The Rate of Leg Fat Oxidation Is Not Attenuated During Incremental Intensity One-Leg Knee Extensor Exercise.

It is not clear if fat oxidation is attenuated at higher exercise intensities, when exercising with a small muscle mass, and therefore, we studied leg fat oxidation during graded one-leg exercise. Ten males (age: 27 ± 2 years, body mass: 82 ± 3 kg, BMI: 24 ± 1 kg m-2, V̇O2max: 49 ± 2 mL min-1 kg-1) performed one-leg exercise at 25% of maximal workload (Wmax) for 30 min, followed by 120-min exercise at 55% Wmax with the contralateral leg, and finally 30-min exercise at 85% Wmax with the first leg. Blood was sampled from an artery and both femoral veins, and blood flow was determined using Doppler ultrasound. Muscle biopsies were obtained before and after 30 min at each workload. One-way RM ANOVA was applied to determine the impact of exercise intensity. Data are expressed as mean ± SEM. From rest through exercise average blood flow (0.4 ± 0.1, 2.1 ± 0.1, 2.6 ± 0.2, 3.7 ± 0.2 L min-1) and oxygen uptake across the leg (0.03 ± 0.01, 0.23 ± 0.02, 0.35 ± 0.03, 0.53 ± 0.04 L min-1) increased with exercise intensity (p < 0.001). Leg RQ (0.76 ± 0.04, 0.86 ± 0.02,0.87 ± 0.01, 0.92 ± 0.01, p < 0.001), leg plasma FA uptake (2 ± 2, 46 ± 8,83 ± 9, 114 ± 16 µmol min-1; p < 0.001) and rate of leg fat oxidation (0.016 ± 0.005, 0.062 ± 0.012, 0.075 ± 0.011, 0.084 ± 0.018 g min-1, p < 0.007) increased with exercise intensity. Muscle-free carnitine content was unchanged from rest at 25% Wmax and decreased after 30 min exercise at 55% and 85% Wmax (17.4 ± 1.6, 16.6 ± 0.7, 14.5 ± 1.2, 10.5 ± 1.0 mmol/kg dry muscle, respectively; p < 0.006). During incremental one-leg exercise, the rate of leg fat oxidation was not attenuated with increasing exercise intensity, probably due to an insufficient muscle metabolic stress response.

Protein dose requirements to maximize skeletal muscle protein synthesis after repeated bouts of resistance exercise in young trained women.

Studies examining the effect of protein (PRO) feeding on post resistance exercise (RE) muscle protein synthesis (MPS) have primarily been performed in men, and little evidence is available regarding the quantity of PRO required to maximally stimulate MPS in trained women following repeated bouts of RE. We therefore quantified acute (4 h and 8 h) and extended (24 h) effects of two bouts of resistance exercise, alongside protein-feeding, in women, and the PRO requirement to maximize MPS. Twenty-four RE trained women (26.6 ± 0.7 years, mean ± SEM) performed two bouts of whole-body RE (3 × 8 repetitions/maneuver at 75% 1-repetition maximum) 4 h apart, with post-exercise ingestion of 15 g, 30 g, or 60 g whey PRO (n = 8/group). Saliva, venous blood, and a vastus lateralis muscle biopsy were taken at 0 h, 4 h, 8 h, and 24 h post-exercise. Plasma leucine and branched chain amino acids were quantified using gas chromatography mass spectrometry (GC-MS) after ingestion of D2 O. Fifteen grams PRO did not alter plasma leucine concentration or myofibrillar synthetic rate (MyoFSR). Thirty and sixty grams PRO increased plasma leucine concentration above baseline (105.5 ± 5.3 µM; 120.2 ± 7.4 µM, respectively) at 4 h (151.5 ± 8.2 µM, p < 0.01; 224.8 ± 16.0 µM, p < 0.001, respectively) and 8 h (176.0 ± 7.3 µM, p < 0.001; 281.7 ± 21.6 µM, p < 0.001, respectively). Ingestion of 30 g PRO increased MyoFSR above baseline (0.068 ± 0.005%/h) from 0 to 4 h (0.140 ± 0.021%/h, p < 0.05), 0 to 8 h (0.121 ± 0.012%/h, p < 0.001), and 0 to 24 h (0.099 ± 0.011%/h, p < 0.01). Ingestion of 60 g PRO increased MyoFSR above baseline (0.063 ± 0.003%/h) from 0 to 4 h (0.109 ± 0.011%/h, p < 0.01), 0 to 8 h (0.093 ± 0.008%/h, p < 0.01), and 0 to 24 h (0.086 ± 0.006%/h, p < 0.01). Post-exercise ingestion of 30 g or 60 g PRO, but not 15 g, acutely increased MyoFSR following two consecutive bouts of RE and extended the anabolic window over 24 h. There was no difference between the 30 g and 60 g responses.

