS) (R, 30%; W, 52%) and intermyofibrillar (IMF) mitochondria (R, 17%; W, 44%) along with reductions in ß-HAD activity (SS: R, 17%; W, 51%; IMF: R, 20%; W, 57%). Pyruvate oxidation was only reduced in SS mitochondria (R, 20%; W, 28%), but this was not attributable solely to PDHE1α, which was reduced in both SS (R, 21%; W, 20%) and IMF mitochondria (R, 15%; W, 43%). These extensive metabolic changes induced by clenbuterol were associated with reductions in PGC-1α (R, 37%; W, 32%) and increases in RIP140 (R, 23%; W, 21%). This is the first evidence that clenbuterol appears to exert its metabolic effects via simultaneous and reciprocal changes in the nuclear receptor coactivator PGC-1α and the nuclear corepressor RIP140.

Inhibition of monocarboxylate transporters (MCT) 1 and 4 reduces exercise capacity in mice.

The concept of lactate shuttle is widely accepted in exercise physiology. Lactate transport is mediated by monocarboxylate transporters (MCT), which enable cells to take up and release lactate. However, the role of lactate during exercise has not yet been fully elucidated. In this study, we investigated the effects of lactate transport inhibition on exercise capacity and metabolism in mice. Here, we demonstrated that MCT1 inhibition by α-cyano-4-hydroxycinnamate administration (4-CIN, 200 mg/g of body weight) reduced the treadmill running duration at 20 m/min. The administration of 4-CIN increased the blood lactate concentration immediately after exercise. With matched exercise duration, the muscle lactate concentration was higher while muscle glycogen content was lower in 4-CIN-administered mice. Further, we showed that MCT4 inhibition by bindarit administration (50 mg/kg of body weight) reduced the treadmill running duration at 40 m/min. Bindarit administration increased the muscle lactate but did not alter the blood lactate and glucose concentrations, as well as muscle glycogen content, immediately after exercise. A negative correlation was observed between exercise duration at 40 m/min and muscle lactate concentration immediately after exercise. Our results suggest that lactate transport via MCT1 and MCT4 plays a pivotal role in sustaining exercise.

Effects of endurance training on metabolic enzyme activity and transporter protein levels in the skeletal muscles of orchiectomized mice.

This study investigated whether endurance training attenuates orchiectomy (ORX)-induced metabolic alterations. At 7 days of recovery after sham operation or ORX surgery, the mice were randomized to remain sedentary or undergo 5 weeks of treadmill running training (15-20 m/min, 60 min, 5 days/week). ORX decreased glycogen concentration in the gastrocnemius muscle, enhanced phosphofructokinase activity in the plantaris muscle, and decreased lactate dehydrogenase activity in the plantaris and soleus muscles. Mitochondrial enzyme activities and protein content in the plantaris and soleus muscles were also decreased after ORX, but preserved, in part, by endurance training. In the treadmill running test (15 m/min, 60 min) after 4 weeks of training, orchiectomized sedentary mice showed impaired exercise performance, which was restored by endurance training. Thus, endurance training could be a potential therapeutic strategy to prevent the hypoandrogenism-induced decline in muscle mitochondrial content and physical performance.

Effects of Lactate Administration on Mitochondrial Respiratory Function in Mouse Skeletal Muscle.

Recent evidence has shown that mitochondrial respiratory function contributes to exercise performance and metabolic health. Given that lactate is considered a potential signaling molecule that induces mitochondrial adaptations, we tested the hypothesis that lactate would change mitochondrial respiratory function in skeletal muscle. Male ICR mice (8 weeks old) received intraperitoneal injection of PBS or sodium lactate (1 g/kg BW) 5 days a week for 4 weeks. Mitochondria were isolated from freshly excised gastrocnemius muscle using differential centrifugation and were used for all analyses. Lactate administration significantly enhanced pyruvate + malate- and glutamate + malate-induced (complex I-driven) state 3 (maximal/ATP synthesis-coupled) respiration, but not state 2 (basal/proton conductance) respiration. In contrast, lactate administration significantly decreased succinate + rotenone-induced (complex II-driven) state 3 and 2 respiration. No significant differences were observed in malate + octanoyl-l-carnitine-induced state 3 or 2 respiration. The enzymatic activity of complex I was tended to increase and those of complexes I + III and IV were significantly increased after lactate administration. No differences were observed in the activities of complexes II or II + III. Moreover, lactate administration increased the protein content of NDUFS4, a subunit of complex I, but not those of the other components. The present findings suggest that lactate alters mitochondrial respiratory function in skeletal muscle.

