Low glycogen and branched-chain amino acid ingestion do not impair anaplerosis during exercise in humans.

We examined the hypothesis that increasing the rate of branched-chain amino acid (BCAA) oxidation, during conditions of low glycogen availability, reduces the level of muscle tricarboxylic acid cycle intermediates (TCAI) by placing a carbon "drain" on the cycle at the level of 2-oxoglutarate. Six men cycled at approximately 70% of maximal oxygen uptake for 15 min under two conditions: 1) low preexercise muscle glycogen (placebo) and 2) low glycogen combined with BCAA ingestion. We have previously shown that BCAA ingestion increased the activity of branched-chain oxoacid dehydrogenase, the rate-limiting enzyme for BCAA oxidation in muscle, compared with low glycogen alone [M. L. Jackman, M. J. Gibala, E. Hultman, and T. E. Graham. Am. J. Physiol. 272 (Endocrinol. Metab. 35): E233-E238, 1997]. Muscle glycogen concentration was 185 +/- 22 and 206 +/- 22 mmol/kg dry wt at rest for the placebo and BCAA-supplemented trials, respectively, and decreased to 109 +/- 18 and 96 +/- 10 mmol/kg dry wt after exercise. The net increase in the total concentration of six measured TCAI ( approximately 95% of TCAI pool) during exercise was not different between trials (3.97 +/- 0. 34 vs. 3.88 +/- 0.34 mmol/kg dry wt for the placebo and BCAA trials, respectively). Muscle 2-oxoglutarate concentration decreased from approximately 0.05 at rest to approximately 0.03 mmol/kg dry wt after exercise in both trials. The magnitude of TCAI pool expansion in both trials was similar to that seen previously in subjects who performed an identical exercise bout after a normal mixed diet [M. J. Gibala, M. A. Tarnopolsky, and T. E. Graham. Am. J. Physiol. 272 (Endocrinol. Metab. 35): E239-E244, 1997]. These data suggest that increasing the rate of BCAA oxidation has no measurable effect on muscle TCAI during exercise with low glycogen in humans. Moreover, it appears that low resting glycogen per se does not impair the increase in TCAI during moderate exercise.     

Effect of carbohydrate ingestion on ammonia metabolism during exercise in humans.

The present study was undertaken to examine the effect of carbohydrate ingestion on plasma and muscle ammonia (NH(3) denotes ammonia and ammonium) accumulation during prolonged exercise. Eleven trained men exercised for 2 h at 65% peak pulmonary oxygen consumption while ingesting either 250 ml of an 8% carbohydrate-electrolyte solution every 15 min (CHO) or an equal volume of a sweet placebo. Blood glucose and plasma insulin levels during exercise were higher in CHO, but plasma hypoxanthine was lower after 120 min (1.7 +/- 0.3 vs. 2.6 +/- 0.1 micromol/l; P < 0. 05). Plasma NH(3) levels were similar at rest and after 30 min of exercise in both trials but were lower after 60, 90, and 120 min of exercise in CHO (62 +/- 9 vs. 76 +/- 9 micromol/l; P < 0.05). Muscle NH(3) levels were similar at rest and after 30 min of exercise but were lower after 120 min of exercise in CHO (1.51 +/- 0.21 vs. 2.07 +/- 0.23 mmol/kg dry muscle; P < 0.05; n = 5). These data are best explained by carbohydrate ingestion reducing muscle NH(3) production from amino acid degradation, although a small reduction in net AMP catabolism within the contracting muscle may also make a minor contribution to the lower tissue NH(3) levels.     

Exercise training increases branched-chain oxoacid dehydrogenase kinase content in human skeletal muscle