Impacts of rat hindlimb Fndc5/irisin overexpression on muscle and adipose tissue metabolism.

Myokines, such as irisin, have been purported to exert physiological effects on skeletal muscle in an autocrine/paracrine fashion. In this study, we aimed to investigate the mechanistic role of in vivo fibronectin type III domain-containing 5 (Fndc5)/irisin upregulation in muscle. Overexpression (OE) of Fndc5 in rat hindlimb muscle was achieved by in vivo electrotransfer, i.e., bilateral injections of Fndc5 harboring vectors for OE rats (n = 8) and empty vector for control rats (n = 8). Seven days later, a bolus of D2O (7.2 mL/kg) was administered via oral gavage to quantify muscle protein synthesis. After an overnight fast, on day 9, 2-deoxy-d-glucose-6-phosphate (2-DG6P; 6 mg/kg) was provided during an intraperitoneal glucose tolerance test (2 g/kg) to assess glucose handling. Animals were euthanized, musculus tibialis cranialis muscles and subcutaneous fat (inguinal) were harvested, and metabolic and molecular effects were evaluated. Muscle Fndc5 mRNA increased with OE (~2-fold; P = 0.014), leading to increased circulating irisin (1.5 ± 0.9 to 3.5 ± 1.2 ng/mL; P = 0.049). OE had no effect on protein anabolism or mitochondrial biogenesis; however, muscle glycogen was increased, along with glycogen synthase 1 gene expression (P = 0.04 and 0.02, respectively). In addition to an increase in glycogen synthase activation in OE (P = 0.03), there was a tendency toward increased glucose transporter 4 protein (P = 0.09). However, glucose uptake (accumulation of 2-DG6P) was identical. Irisin elicited no endocrine effect on mitochondrial biogenesis or uncoupling proteins in white adipose tissue. Hindlimb overexpression led to physiological increases in Fndc5/irisin. However, our data indicate limited short-term impacts of irisin in relation to muscle anabolism, mitochondrial biogenesis, glucose uptake, or adipose remodeling.

Muscle thickness correlates to muscle cross-sectional area in the assessment of strength training-induced hypertrophy.

Muscle thickness (MT) measured by ultrasound has been used to estimate cross-sectional area (measured by CT and MRI) at a single time point. We tested whether MT could be used as a valid marker of MRI determined muscle anatomical cross-sectional area (ACSA) and volume changes following resistance training (RT). Nine healthy, young, male volunteers (24 ± 2 y.o., BMI 24.1 ± 2.8 kg/m2 ) had vastus lateralis (VL) muscle volume (VOL) and ACSAmid (at 50% of femur length, FL) assessed by MRI, and VL MT measured by ultrasound at 50% FL. Measurements were taken at baseline and after 12 weeks of isokinetic RT. Differences between baseline and post-training were assessed by Student's paired t test. The relationships between MRI and ultrasound measurements were tested by Pearson's correlation. After RT, MT increased by 7.5 ± 6.1% (P < .001), ACSAmid by 5.2 ± 5% (P < .001), and VOL by 5.0 ± 6.9% (P < .05) (values: means ± SD). Positive correlations were found, at baseline and 12 weeks, between MT and ACSAmid (r = .82, P < .001 and r = .73, P < .001, respectively), and between MT and VOL (r = .76, P < .001 and r = .73, P < .001, respectively). The % change in MT with training was correlated with % change in ACSAmid (r = .69, P < .01), but not % change in VOL (r = .33, P > .05). These data support evidence that MT is a reliable index of muscle ACSAmid and VOL at a single time point. MT changes following RT are associated with parallel changes in muscle ACSAmid but not with the changes in VOL, highlighting the impact of RT on regional hypertrophy.

