Effects of PDH activation by dichloroacetate in human skeletal muscle during exercise in hypoxia.

During the onset of exercise in hypoxia, the increased lactate accumulation is associated with a delayed activation of pyruvate dehydrogenase (PDH; Parolin ML, Spreit LL, Hultman E, Hollidge-Horvat MG, Jones NL, and Heigenhauser GJF. Am J Physiol Endocrinol Metab 278: E522-E534, 2000). The present study investigated whether activation of PDH with dichloroacetate (DCA) before exercise would reduce lactate accumulation during exercise in acute hypoxia by increasing oxidative phosphorylation. Six subjects cycled on two occasions for 15 min at 55% of their normoxic maximal oxygen uptake after a saline (control) or DCA infusion while breathing 11% O(2). Muscle biopsies of the vastus lateralis were taken at rest and after 1 and 15 min of exercise. DCA increased PDH activity at rest and at 1 min of exercise, resulting in increased acetyl-CoA concentration and acetylcarnitine concentration at rest and at 1 min. In the first minute of exercise, there was a trend toward a lower phosphocreatine (PCr) breakdown with DCA compared with control. Glycogenolysis was lower with DCA, resulting in reduced lactate concentration ([lactate]), despite similar phosphorylase a mole fractions and posttransformational regulators. During the subsequent 14 min of exercise, PDH activity was similar, whereas PCr breakdown and muscle [lactate] were reduced with DCA. Glycogenolysis was lower with DCA, despite similar mole fractions of phosphorylase a, and was due to reduced posttransformational regulators. The results from the present study support the hypothesis that lactate production is due in part to metabolic inertia and cannot solely be explained by an oxygen limitation, even under conditions of acute hypoxia.

Regulation of glycogen phosphorylase and PDH during exercise in human skeletal muscle during hypoxia.

The present study examined the acute effects of hypoxia on the regulation of skeletal muscle metabolism at rest and during 15 min of submaximal exercise. Subjects exercised on two occasions for 15 min at 55% of their normoxic maximal oxygen uptake while breathing 11% O(2) (hypoxia) or room air (normoxia). Muscle biopsies were taken at rest and after 1 and 15 min of exercise. At rest, no effects on muscle metabolism were observed in response to hypoxia. In the 1st min of exercise, glycogenolysis was significantly greater in hypoxia compared with normoxia. This small difference in glycogenolysis was associated with a tendency toward a greater concentration of substrate, free P(i), in hypoxia compared with normoxia. Pyruvate dehydrogenase activity (PDH(a)) was lower in hypoxia at 1 min compared with normoxia, resulting in a reduced rate of pyruvate oxidation and a greater lactate accumulation. During the last 14 min of exercise, glycogenolysis was greater in hypoxia despite a lower mole fraction of phosphorylase a. The greater glycogenolytic rate was maintained posttransformationally through significantly higher free [AMP] and [P(i)]. At the end of exercise, PDH(a) was greater in hypoxia compared with normoxia, contributing to a greater rate of pyruvate oxidation. Because of the higher glycogenolytic rate in hypoxia, the rate of pyruvate production continued to exceed the rate of pyruvate oxidation, resulting in significant lactate accumulation in hypoxia compared with no further lactate accumulation in normoxia. Hence, the elevated lactate production associated with hypoxia at the same absolute workload could in part be explained by the effects of hypoxia on the activities of the rate-limiting enzymes, phosphorylase and PDH, which regulate the rates of pyruvate production and pyruvate oxidation, respectively.

Effect of induced metabolic alkalosis on human skeletal muscle metabolism during exercise.