The branched-chain oxoacid dehydrogenase complex (BCOAD) is rate determining for the oxidation of branched-chain amino acids (BCAAs) in skeletal muscle. Exercise training blunts the acute exercise-induced activation of BCOAD (BCOADa) in human skeletal muscle (McKenzie S, Phillips SM, Carter SL, Lowther S, Gibala MJ, Tarnopolsky MA. Am J Physiol Endocrinol Metab 278: E580-E587, 2000); however, the mechanism is unknown. We hypothesized that training would increase the muscle protein content of BCOAD kinase, the enzyme responsible for inactivation of BCOAD by phosphorylation. Twenty subjects [23 +/- 1 yr; peak oxygen uptake (.VO(2peak)) = 41 +/- 2 ml.kg(-1).min(-1)] performed 6 wk of either high-intensity interval or continuous moderate-intensity training on a cycle ergometer (n = 10/group). Before and after training, subjects performed 60 min of cycling at 65% of pretraining .VO(2peak), and needle biopsy samples (vastus lateralis) were obtained before and immediately after exercise. The effect of training was demonstrated by an increased .VO(2peak), increased citrate synthase maximal activity, and reduced muscle glycogenolysis during exercise, with no difference between groups (main effects, P < 0.05). BCOADa was lower after training (main effect, P < 0.05), and this was associated with a approximately 30% increase in BCOAD kinase protein content (main effect, P < 0.05). We conclude that the increased protein content of BCOAD kinase may be involved in the mechanism for reduced BCOADa after exercise training in human skeletal muscle. These data also highlight differences in models used to study the regulation of skeletal muscle BCAA metabolism, since exercise training was previously reported to increase BCOADa during exercise and decrease BCOAD kinase content in rats (Fujii H, Shimomura Y, Murakami T, Nakai N, Sato T, Suzuki M, Harris RA. Biochem Mol Biol Int 44: 1211-1216, 1998).     

Suppressing lipolysis increases interleukin-6 at rest and during prolonged moderate-intensity exercise in humans.

IL-6 induces lipolysis when administered to humans. Consequently, it has been hypothesized that IL-6 is released from skeletal muscle during exercise to act in a "hormonelike" manner and increase lipolysis from adipose tissue to supply the muscle with substrate. In the present study, we hypothesized that suppressing lipolysis, and subsequent free fatty acid (FFA) availability, would result in a compensatory elevation in IL-6 at rest and during exercise. First, we had five healthy men ingest nicotinic acid (NA) at 30-min intervals for 120 min at rest [10 mg/kg body mass (initial dose), 5 mg/kg body mass (subsequent doses)]. Plasma was collected and analyzed for FFA and IL-6. After 120 min, plasma FFA concentration was attenuated (0 min: 0.26 +/- 0.05 mmol/l; 120 min: 0.09 +/- 0.02 mmol/l; P < 0.01), whereas plasma IL-6 was concomitantly increased approximately eightfold (0 min: 0.75 +/- 0.18 pg/ml; 120 min: 6.05 +/- 0.89 pg/ml; P < 0.001). To assess the effect of lipolytic suppression on the exercise-induced IL-6 response, seven active, but not specifically trained, men performed two experimental exercise trials with (NA) or without [control (Con)] NA ingestion 60 min before (10 mg/kg body mass) and throughout (5 mg/kg body mass every 30 min) exercise. Blood samples were obtained before ingestion, 60 min after ingestion, and throughout 180 min of cycling exercise at 62 +/- 5% of maximal oxygen consumption. IL-6 gene expression, in muscle and adipose tissue sampled at 0, 90, and 180 min, was determined by using semiquantitative real-time PCR. IL-6 mRNA increased in Con (rest vs. 180 min; P < 0.01) approximately 13-fold in muscle and approximately 42-fold in fat with exercise. NA increased (rest vs. 180 min; P < 0.01) IL-6 mRNA 34-fold in muscle, but the treatment effect was not statistically significant (Con vs. NA, P = 0.1), and 235-fold in fat (Con vs. NA, P < 0.01). Consistent with the study at rest, NA completely suppressed plasma FFA (180 min: Con, 1.42 +/- 0.07 mmol/l; NA, 0.10 +/- 0.01 mmol/l; P < 0.001) and increased plasma IL-6 (180 min: Con, 9.81 +/- 0.98 pg/ml; NA, 19.23 +/- 2.50 pg/ml; P < 0.05) during exercise. In conclusion, these data demonstrate that circulating IL-6 is markedly elevated at rest and during prolonged moderate-intensity exercise when lipolysis is suppressed.     

Progressive increase in human skeletal muscle AMPKalpha2 activity and ACC phosphorylation during exercise