A novel D2O tracer method to quantify RNA turnover as a biomarker of de novo ribosomal biogenesis, in vitro, in animal models, and in human skeletal muscle.

Current methods to quantify in vivo RNA dynamics are limited. Here, we developed a novel stable isotope (D2O) methodology to quantify RNA synthesis (i.e., ribosomal biogenesis) in cells, animal models, and humans. First, proliferating C2C12 cells were incubated in D2O-enriched media and myotubes ±50 ng/ml IGF-I. Second, rat quadriceps (untrained, n = 9; 7-wk interval-"like" training, n = 13) were collected after ~3-wk D2O (70 atom %) administration, with body-water enrichment monitored via blood sampling. Finally, 10 (23 ± 1 yr) men consumed 150-ml D2O followed by 50 ml/wk and undertook 6-wk resistance exercise (6 × 8 repetitions, 75% 1-repetition maximum 3/wk) with body-water enrichment monitored by saliva sampling and muscle biopsies (for determination of RNA synthesis) at 0, 3, and 6 wk. Ribose mole percent excess (r-MPE) from purine nucleotides was analyzed via GC-MS/MS. Proliferating C2C12 cell r-MPE exhibited a rise to plateau, whereas IGF-I increased myotube RNA from 76 ± 3 to 123 ± 3 ng/µl and r-MPE by 0.39 ± 0.1% (both P < 0.01). After 3 wk, rat quadriceps r-MPE had increased to 0.25 ± 0.01% (P < 0.01) and was greater with running exercise (0.36 ± 0.02%; P < 0.01). Human muscle r-MPE increased to 0.06 ± 0.01 and 0.13 ± 0.02% at 3/6 wk, respectively, equating to synthesis rates of ~0.8%/day, increasing with resistance exercise to 1.7 ± 0.3%/day (P < 0.01) and 1.2 ± 0.1%/day (P < 0.05) at 3/6 wk, respectively. Therefore, we have developed and physiologically validated a novel technique to explore ribosomal biogenesis in a multimodal fashion.

The effect of age and unilateral leg immobilization for 2 weeks on substrate utilization during moderate-intensity exercise in human skeletal muscle.

KEY POINTS: This study aimed to provide molecular insight into the differential effects of age and physical inactivity on the regulation of substrate metabolism during moderate-intensity exercise. Using the arteriovenous balance technique, we studied the effect of immobilization of one leg for 2 weeks on leg substrate utilization in young and older men during two-legged dynamic knee-extensor moderate-intensity exercise, as well as changes in key proteins in muscle metabolism before and after exercise. Age and immobilization did not affect relative carbohydrate and fat utilization during exercise, but the older men had higher uptake of exogenous fatty acids, whereas the young men relied more on endogenous fatty acids during exercise. Using a combined whole-leg and molecular approach, we provide evidence that both age and physical inactivity result in intramuscular lipid accumulation, but this occurs only in part through the same mechanisms. ABSTRACT: Age and inactivity have been associated with intramuscular triglyceride (IMTG) accumulation. Here, we attempt to disentangle these factors by studying the effect of 2 weeks of unilateral leg immobilization on substrate utilization across the legs during moderate-intensity exercise in young (n = 17; 23 ± 1 years old) and older men (n = 15; 68 ± 1 years old), while the contralateral leg served as the control. After immobilization, the participants performed two-legged isolated knee-extensor exercise at 20 ± 1 W (∼50% maximal work capacity) for 45 min with catheters inserted in the brachial artery and both femoral veins. Biopsy samples obtained from vastus lateralis muscles of both legs before and after exercise were used for analysis of substrates, protein content and enzyme activities. During exercise, leg substrate utilization (respiratory quotient) did not differ between groups or legs. Leg fatty acid uptake was greater in older than in young men, and although young men demonstrated net leg glycerol release during exercise, older men showed net glycerol uptake. At baseline, IMTG, muscle pyruvate dehydrogenase complex activity and the protein content of adipose triglyceride lipase, acetyl-CoA carboxylase 2 and AMP-activated protein kinase (AMPK)γ3 were higher in young than in older men. Furthermore, adipose triglyceride lipase, plasma membrane-associated fatty acid binding protein and AMPKγ3 subunit protein contents were lower and IMTG was higher in the immobilized than the contralateral leg in young and older men. Thus, immobilization and age did not affect substrate choice (respiratory quotient) during moderate exercise, but the whole-leg and molecular differences in fatty acid mobilization could explain the age- and immobilization-induced IMTG accumulation.