The purpose of the study was to examine the roles of active pyruvate dehydrogenase (PDH(a)), glycogen phosphorylase (Phos), and their regulators in lactate (Lac(-)) metabolism during incremental exercise after ingestion of 0.3 g/kg of either NaHCO(3) [metabolic alkalosis (ALK)] or CaCO(3) [control (CON)]. Subjects (n = 8) were studied at rest, rest postingestion, and during constant rate cycling at three stages (15 min each): 30, 60, 75% of maximal O(2) uptake (VO(2 max)). Radial artery and femoral venous blood samples, leg blood flow, and biopsies of the vastus lateralis were obtained during each power output. ALK resulted in significantly (P < 0.05) higher intramuscular Lac(-) concentration ([Lac(-)]; ALK 72.8 vs. CON 65.2 mmol/kg dry wt), arterial whole blood [Lac(-)] (ALK 8.7 vs. CON 7.0 mmol/l), and leg Lac(-) efflux (ALK 10.0 vs. CON 4.2 mmol/min) at 75% VO(2 max). The increased intramuscular [Lac(-)] resulted from increased pyruvate production due to stimulation of glycogenolysis at the level of Phos a and phosphofructokinase due to allosteric regulation mediated by increased free ADP (ADP(f)), free AMP (AMP(f)), and free P(i) concentrations. PDH(a) increased with ALK at 60% VO(2 max) but was similar to CON at 75% VO(2 max). The increased PDH(a) may have resulted from alterations in the acetyl-CoA, ADP(f), pyruvate, NADH, and H(+) concentrations leading to a lower relative activity of PDH kinase, whereas the similar values at 75% VO(2 max) may have reflected maximal activation. The results demonstrate that imposed metabolic alkalosis in skeletal muscle results in acceleration of glycogenolysis at the level of Phos relative to maximal PDH activation, resulting in a mismatch between the rates of pyruvate production and oxidation resulting in an increase in Lac(-) production.

Effect of induced metabolic acidosis on human skeletal muscle metabolism during exercise.

The roles of pyruvate dehydrogenase (PDH), glycogen phosphorylase (Phos), and their regulators in lactate (Lac(-)) metabolism were examined during incremental exercise after ingestion of 0.3 g/kg of either NH(4)Cl [metabolic acidosis (ACID)] or CaCO(3) [control (CON)]. Subjects were studied at rest, at rest postingestion, and during continuous steady-state cycling at three stages (15 min each): 30, 60, and 75% of maximal oxygen uptake. Radial artery and femoral venous blood samples, leg blood flow, and biopsies of the vastus lateralis were obtained during each power output. ACID resulted in significantly lower intramuscular concentration of [Lac(-)] (ACID 40.8 vs. CON 56.9 mmol/kg dry wt), arterial whole blood [Lac(-)] (ACID 4.7 vs. CON 6.5 mmol/l), and leg Lac(-) efflux (ACID 3.05 vs. CON 6.98 mmol. l(-1). min(-1)). The reduced intramuscular [Lac(-)] resulted from decreases in pyruvate production due to inhibition of glycogenolysis, at the level of Phos a, and phosphofructokinase, together with an increase in the amount of pyruvate oxidized relative to the total produced. The reduction in Phos a activity was mediated through decreases in transformation, decreases in free inorganic phosphate concentration, and decreases in the posttransformational allosteric regulator free AMP. Reduced PDH activity occurred with ACID and may have resulted from alterations in the concentrations of acetyl-CoA, free ADP, pyruvate, NADH, and H(+), leading to greater relative activity of the kinase. The results demonstrate that imposed metabolic acidosis in skeletal muscle results in decreased Lac(-) production due to inhibition of glycogenolysis at the level of Phos and increased pyruvate oxidation at PDH.

Effects of dichloroacetate infusion on human skeletal muscle metabolism at the onset of exercise.