The effect of prolonged moderate-intensity exercise on human skeletal muscle AMP-activated protein kinase (AMPK)alpha1 and -alpha2 activity and acetyl-CoA carboxylase (ACCbeta) and neuronal nitric oxide synthase (nNOSmu) phosphorylation was investigated. Seven active healthy individuals cycled for 30 min at a workload requiring 62.8 +/- 1.3% of peak O(2) consumption (VO(2 peak)) with muscle biopsies obtained from the vastus lateralis at rest and at 5 and 30 min of exercise. AMPKalpha1 activity was not altered by exercise; however, AMPKalpha2 activity was significantly (P < 0.05) elevated after 5 min (approximately 2-fold), and further elevated (P < 0.05) after 30 min (approximately 3-fold) of exercise. ACCbeta phosphorylation was increased (P < 0.05) after 5 min (approximately 18-fold compared with rest) and increased (P < 0.05) further after 30 min of exercise (approximately 36-fold compared with rest). Increases in AMPKalpha2 activity were significantly correlated with both increases in ACCbeta phosphorylation and reductions in muscle glycogen content. Fat oxidation tended (P = 0.058) to increase progressively during exercise. Muscle creatine phosphate was lower (P < 0.05), and muscle creatine, calculated free AMP, and free AMP-to-ATP ratio were higher (P < 0.05) at both 5 and 30 min of exercise compared with those at rest. At 30 min of exercise, the values of these metabolites were not significantly different from those at 5 min of exercise. Phosphorylation of nNOSmu was variable, and despite the mean doubling with exercise, statistically significance was not achieved (P = 0.304). Western blots indicated that AMPKapproximately 2 was associated with both nNOSmu and ACCbeta consistent with them both being substrates of AMPKalpha2 in vivo. In conclusion, AMPKalpha2 activity and ACCbeta phosphorylation increase progressively during moderate exercise at approximately 60% of VO(2 peak) in humans, with these responses more closely coupled to muscle glycogen content than muscle AMP/ATP ratio.     

Hormone-sensitive lipase activity and fatty acyl-CoA content in human skeletal muscle during prolonged exercise.

Hormone-sensitive lipase (HSL) catalyzes the hydrolysis of intramuscular triacylglycerols (IMTGs), but HSL regulation is poorly understood in skeletal muscle. The present study measured human skeletal muscle HSL activity at rest and during 120 min of cycling at 60% of peak O2 uptake. Several putative HSL regulators were also measured, including muscle long-chain fatty acyl-CoA (LCFA CoA) and free AMP contents and plasma epinephrine and insulin concentrations. HSL activity increased from resting levels by 10 min of exercise (from 2.09 +/- 0.19 to 2.56 +/- 0.22 mmol. min-1x kg dry mass-1, P < 0.05), increased further by 60 min (to 3.12 +/- 0.27 mmol x min-1x kg dry mass-1, P < 0.05), and decreased to near-resting rates after 120 min of cycling. Skeletal muscle LCFA CoA increased (P < 0.05) above rest by 60 min (from 15.9 +/- 3.0 to 50.4 +/- 7.9 micromol/kg dry mass) and increased further by 120 min. Estimated free AMP increased (P < 0.05) from rest to 60 min and was approximately 20-fold greater than that at rest by 120 min. Epinephrine was increased above rest (P < 0.05) at 60 (1.47 +/- 0.15 nM) and 120 min (4.87 +/- 0.76 nM) of exercise. Insulin concentrations decreased rapidly and were lower than resting levels by 10 min and continued to decrease throughout exercise. In summary, HSL activity was increased from resting levels by 10 min, increased further by 60 min, and decreased to near-resting values by 120 min. The increased HSL activity at 60 min was associated with the stimulating effect of increased epinephrine and decreased insulin levels. After 120 min, the decreased HSL activity was associated with the proposed inhibitory effects of increased free AMP. The accumulation of LCFA CoA in the 2nd h of exercise may also have reduced the flux through HSL and accounted for the reduction in IMTG utilization previously observed late in prolonged exercise.     

Effects of carbohydrate feedings on plasma free tryptophan and branched-chain amino acids during prolonged cycling

Brain serotonin (5-hydroxytryptamine, 5-HT) has been suggested to be involved in central fatigue during prolonged exercise. Changes in the ratio of plasma free tryptophan (free Trp) to branched-chain amino acids (BCAA) are associated with altered brain 5-HT synthesis. The purposes of this study were to describe systematically the effects of prolonged exercise on changes in plasma free Trp and BCAA and to examine the effects of carbohydrate (CHO) feedings on these same variables. Eight well-trained men [VO2max = 57.8 (SE 4.1) ml kg-1 min-1] cycled for up to 255 min at a power output corresponding to VO2 at lactate threshold (approximately 68% VO2max) on three occasions separated by at least 1 week. Subjects drank 5 ml kg-1 body wt-1 of either a water placebo, or a liquid beverage containing a moderate (6% CHO) or high (12% CHO) concentration of carbohydrate beginning at min 14 of exercise and every 30 min thereafter. Exercise time to fatigue was shorter in subjects receiving placebo [190 (SE 4) min] as compared to 6% CHO [235 (SE 10) min] and 12% CHO [234 (SE 9) min] (P < 0.05). Glucose and insulin decreased in the placebo group, and free Trp, free-Trp/BCAA, and free fatty acids increased approximately five- to sevenfold (P < 0.05). These changes were attenuated in a dose-related manner by the carbohydrate drinks.(ABSTRACT TRUNCATED AT 250 WORDS)     