Caloric restriction improves glycaemic control without reducing plasma branched-chain amino acids or keto-acids in obese men.

Higher plasma leucine, isoleucine and valine (BCAA) concentrations are associated with diabetes, obesity and insulin resistance (IR). Here, we evaluated the effects of 6-weeks very-low calorie diet (VLCD) upon fasting BCAA in overweight (OW) non-diabetic men, to explore associations between circulating BCAA and IR, before and after a weight loss intervention. Fasting plasma BCAAs were quantified in an OW (n = 26; BMI 32.4 ± 3 kg/m2; mean age 44 ± 9 y) and a normal-weight (NW) group (n = 26; BMI 24 ± 3.1 kg/m2; mean age 32 ± 12.3 y). Ten of the OW group (BMI 32.2 ± 4 kg/m2; 46 ± 8 y) then underwent 6-weeks of VLCD (600-800 kcal/day). Fasting plasma BCAA (gas chromatography-mass spectrometry), insulin sensitivity (HOMA-IR) and body-composition (DXA) were assessed before and after VLCD. Total BCAA were higher in OW individuals (sum leucine/isoleucine/valine: 457 ± 85 µM) compared to NW control individuals (365 ± 78 µM, p < 0.001). Despite significant weight loss (baseline 103.9 ± 12.3 to 93 ± 9.6 kg and BMI 32.2 ± 4 to 28.9 ± 3.6 kg/m2), no changes were observed in BCAAs after 6-weeks of VLCD. Moreover, although VLCD resulted in a significant reduction in HOMA-IR (baseline 1.19 ± 0.62 to 0.51 ± 0.21 post-VLCD; p < 0.001), Pearson's r revealed no relationships between BCAA and HOMA-IR, either before (leucine R2: 2.49e-005, p = 0.98; isoleucine R2: 1.211-e006, p = 0.9; valine R2: 0.004, p = 0.85) or after VLCD (leucine R2: 0.003, p = 0.86; isoleucine R2: 0.006, p = 0.82; valine R2: 0.002, p = 0.65). Plasma BCAA are higher in OW compared to NW individuals. However, while 6-weeks VLCD reduced body weight and IR in OW individuals, this was not associated with reductions in BCAA. This suggests that studies demonstrating links between BCAA and insulin resistance in OW individuals, are complex and are not normalised by simply losing weight.

Dietary supplements for athletes: emerging trends and recurring themes.

Dietary supplements are widely used at all levels of sport. Changes in patterns of supplement use are taking place against a background of changes in the regulatory framework that governs the manufacture and distribution of supplements in the major markets. Market regulation is complicated by the increasing popularity of Internet sales. The need for quality control of products to ensure they contain the listed ingredients in the stated amount and to ensure the absence of potentially harmful substances is recognized. This latter category includes compounds prohibited under anti-doping regulations. Several certification programmes now provide testing facilities for manufacturers of both raw ingredients and end products to ensure the absence of prohibited substances. Athletes should carry out a cost-benefit analysis for any supplement they propose to use. For most supplements, the evidence is weak, or even completely absent. A few supplements, including caffeine, creatine, and bicarbonate, are supported by a strong research base. Difficulties arise when new evidence appears to support novel supplements: in recent years, ß-alanine has become popular, and the use of nitrate and arginine is growing. Athletes seldom wait until there is convincing evidence of efficacy or of safety, but caution is necessary to minimize risk.