This study investigated whether dichloroacetate (DCA) decreases the reliance on substrate level phosphorylation during the transition from rest to moderate-intensity exercise in humans. Nine subjects cycled at approximately 65% of maximal oxygen uptake (VO(2 max)) after a saline or DCA (100 mg/kg body wt) infusion, with muscle biopsies taken at rest and at 30 s and 2 and 10 min of exercise. DCA infusion increased pyruvate dehydrogenase (PDH) activation at rest (4.0 +/- 0.3 vs. 0.9 +/- 0.1 mmol. kg wet wt(-1). min(-1)) and at 30 s (3.6 +/- 0.2 vs. 2.5 +/- 0.4 mmol. kg(-1). min(-1)) of exercise. As a result, acetyl-CoA (45.9 +/- 5.9 vs. 11.3 +/- 1.5 micromol/kg dry wt) and acetylcarnitine (13.1 +/- 1.0 vs. 1.6 +/- 0.3 mmol/kg dry wt) were markedly increased by DCA infusion at rest. These differences were maintained at 30 s and 2 min for both acetyl-CoA and acetylcarnitine. Resting muscle lactate and phosphocreatine (PCr) were not different between trials, but DCA infusion resulted in lower lactate accumulation throughout exercise, especially at 2 min (21.6 +/- 3.1 vs. 44.6 +/- 8.0 mmol/kg dry wt). PCr utilization in the initial 30 s (16.9 +/- 0.4 vs. 31.7 +/- 2.6 mmol/kg dry wt) and 2 min (27.8 +/- 4.7 vs. 45.1 +/- 2.6 mmol/kg dry wt) of exercise was decreased with DCA. This resulted in a lower accumulation of free inorganic phosphate at 30 s (25.4 +/- 2.0 vs. 36.4 +/- 2.8 mmol/kg dry wt) and 2 min (34.6 +/- 4.7 vs. 50.5 +/- 2.2 mmol/kg dry wt) with DCA and decreased glycogenolysis over 10 min. The data from this study support the hypothesis that increased provision of substrate by DCA infusion increases oxidative metabolism during the rest-to-work transition, resulting in decreased PCr utilization and an improved cellular energy state at the onset of exercise. The transitory improvement in energy state decreased glycogenolysis and lactate accumulation during moderate-intensity exercise.

Effects of short-term submaximal training in humans on muscle metabolism in exercise.

Muscle metabolism, including the role of pyruvate dehydrogenase (PDH) in muscle lactate (Lac-) production, was examined during incremental exercise before and after 7 days of submaximal training on a cycle ergometer [2 h daily at 60% peak O2 uptake (VO2 max)]. Subjects were studied at rest and during continuous steady-state cycling at three stages (15 min each): 30, 65, and 75% of the pretraining VO2 max. Blood was sampled from brachial artery and femoral vein, and leg blood flow was measured by thermodilution. Biopsies of the vastus lateralis were obtained at rest and during steady-state exercise at the end of each stage. VO2 max, leg O2 uptake, and the maximum activities of citrate synthase and PDH were not altered by training; muscle glycogen concentration was higher. During rest and cycling at 30% VO2 max, muscle Lac- concentration ([Lac-]) and leg efflux were similar. At 65% VO2 max, muscle [Lac-] was lower (11.9 +/- 3.2 vs. 20.0 +/- 5.8 mmol/kg dry wt) and Lac- efflux was less [-0.22 +/- 0.24 (one leg) vs. 1.42 +/- 0.33 mmol/min] after training. Similarly, at 75% VO2 max, lower muscle [Lac-] (17.2 +/- 4.4 vs. 45.2 +/- 6.6 mmol/kg dry wt) accompanied less release (0.41 +/- 0.53 vs. 1.32 +/- 0.65 mmol/min) after training. PDH in its active form (PDHa) was not different between conditions. Calculated pyruvate production at 75% VO2 max fell by 33%, pyruvate reduction to lactate fell by 59%, and pyruvate oxidation fell by 24% compared with before training. Muscle contents of coenzyme A and phosphocreatine were higher during exercise after training. Lower muscle lactate production after training resulted from improved matching of glycolytic and PDHa fluxes, independently of changes in muscle O2 consumption, and was associated with greater phosphorylation potential.

Effects of increased fat availability on fat-carbohydrate interaction during prolonged exercise in men.