Exercise intensity: effect on postexercise O2 uptake in trained and untrained women

Despite many reports of long-lasting elevation of metabolism after exercise, little is known regarding the effects of exercise intensity and duration on this phenomenon. This study examined the effect of a constant duration (30 min) of cycle ergometer exercise at varied intensity levels [50 and 70% of maximal O2 consumption (VO2max)] on 3-h recovery of oxygen uptake (VO2). VO2 and respiratory exchange ratios were measured by open-circuit spirometry in five trained female cyclists (age 25 +/- 1.7 yr) and five untrained females (age 27 +/- 0.8 yr). Postexercise VO2 measured at intervals for 3 h after exercise was greater (P less than 0.01) after exercise at 50% VO2max in trained (0.40 +/- 0.01 l/min) and untrained subjects (0.39 +/- 0.01 l/min) than after 70% VO2max in (0.31 +/- 0.02 l/min) and untrained subjects (0.29 +/- 0.02 l/min). The lower respiratory exchange ratio values (P less than 0.01) after 50% VO2max in trained (0.78 +/- 0.01) and untrained subjects (0.80 +/- 0.01) compared with 70% VO2max in trained (0.81 +/- 0.01) and untrained subjects (0.83 +/- 0.01) suggest that an increase in fat metabolism may be implicated in the long-term elevation of metabolism after exercise. This was supported by the greater estimated fatty acid oxidation (P less than 0.05) after 50% VO2max in trained (147 +/- 4 mg/min) and untrained subjects (133 +/- 9 mg/min) compared with 70% VO2max in trained (101 +/- 6 mg/min) and untrained subjects (85 +/- 7 mg/min).     

Influence of a 36 h fast followed by refeeding with glucose, glycerol or placebo on metabolism and performance during prolonged exercise in man

Five men were studied during exercise to exhaustion on an electrically braked cycle ergometer at 70% of VO2max. The four experimental treatments were as follows: fasted for 36 h (A); fasted (36 h) and refed with glucose (B) or glycerol (C); postabsorptive (overnight fast, D). In B and C the subjects were given a drink containing glucose or glycerol (1g per kg body weight) 45 min before starting exercise. A placebo drink was given 45 min before exercise on treatments A and D. Despite an increased availability of circulating free fatty acids, beta-hydroxybutyrate and glycerol exercise time to exhaustion was significantly lower after fasting (treatment A 77.7 +/- 6.8 min) compared with treatment D (119.5 +/- 5.8 min). Refeeding with glucose or glycerol did not significantly improve performance (92.4 +/- 11.8 min and 80.8 +/- 3.6 min respectively) compared with treatment A and lowered circulating levels of FFA and beta-HB during exercise compared with A. Despite the probability of low liver glycogen levels after fasting, none of the subjects became hypoglycaemic (blood glucose less than 4 mmol.l-1) during exercise and their blood lactate concentrations were not high at exhaustion. Plasma levels of branched chain amino acids (BCAA) decreased progressively during exercise on treatments A, B and C and were considerably lower at exhaustion compared with treatment D. Falling plasma concentrations of BCAA during prolonged exercise may be implicated in the generation of central fatigue.     

Intake of branched-chain amino acids influences the levels of MAFbx mRNA and MuRF-1 total protein in resting and exercising human muscle

Resistance exercise and amino acids are two major factors that influence muscle protein turnover. Here, we examined the effects of resistance exercise and branched-chain amino acids (BCAA), individually and in combination, on the expression of anabolic and catabolic genes in human skeletal muscle. Seven subjects performed two sessions of unilateral leg press exercise with randomized supplementation with BCAA or flavored water. Biopsies were collected from the vastus lateralis muscle of both the resting and exercising legs before and repeatedly after exercise to determine levels of mRNA, protein phosphorylation, and amino acid concentrations. Intake of BCAA reduced (P < 0.05) MAFbx mRNA by 30 and 50% in the resting and exercising legs, respectively. The level of MuRF-1 mRNA was elevated (P < 0.05) in the exercising leg two- and threefold under the placebo and BCAA conditions, respectively, whereas MuRF-1 total protein increased by 20% (P < 0.05) only in the placebo condition. Phosphorylation of p70(S6k) increased to a larger extent (～2-fold; P < 0.05) in the early recovery period with BCAA supplementation, whereas the expression of genes regulating mTOR activity was not influenced by BCAA. Muscle levels of phenylalanine and tyrosine were reduced (13-17%) throughout recovery (P < 0.05) in the placebo condition and to a greater extent (32-43%; P < 0.05) following BCAA supplementation in both resting and exercising muscle. In conclusion, BCAA ingestion reduced MAFbx mRNA and prevented the exercise-induced increase in MuRF-1 total protein in both resting and exercising leg. Further-more, resistance exercise differently influenced MAFbx and MuRF-1 mRNA expression, suggesting both common and divergent regulation of these two ubiquitin ligases.     