Age-related changes in muscle architecture and metabolism in humans: The likely contribution of physical inactivity to age-related functional decline.

In the United Kingdom (UK), it is projected that by 2035 people aged >65 years will make up 23 % of the population, with those aged >85 years accounting for 5% of the total population. Ageing is associated with progressive changes in muscle metabolism and a decline in functional capacity, leading to a loss of independence. Muscle metabolic changes associated with ageing have been linked to alterations in muscle architecture and declines in muscle mass and insulin sensitivity. However, the biological features often attributed to muscle ageing are also seen in controlled studies of physical inactivity (e.g. reduced step-count and bed-rest), and it is currently unclear how many of these ageing features are due to ageing per se or sedentarism. This is particularly relevant at a time of home confinements reducing physical activity levels during the Covid-19 pandemic. Current knowledge gaps include the relative contribution that physical inactivity plays in the development of many of the negative features associated with muscle decline in older age. Similarly, data demonstrating positive effects of government recommended physical activity guidelines on muscle health are largely non-existent. It is imperative therefore that research examining interactions between ageing, physical activity and muscle mass and metabolic health is prioritised so that it can inform on the "normal" muscle ageing process and on strategies for improving health span and well-being. This review will focus on important changes in muscle architecture and metabolism that accompany ageing and highlight the likely contribution of physical inactivity to these changes.

Optimization of insulin-mediated creatine retention during creatine feeding in humans.

The aim of this study was to determine whether creatine ingested in combination with relatively small quantities of essential amino acids, simple sugars, and protein would stimulate insulin release and augment whole-body creatine retention to the same extent as a large bolus of simple sugars. Seven young, healthy males underwent three randomized, 3-day experimental trials. Each day, 24-h urine collections were made, and on the second day participants received 5 g creatine + water (creatine trial), 5 g creatine + approximately 95 g dextrose (creatine + carbohydrate) or 5 g creatine + 14 g protein hydrolysate, 7 g leucine, 7 g phenylalanine, and 57 g dextrose (creatine + protein, amino acids, and carbohydrate) via naso-gastric tube at three equally spaced intervals. Blood samples were collected at predetermined intervals after the first and third naso-gastric bolus. After administration of the first and third bolus, serum insulin concentration was increased by 15 min (P < 0.05) in the creatine + carbohydrate and creatine + protein, amino acids, and carbohydrate trials compared with creatine alone, and plasma creatine increased more following creatine alone (15 min, P < 0.05) than in the creatine + carbohydrate and creatine + protein, amino acids, and carbohydrate trials. Urinary creatine excretion was greater with creatine alone (P < 0.05) than with creatine + carbohydrate and creatine + protein, amino acids, and carbohydrate. Administration of creatine + protein, amino acids, and carbohydrate can stimulate insulin release and augment whole-body creatine retention to the same extent as when larger quantities of simple sugars are ingested.

The involvement of the ubiquitin proteasome system in human skeletal muscle remodelling and atrophy.

Skeletal muscle exhibits great plasticity in response to altered activity levels, ultimately resulting in tissue remodelling and substantial changes in mass. Animal research would suggest that the ubiquitin proteasome system, in particular the ubiquitin ligases MAFbx/atrogin-1 and MuRF1, are instrumental to the processes underlying these changes. This review article therefore examines the role of proteasomal-mediated protein degradation in human skeletal muscle in health and disease. Specifically, the effects of exercise, disuse and inflammatory disease states on the ubiquitin proteasome system in human skeletal muscle are examined. The article also identifies several inconsistencies between published human studies and data obtained from animal models of muscle atrophy, highlighting the need for a more comprehensive examination of the molecular events responsible for modulating muscle mass in humans.

Maximal-intensity exercise does not fully restore muscle pyruvate dehydrogenase complex activation after 3 days of high-fat dietary intake.