Effect of training and detraining on monocarboxylate transporter (MCT) 1 and MCT4 in Thoroughbred horses.

The aim of this study was to investigate the effects of training and detraining on the monocarboxylate transporter (MCT) 1 and MCT4 levels in the gluteus medius muscle of Thoroughbred horses. Twelve Thoroughbred horses were used for the analysis. For 18 weeks, all the horses underwent high-intensity training (HIT), with running at 90-110% maximal oxygen consumption (VO2 max ) for 3 min, 5 days week(-1). Thereafter, the horses either underwent detraining for 6 weeks by either 3 min of moderate-intensity training (MIT) at 70% VO2 max, 5 days week(-1) (HIT-MIT group) or stall rest (HIT-SR group). The horses underwent an incremental exercise test, VO2 max was measured and resting muscle samples were obtained from the middle gluteus muscle at 0, 18 and 24 weeks. The content of MCT1 and MCT4 proteins increased after 18 weeks of HIT. At the end of this period, an increase was noted in the citrate synthase activity, while phosphofructokinase activity remained unchanged. After 6 weeks of detraining, all these indexes returned to the pretraining levels in the HIT-SR group. However, in the HIT-MIT group, the increase in the MCT1 protein content and citrate synthase activity was maintained after 6 weeks of MIT, while the MCT4 protein content decreased to the pretraining value. These results suggest that the content of MCT1 and MCT4 proteins increases after HIT in Thoroughbred horses. In addition, the increase in the MCT1 protein content and oxidative capacity induced by HIT can be maintained by MIT of 70% VO2 max, but the increase in the MCT4 protein content cannot be maintained by MIT.

Performance, Metabolic, and Neuromuscular Consequences of Repeated Wingates in Hypoxia and Normoxia: A Pilot Study.

BACKGROUND: Compared with normoxia, repeated short (5-10 s) sprints (>10 efforts) with incomplete recovery (≤30 s) in hypoxia likely cause substantial performance reduction accompanied by larger metabolic disturbances and magnitude of neuromuscular fatigue. However, the effects of hypoxia on performance of repeated long (30 s) "all-out" efforts with near complete recovery (4.5 min) and resulting metabolic and neuromuscular adjustments remain unclear. PURPOSE: The intention was to compare acute performance, metabolic, and neuromuscular responses across repeated Wingates between hypoxia and normoxia. METHODS: On separate visits, 6 male participants performed 4 × 30-second Wingate efforts with 4.5-minute recovery in either hypoxia (fraction of inspired oxygen: 0.145) or normoxia. Responses to exercise (muscle and arterial oxygenation trends, heart rate, and blood lactate concentration) and the integrity of neuromuscular function in the knee extensors were assessed for each exercise bout. RESULTS: Mean (P = .80) and peak (P = .92) power outputs, muscle oxygenation (P = .88), blood lactate concentration (P = .72), and perceptual responses (all Ps > .05) were not different between conditions. Arterial oxygen saturation was significantly lower, and heart rate higher, in hypoxia versus normoxia (P < .001). Maximal voluntary contraction force and peripheral fatigue indices (peak twitch force and doublets at low and high frequencies) decreased across efforts (all Ps < .001) irrespective of conditions (all Ps > .05). CONCLUSION: Despite heightened arterial hypoxemia and cardiovascular solicitation, hypoxic exposure during 4 repeated 30-second Wingate efforts had no effect on performance and accompanying metabolic and neuromuscular adjustments.

Lactate administration does not affect denervation-induced loss of mitochondrial content and muscle mass in mice.

Lactate is considered to be a signaling molecule that induces mitochondrial adaptation and muscle hypertrophy. The purpose of this study was to examine whether lactate administration attenuates denervation-induced loss of mitochondrial content and muscle mass. Eight-week-old male Institute of Cancer Research mice underwent unilateral sciatic nerve transection surgery. The contralateral hindlimb served as a sham-operated control. From the day of surgery, mice were injected intraperitoneally with PBS or sodium lactate (equivalent to 1 g·kg-1 body weight) once daily for 9 days. After 10 days of denervation, gastrocnemius muscle weight decreased to a similar extent in both the PBS- and lactate-injected groups. Denervation significantly decreased mitochondrial enzyme activity, protein content, and MCT4 protein content in the gastrocnemius muscle. However, lactate administration did not have any significant effects. The current observations suggest that daily lactate administration for 9 days does not affect denervation-induced loss of mitochondrial content and muscle mass.