Adrenergic regulation of HSL serine phosphorylation and activity in human skeletal muscle during the onset of exercise

Skeletal muscle hormone-sensitive lipase (HSL) activity is increased by contractions and increases in blood epinephrine (EPI) concentrations and cyclic AMP activation of the adrenergic pathway during prolonged exercise. To determine the importance of hormonal stimulation of HSL activity during the onset of moderate- and high-intensity exercise, nine men [age 24.3 +/- 1.2 yr, 80.8 +/- 5.0 kg, peak oxygen consumption (VO2 peak) 43.9 +/- 3.6 ml x kg(-1) x min(-1)] cycled for 1 min at approximately 65% VO2 peak, rested for 60 min, and cycled at approximately 90% VO2 peak for 1 min. Skeletal muscle biopsies were taken pre- and postexercise, and arterial blood was sampled throughout exercise. Arterial EPI increased (P < 0.05) postexercise at 65% (0.45 +/- 0.10 to 0.78 +/- 0.27 nM) and 90% VO2 peak (0.57 +/- 0.34 to 1.09 +/- 0.50 nM). HSL activity increased (P < 0.05) following 1 min of exercise at 65% VO2 peak [1.05 +/- 0.39 to 1.78 +/- 0.54 mmol x min(-1) x kg dry muscle (dm)(-1)] and 90% VO2 peak (1.07 +/- 0.24 to 1.91 +/- 0.62 mmol x min(-1) x kg dm(-1)). Cyclic AMP content also increased (P < 0.05) at both exercise intensities (65%: 1.52 +/- 0.67 to 2.75 +/- 1.12, 90%: 1.85 +/- 0.65 to 2.64 +/- 0.93 micromol/kg dm). HSL Ser660 phosphorylation (approximately 55% increase) and ERK1/2 phosphorylation ( approximately 33% increase) were augmented following exercise at both intensities, whereas HSL Ser563 and Ser565 phosphorylation were not different from rest. The results indicate that increases in arterial EPI concentration during the onset of moderate- and high-intensity exercise increase cyclic AMP content, which results in the phosphorylation of HSL Ser660. This adrenergic stimulation contributes to the increase in HSL activity that occurs in human skeletal muscle in the first minute of exercise at 65% and 90% VO2 peak.     

Effect of aerobic capacity on the T(2) increase in exercised skeletal muscle.

The increase in nuclear magnetic resonance transverse relaxation time (T(2)) of muscle water measured by magnetic resonance imaging after exercise has been correlated with work rate in human subjects. This study compared the T(2) increase in thigh muscles of trained (cycling VO(2 max) = 54.4 +/- 2.7 ml O(2). kg(-1). min(-1), mean +/- SE, n = 8, 4 female) vs. sedentary (31.7 +/- 0.9 ml O(2). kg(-1). min(-1), n = 8, 4 female) subjects after cycling exercise for 6 min at 50 and 90% of the subjects' individually determined VO(2 max). There was no significant difference between groups in the T(2) increase measured in quadriceps muscles within 3 min after the exercises, despite the fact that the absolute work rates were 60% higher in the trained group (253 +/- 15 vs. 159 +/- 21 W for the 90% exercise). In both groups, the increase in T(2) of vastus muscles was twofold greater after the 90% exercise than after the 50% exercise. The recovery of T(2) after the 90% exercise was significantly faster in vastus muscles of the trained compared with the sedentary group (mean recovery half-time 11.9 +/- 1.2 vs. 23.3 +/- 3.7 min). The results show that the increase in muscle T(2) varies with work rate relative to muscle maximum aerobic power, not with absolute work rate.     