BACKGROUND & AIMS: Exercise activates muscle pyruvate dehydrogenase complex (PDC), but moderate intensity exercise fails to fully activate muscle PDC after high-fat diet [1]. We investigated whether maximal intensity exercise overcomes this inhibition. METHODS: Quadriceps femoris muscle biopsy samples were obtained from healthy males at rest, and after 46 and 92 electrically-evoked maximal intermittent isometric contractions, which were preceded by 3 days of either low- (18%) or high- (69%) isocaloric dietary fat intake (LFD and HFD, respectively). RESULTS: The ratio of PDCa (active form) to total PDCt (fully activated) at rest was 50% less after HFD (0.32 ± 0.01 vs 0.15 ± 0.01; P < 0.05). This ratio increased to 0.77 ± 0.06 after 46 contractions (P < 0.001) and to 0.98 ± 0.07 after 92 contractions (P < 0.001) in LFD. The corresponding values after HFD were less (0.54 ± 0.06; P < 0.01 and 0.70 ± 0.07; P < 0.01, respectively). Resting muscle acetyl-CoA and acetylcarnitine content was greater after HFD than LFD (both P < 0.05), but their rate of accumulation in the former was reduced during contraction. Muscle lactate content after 92 contractions was 30% greater after HFD (P < 0.05). Muscle force generation during contraction was no different between interventions, but HFD lengthened muscle relaxation time (P < 0.05). Daily urinary total carnitine excretion after HFD was 2.5-fold greater than after LFD (P < 0.01). CONCLUSIONS: A bout of maximal intense exercise did not overcome dietary fat-mediated inhibition of muscle pyruvate dehydrogenase complex activation, and was associated with greater muscle lactate accumulation, as a result of lower PDC flux, and increased muscle relaxation time.

Temporal changes in the involvement of pyruvate dehydrogenase complex in muscle lactate accumulation during lipopolysaccharide infusion in rats.

A characteristic manifestation of sepsis is muscle lactate accumulation. This study examined any putative (causative) association between pyruvate dehydrogenase complex (PDC) inhibition and lactate accumulation in the extensor digitorum longus (EDL) muscle of rats infused with lipopolysaccharide (LPS), and explored the involvement of increased transcription of muscle-specific pyruvate dehydrogenase kinase (PDK) isoenzymes. Conscious, male Sprague-Dawley rats were infused i.v. with saline (0.4 ml h(-1), control) or LPS (150 mug kg(-1) h(-1)) for 2 h, 6 h or 24 h (n = 6-8). Muscle lactate concentration was elevated after 2, 6 and 24 h LPS infusion. Muscle PDC activity was the same at 2 h and 6 h, but was 65% lower after 24 h of LPS infusion (P < 0.01), when there was a 47% decrease in acetylcarnitine concentration (P < 0.05), and a 24-fold increase in PDK4 mRNA expression (P < 0.001). These changes were preceded by marked increases in tumour necrosis factor-alpha and interleukin-6 mRNA expression at 2 h. The findings indicate that the early (2 and 6 h) elevation in muscle lactate concentration during LPS infusion was not attributable to limited muscle oxygen availability or ATP production (evidenced by unchanged ATP and phosphocreatine (PCr) concentrations) or to PDC inhibition, whereas after 24 h, muscle lactate accumulation appears to have resulted from PDC activation status limiting pyruvate flux, most probably due to cytokine-mediated up-regulation of PDK4 transcription.

The plasma ammonia response to cycle exercise in COPD.

The plasma ammonia response to exercise in chronic obstructive pulmonary disease (COPD) was examined and the relationship between plasma ammonia concentration and muscle adenine nucleotide metabolism was explored. In total, 25 stable COPD patients and 13 similar-aged controls underwent incremental and constant-work rate cycle exercise tests. Arterialised venous blood was sampled at rest, at 1-min intervals during exercise and

Increased acetyl group availability enhances contractile function of canine skeletal muscle during ischemia.