The study examined the existence and regulation of fat-carbohydrate interaction during low- and moderate-intensity exercise. Eight males cycled for 10 min at 40% and 60 min at 65% maximal O2 uptake (VO2max) while infused with either Intralipid and heparin (Int) or saline (Con). Before exercise, plasma arterial free fatty acid (FFA) was 0.69 +/- 0.04 mM (Int) vs. 0.25 +/- 0.04 mM (Con). Muscle biopsies were taken at rest and at 10, 20, and 70 min of exercise. Arterial and femoral venous blood samples and expired gases were collected simultaneously throughout exercise, and blood flow was estimated from pulmonary O2 uptake and the leg arterial-venous O2 difference. Respiratory exchange ratio was higher in Con (0.94 +/- 0.01) compared with Int (0.91 +/- 0.01). Mean net leg FFA uptake was higher in Int (0.16 +/- 0.03 vs. 0.04 +/- 0.01 mmol/min), and net lactate efflux was reduced (Int, 1.55 +/- 0.36 vs. Con, 3.07 +/- 0.47 mmol/min). Leg net glucose uptake was unaffected by Int. Muscle glycogen degradation was 23% lower in Int [230 +/- 29 vs. 297 +/- 36 mmol glucosyl units/kg dry muscle (dm)]. Pyruvate dehydrogenase activity in the a form (PDHa) was lower during Int (1.61 +/- 0.17 vs. 2.22 +/- 0.24 mmol.min-1.kg wet muscle-1), and muscle citrate was higher (0.59 +/- 0.04 vs. 0.48 +/- 0.04 mmol/kg dm). Muscle lactate, phosphocreatine, ATP, acetyl-CoA, acetyl-carnitine, and P(i) were unaffected by Int. Calculated free AMP was significantly lower in Int compared with Con at 70 min of exercise (3.3 +/- 0.8 vs. 1.5 +/- 0.3 mumol/kg dm). The high FFA-induced reduction in glycogenolysis and carbohydrate oxidation at 65% VO2max appears to be due to regulation at several sites. The reduced flux through phosphorylase and phosphofructokinase during Int may have been due to reduced free AMP accumulation and increased cytoplasmic citrate. The mechanism for reduced PDH transformation to the a form is unknown but suggests reduced flux through PDH.

Skeletal muscle pyruvate dehydrogenase activity during maximal exercise in humans.

The regulation of the active form of pyruvate dehydrogenase (PDHa) and related metabolic events were examined in human skeletal muscle during repeated bouts of maximum exercise. Seven subjects completed three consecutive 30-s bouts of maximum isokinetic cycling, separated by 4 min of recovery. Biopsies of the vastus lateralis were taken before and immediately after each bout. PDHa increased from 0.45 +/- 0.15 to 2.96 +/- 0.38, 1.10 +/- 0.11 to 2.91 +/- 0.11, and 1.28 +/- 0.18 to 2.82 +/- 0.32 mmol.min-1.kg wet wt-1 during bouts 1, 2, and 3, respectively. Glycolytic flux was 13-fold greater than PDHa in bouts 1 and 2 and 4-fold greater during bout 3. This discrepancy between the rate of pyruvate production and oxidation resulted in substantial lactate accumulation to 89.5 +/- 11.6 in bout 1, 130.8 +/- 13.8 in bout 2, and 106.6 +/- 10.1 mmol/kg dry wt in bout 3. These events coincided with an increase in the mitochondrial oxidation state, as reflected by a fall in mitochondrial NADH/NAD, indicating that muscle lactate production during exercise was not an O2-dependent process in our subjects. During exercise the primary factor regulating PDHa transformation was probably intracellular Ca2+. In contrast, the primary regulatory factors causing greater PDHa during recovery were lower ATP/ADP and NADH/NAD and increased concentrations of pyruvate and H+. Greater PDHa during recovery facilitated continued oxidation of the lactate load between exercise bouts.