Effect of carbohydrate feedings during high-intensity exercise

To determine the upper limits of steady-state exercise performance and carbohydrate oxidation late in exercise, seven trained men were studied on two occasions during prolonged cycling that alternated every 15 min between approximately 60% and approximately 85% of VO2max. When fed a sweet placebo throughout exercise, plasma glucose and respiratory exchange ratio (R) declined (P less than 0.05) from 5.0 +/- 0.1 mM and 0.91 +/- 0.01 after 30 min (i.e., at 85% VO2max) to 3.7 +/- 0.3 mM and 0.79 +/- 0.01 at fatigue (i.e., when the subjects were unable to continue exercise at 60% VO2max). Carbohydrate feeding throughout exercise (1 g/kg at 10 min, then 0.6 g/kg every 30 min) increased plasma glucose to approximately 6 mM and partially prevented this decline in carbohydrate oxidation, allowing the men to perform 19% more work (2.74 +/- 0.13 vs. 2.29 +/- 0.09 MJ, P less than 0.05) before fatiguing. Even when fed carbohydrate, however, by the 3rd h of exercise, R had fallen from 0.92 to 0.87, accompanied by a reduction in exercise intensity from approximately 85% to approximately 75% VO2max (both P less than 0.05). These data indicate that carbohydrate feedings enable trained cyclists to exercise at up to 75% VO2max and to oxidize carbohydrate at up to 2 g/min during the later stages of prolonged intense exercise.     

Influence of carnitine supplementation on muscle substrate and carnitine metabolism during exercise

We examined 1) the effect of L-carnitine supplementation on free fatty acid (FFA) utilization during exercise and 2) exercise-induced alterations in plasma levels and skeletal muscle exchange of carnitine. Seven moderately trained human male subjects serving as their own controls participated in two bicycle exercise sessions (120 min, 50% of VO2max). The second exercise was preceded by 5 days of oral carnitine supplementation (CS; 5 g daily). Despite a doubling of plasma carnitine levels, with CS, there were no effects on exercise-induced changes in arterial levels and turnover of FFA, the relation between leg FFA inflow and FFA uptake, or the leg exchange of other substrates. Heart rate during exercise after CS decreased 7-8%, but O2 uptake was unchanged. Exercise before CS induced a fall from 33.4 +/- 1.6 to 30.8 +/- 1.0 (SE) mumol/l in free plasma carnitine despite a release (2.5 +/- 0.9 mumol/min) from the leg. Simultaneously, acylated plasma carnitine rose from 5.0 +/- 1.0 to 14.2 +/- 1.4 mumol/l, with no evidence of leg release. Consequently, total plasma carnitine increased. We concluded that in healthy subjects CS does not influence muscle substrate utilization either at rest or during prolonged exercise and that free carnitine released from muscle during exercise is presumably acylated in the liver and released to plasma.     

Skeletal muscle metabolism is unaffected by DCA infusion and hyperoxia after onset of intense aerobic exercise

This study investigated whether hyperoxic breathing (100% O(2)) or increasing oxidative substrate supply [dichloroacetate (DCA) infusion] would increase oxidative phosphorylation and reduce the reliance on substrate phosphorylation at the onset of high-intensity aerobic exercise. Eight male subjects cycled at 90% maximal O(2) uptake (VO(2 max)) for 90 s in three randomized conditions: 1) normoxic breathing and saline infusion over 1 h immediately before exercise (CON), 2) normoxic breathing and saline infusion with DCA (100 mg/kg body wt), and 3) hyperoxic breathing for 20 min at rest and during exercise and saline infusion (HYP). Muscle biopsies from the vastus lateralis were sampled at rest and after 30 and 90 s of exercise. DCA infusion increased pyruvate dehydrogenase (PDH) activation above CON and HYP (3.10 +/- 0.23, 0.56 +/- 0.08, 0.69 +/- 0.05 mmol x kg wet muscle(-1) x min(-1), respectively) and significantly increased both acetyl-CoA and acetylcarnitine (11.0 +/- 0.7, 2.0 +/- 0.5, 2.2 +/- 0.5 mmol/kg dry muscle, respectively) at rest. However, DCA and HYP did not alter phosphocreatine degradation and lactate accumulation and, therefore, the reliance on substrate phosphorylation during 30 s (CON, 51.2 +/- 5.4; DCA, 56.5 +/- 7.1; HYP, 69.5 +/- 6.3 mmol ATP/kg dry muscle) and 90 s of exercise (CON, 90.6 +/- 9.5; DCA, 107.2 +/- 13.0; HYP, 101.2 +/- 15.2 mmol ATP/kg dry muscle). These data suggest that the rate of oxidative phosphorylation at the onset of exercise at 90% VO(2 max) is not limited by oxygen availability to the active muscle or by substrate availability (metabolic inertia) at the level of PDH in aerobically trained subjects.     