Skeletal muscle contractile function is impaired during acute ischemia such as that experienced by peripheral vascular disease patients. We therefore, examined the effects of dichloroacetate, which can alter resting metabolism, on canine gracilis muscle contractile function during constant flow ischemia. Pretreatment with dichloroacetate increased resting pyruvate dehydrogenase complex activity and resting acetylcarnitine concentration by approximately 4- and approximately 10-fold, respectively. After 20-min contraction the control group had demonstrated an approximately 40% reduction in isomeric tension whereas the dichloroacetate group had fatigued by approximately 25% (P < 0.05). Dichloroacetate resulted in less lactate accumulation (10.3 +/- 3.0 vs 58.9 +/- 10.5 mmol.kg-1 dry muscle [dm], P < 0.05) and phosphocreatine hydrolysis (15.6 +/- 6.3 vs 33.8 +/- 9.0 mmol.kg-1 dm, P < 0.05) during contraction. Acetylcarnitine concentration fell during contraction by 5.4 +/- 1.8 mmol.kg-1 dm in the dichloroacetate group but increased by 10.0 +/- 1.9 mmol.kg-1 dm in the control group. In conclusion, dichloroacetate enhanced contractile function during ischemia, independently of blood flow, such that it appears oxidative ATP regeneration is limited by pyruvate dehydrogenase complex activity and acetyl group availability.

Substrate availability limits human skeletal muscle oxidative ATP regeneration at the onset of ischemic exercise.

We have demonstrated previously that dichloroacetate can attenuate skeletal muscle fatigue by up to 35% in a canine model of peripheral ischemia (Timmons, J.A., S.M. Poucher, D. Constantin-Teodosiu, V. Worrall, I.A. Macdonald, and P.L. Greenhaff. 1996. J. Clin. Invest. 97:879-883). This was thought to be a consequence of dichloroacetate increasing acetyl group availability early during contraction. In this study we characterized the metabolic effects of dichloroacetate in a human model of peripheral muscle ischemia. On two separate occasions (control-saline or dichloroacetate infusion), nine subjects performed 8 min of single-leg knee extension exercise at an intensity aimed at achieving volitional exhaustion in approximately 8 min. During exercise each subject's lower limbs were exposed to 50 mmHg of positive pressure, which reduces blood flow by approximately 20%. Dichloroacetate increased resting muscle pyruvate dehydrogenase complex activation status by threefold and elevated acetylcarnitine concentration by fivefold. After 3 min of exercise, phosphocreatine degradation and lactate accumulation were both reduced by approximately 50% after dichloroacetate pretreatment, when compared with control conditions. However, after 8 min of exercise no differences existed between treatments. Therefore, it would appear that dichloroacetate can delay the accumulation of metabolites which lead to the development of skeletal muscle fatigue during ischemia but does not alter the metabolic profile when a maximal effort is approached.

Effect of oral creatine supplementation on human muscle GLUT4 protein content after immobilization.

The purpose of this study was to investigate the effect of oral creatine supplementation on muscle GLUT4 protein content and total creatine and glycogen content during muscle disuse and subsequent training. A double-blind placebo-controlled trial was performed with 22 young healthy volunteers. The right leg of each subject was immobilized using a cast for 2 weeks, after which subjects participated in a 10-week heavy resistance training program involving the knee-extensor muscles (three sessions per week). Half of the subjects received creatine monohydrate supplements (20 g daily during the immobilization period and 15 and 5 g daily during the first 3 and the last 7 weeks of rehabilitation training, respectively), whereas the other 11 subjects ingested placebo (maltodextrine). Muscle GLUT4 protein content and glycogen and total creatine concentrations were assayed in needle biopsy samples from the vastus lateralis muscle before and after immobilization and after 3 and 10 weeks of training. Immobilization decreased GLUT4 in the placebo group (-20%, P < 0.05), but not in the creatine group (+9% NS). Glycogen and total creatine were unchanged in both groups during the immobilization period. In the placebo group, during training, GLUT4 was normalized, and glycogen and total creatine were stable. Conversely, in the creatine group, GLUT4 increased by approximately 40% (P < 0.05) during rehabilitation. Muscle glycogen and total creatine levels were higher in the creatine group after 3 weeks of rehabilitation (P < 0.05), but not after 10 weeks of rehabilitation. We concluded that 1) oral creatine supplementation offsets the decline in muscle GLUT4 protein content that occurs during immobilization, and 2) oral creatine supplementation increases GLUT4 protein content during subsequent rehabilitation training in healthy subjects.