Branched-chain amino acid supplementation during bed rest: effect on recovery.

Bed rest is associated with a loss of protein from the weight-bearing muscle. The objectives of this study are to determine whether increasing dietary branched-chain amino acids (BCAAs) during bed rest improves the anabolic response after bed rest. The study consisted of a 1-day ambulatory period, 14 days of bed rest, and a 4-day recovery period. During bed rest, dietary intake was supplemented with either 30 mmol/day each of glycine, serine, and alanine (group 1) or with 30 mmol/day each of the three BCAAs (group 2). Whole body protein synthesis was determined with U-(15)N-labeled amino acids, muscle, and selected plasma protein synthesis with l-[(2)H(5)]phenylalanine. Total glucose production and gluconeogenesis from alanine were determined with l-[U-(13)C(3)]alanine and [6,6-(2)H(2)]glucose. During bed rest, nitrogen (N) retention was greater with BCAA feeding (56 +/- 6 vs. 26 +/- 12 mg N. kg(-1). day(-1), P < 0.05). There was no effect of BCAA supplementation on either whole body, muscle, or plasma protein synthesis or the rate of 3-MeH excretion. Muscle tissue free amino acid concentrations were increased during bed rest with BCAA (0.214 +/- 0.066 vs. 0.088 +/- 0.12 nmol/mg protein, P < 0.05). Total glucose production and gluconeogenesis from alanine were unchanged with bed rest but were significantly reduced (P < 0.05) with the BCAA group in the recovery phase. In conclusion, the improved N retention during bed rest is due, at least in part, to accretion of amino acids in the tissue free amino acid pools. The amount accreted is not enough to impact protein kinetics in the recovery phase but does improve N retention by providing additional essential amino acids in the early recovery phase.     

Decreased PDH activation and glycogenolysis during exercise following fat adaptation with carbohydrate restoration

Five days of a high-fat diet while training, followed by 1 day of carbohydrate (CHO) restoration, increases rates of whole body fat oxidation and decreases CHO oxidation during aerobic cycling. The mechanisms responsible for these shifts in fuel oxidation are unknown but involve up- and downregulation of key regulatory enzymes in the pathways of skeletal muscle fat and CHO metabolism, respectively. This study measured muscle PDH and HSL activities before and after 20 min of cycling at 70% VO2peak and 1 min of sprinting at 150% peak power output (PPO). Estimations of muscle glycogenolysis were made during the initial minute of exercise at 70% VO2peak and during the 1-min sprint. Seven male cyclists undertook this exercise protocol on two occasions. For 5 days, subjects consumed in random order either a high-CHO (HCHO) diet (10.3 g x kg(-1) x day(-1) CHO, or approximately 70% of total energy intake) or an isoenergetic high-fat (FAT-adapt) diet (4.6 g x kg(-1) x day(-1) FAT, or 67% of total energy) while undertaking supervised aerobic endurance training. On day 6 for both treatments, subjects ingested an HCHO diet and rested before their experimental trials on day 7. This CHO restoration resulted in similar resting glycogen contents (FAT-adapt 873 +/- 121 vs. HCHO 868 +/- 120 micromol glucosyl units/g dry wt). However, the respiratory exchange ratio was lower during cycling at 70% VO2peak in the FAT-adapt trial, which resulted in an approximately 45% increase and an approximately 30% decrease in fat and CHO oxidation, respectively. PDH activity was lower at rest and throughout exercise at 70% VO2peak (1.69 +/- 0.25 vs. 2.39 +/- 0.19 mmol x kg wet wt(-1) x min(-1)) and the 1-min sprint in the FAT-adapt vs. the HCHO trial. Estimates of glycogenolysis during the 1st min of exercise at 70% VO2peak and the 1-min sprint were also lower after FAT-adapt (9.1 +/- 1.1 vs. 13.4 +/- 2.1 and 37.3 +/- 5.1 vs. 50.5 +/- 2.7 glucosyl units x kg dry wt(-1) x min(-1)). HSL activity was approximately 20% higher (P = 0.12) during exercise at 70% VO2peak after FAT-adapt. Results indicate that previously reported decreases in whole body CHO oxidation and increases in fat oxidation after the FAT-adapt protocol are a function of metabolic changes within skeletal muscle. The metabolic signals responsible for the shift in muscle substrate use during cycling at 70% VO2peak remain unclear, but lower accumulation of free ADP and AMP after the FAT-adapt trial may be responsible for the decreased glycogenolysis and PDH activation during sprinting.     

Metabolic adaptations to training precede changes in muscle mitochondrial capacity.

To determine whether increases in muscle mitochondrial capacity are necessary for the characteristic lower exercise glycogen loss and lactate concentration observed during exercise in the trained state, we have employed a short-term training model involving 2 h of cycling per day at 67% maximal O2 uptake (VO2max) for 5-7 consecutive days. Before and after training, biopsies were extracted from the vastus lateralis of nine male subjects during a continuous exercise challenge consisting of 30 min of work at 67% VO2max followed by 30 min at 76% VO2max. Analysis of samples at 0, 15, 20, and 60 min indicated a pronounced reduction (P less than 0.05) in glycogen utilization after training. Reductions in glycogen utilization were accompanied by reductions (P less than 0.05) in muscle lactate concentration (mmol/kg dry wt) at 15 min [37.4 +/- 9.3 (SE) vs. 20.2 +/- 5.3], 30 min (30.5 +/- 6.9 vs. 17.6 +/- 3.8), and 60 min (26.5 +/- 5.8 vs. 17.8 +/- 3.5) of exercise. Maximal aerobic power, VO2max (l/min) was unaffected by the training (3.99 +/- 0.21 vs. 4.05 +/- 0.26). Measurements of maximal activities of enzymes representative of the citric acid cycle (succinic dehydrogenase and citrate synthase) were similar before and after the training. It is concluded that, in the voluntary exercising human, altered metabolic events are an early adaptive response to training and need not be accompanied by changes in muscle mitochondrial capacity.     

Carbohydrate metabolism in human skeletal muscle during exercise is not regulated by G-1,6-P2

Glucose 1,6-bisphosphate (G-1,6-P2) is a potent activator of phosphofructokinase (PFK) and an inhibitor of hexokinase in vitro. It has been suggested that increases in G-1,6-P2 are a main means by which PFK can achieve significant catalytic function in vivo despite falling pH and that increases in G-1,6-P2 will inhibit hexokinase in vivo. The purpose of the present study was to determine whether contraction-induced changes in flux through PFK and hexokinase are associated with changes in G-1,6-P2 in skeletal muscle. Ten men performed bicycle exercise for 10 min at 40 and 75% of maximal O2 uptake (VO2max) and to fatigue [4.8 +/- 0.6 (SE) min] at 100% VO2max. Biopsies were obtained from the quadriceps femoris muscle at rest and after each work load and analyzed for G-1,6-P2. G-1,6-P2 averaged 111 +/- 13 mumol/kg dry wt at rest and 121 +/- 16, 123 +/- 15, and 123 +/- 11 mumol/kg dry wt after the low-, moderate-, and high-intensity exercise bouts, respectively (P less than 0.05 for all means vs. rest). Flux through PFK was estimated to increase exponentially as the exercise intensity increased and muscle pH decreased at the higher work loads, whereas flux through hexokinase was estimated to increase during exercise at 40 and 75% VO2max but decrease sharply at 100% VO2max. These data demonstrate that flux through neither PFK nor hexokinase is mediated by changes in G-1,6-P2 in human skeletal muscle during short-term dynamic exercise.     

Morning-to-evening differences in oxygen uptake kinetics in short-duration cycling exercise

This study analyzed diurnal variations in oxygen (O(2)) uptake kinetics and efficiency during a moderate cycle ergometer exercise. Fourteen physically active diurnally active male subjects (age 23+/-5 yrs) not specifically trained at cycling first completed a test to determine their ventilatory threshold (T(vent)) and maximal oxygen consumption (VO(2max)); one week later, they completed four bouts of testing in the morning and evening in a random order, each separated by at least 24 h. For each period of the day (07:00-08:30 h and 19:00-20:30 h), subjects performed two bouts. Each bout was composed of a 5 min cycling exercise at 45 W, followed after 5 min rest by a 10 min cycling exercise at 80% of the power output associated with T(vent). Gas exchanges were analyzed breath-by-breath and fitted using a mono-exponential function. During moderate exercise, the time constant and amplitude of VO(2) kinetics were significantly higher in the morning compared to the evening. The net efficiency increased from the morning to evening (17.3+/-4 vs. 20.5+/-2%; p<0.05), and the variability of cycling cadence was greater during the morning than evening (+34%; p<0.05). These findings suggest that VO(2) responses are affected by the time of day and could be related to variability in muscle activity pattern.