Does dietary creatine supplementation play a role in skeletal muscle metabolism and performance?

Fatigue sustained during short-term, high-intensity exercise in humans is associated with the inability of skeletal muscle to maintain a high rate of anaerobic ATP production from phosphocreatine hydrolysis. Ingestion of creatine monohydrate at a rate of 20 g/d for 5-6 d was shown to increase the total creatine concentration of human skeletal muscle by approximately 25 mmol/kg dry mass, some 30% of this in phosphorylated form as phosphocreatine. A positive relation was then shown between muscle creatine uptake and improvements in performance during repeated bouts of maximal exercise. However, there is no evidence that increasing intake > 20-30 g/d for 5-6 d has any potentiating effect on creatine uptake or performance. In individuals in whom the initial total creatine concentration already approached 150 mmol/kg dry mass, neither creatine uptake nor an effect on phosphocreatine resynthesis or performance was found after supplementation. Loss of ATP during heavy anaerobic exercise was found to decline after creatine ingestion, despite an increase in work production. These results suggest that improvements in performance are due to parallel improvements in ATP resynthesis during exercise as a consequence of increased phosphocreatine availability. Creatine uptake is augmented by combining creatine supplementation with exercise and with carbohydrate ingestion.

Dual effects of dichloroacetate on cardiac ischaemic preconditioning in the rat isolated perfused heart.

1. Ischaemic cardiac preconditioning represents an important cardioprotective mechanism which limits myocardial ischaemic damage. The aim of this investigation was to assess the impact of dichloroacetate (DCA), a pyruvate dehydrogenase complex activator, on preconditioning. 2. Rat isolated hearts were perfused by use of the Langendorff technique, and were subjected to either preconditioning (3 x 4 or 3 x 6 min ischaemia) or continuous perfusion, followed by 30 min global ischaemia and 60 min reperfusion. DCA (3 mM) was either given throughout the protocol (pretreatment), during reperfusion only (post-treatment), or not at all. Throughout reperfusion mechanical performance was assessed as the rate-pressure product (RPP: left ventricular developed pressure x heart rate). 3. In non-preconditioned control hearts, mechanical performance was substantially (P < 0.001) depressed on reperfusion (the RPP after 60 min of reperfusion (RPP(t=60)) was 4,246+/-974 mmHg beats min(-1) compared to baseline value of 21,297+/-1,728 mmHg beats min(-1)). Preconditioning with either 3 x 4 min or 3 x 6 min cycles caused significant protection, as shown by enhanced recovery (RPP(t=60) = 7,818+/-1,138, P < 0.05, and 11,123+/-587 mmHg beats min(-1), P < 0.001, respectively). 4. Addition of DCA (3 mM) to hearts under baseline conditions significantly (P < 0.001) enhanced systolic function with an increased left ventricular developed pressure of 108+/-5 mmHg compared to 88.3+/-3.0 mmHg in the controls. 5. Pretreatment with 3 mM DCA had no effect on recovery of mechanical performance in the non-preconditioned hearts (RPP(t=60) = 3,640+/-1,235 mmHg beats min(-1)) while the beneficial effects of preconditioning were reduced in the preconditioned hearts (3 x 4 min: RPP(t=60) = 2,919+/-1,060 mmHg beats min(-1); 3 x 6 min: RPP(t=60) = 8,032+/-1,367 mmHg beats min(-1)). Therefore, DCA had increased the threshold for preconditioning. 6. By contrast, post-treatment of hearts with 3 mM DCA substantially improved recovery on reperfusion in all groups (RPP(t=60) = 5,827+/-1,328 (non-preconditioned), 14,022+/-3,743 (3 x 4 min; P < 0.01) and 23,219+/-1,374 (3 x 6 min; P < 0.001) mmHg beats min(-1)). 7. The results of the present investigation clearly show that pretreatment with DCA enhances baseline cardiac mechanical performance but increases the threshold for cardiac preconditioning. However, post-treatment with DCA substantially augments the beneficial effects of preconditioning.