Beetroot juice supplementation reduces whole body oxygen consumption but does not improve indices of mitochondrial efficiency in human skeletal muscle.

KEY POINTS: Oral consumption of nitrate (NO3(-)) in beetroot juice has been shown to decrease the oxygen cost of submaximal exercise; however, the mechanism of action remains unresolved. We supplemented recreationally active males with beetroot juice to determine if this altered mitochondrial bioenergetics. Despite reduced submaximal exercise oxygen consumption, measures of mitochondrial coupling and respiratory efficiency were not altered in muscle. In contrast, rates of mitochondrial hydrogen peroxide (H2O2) emission were increased in the absence of markers of lipid or protein oxidative damage. These results suggest that improvements in mitochondrial oxidative metabolism are not the cause of beetroot juice-mediated improvements in whole body oxygen consumption. ABSTRACT: Ingestion of sodium nitrate (NO3(-)) simultaneously reduces whole body oxygen consumption (V̇O2) during submaximal exercise while improving mitochondrial efficiency, suggesting a causal link. Consumption of beetroot juice (BRJ) elicits similar decreases in V̇O2 but potential effects on the mitochondria remain unknown. Therefore we examined the effects of 7-day supplementation with BRJ (280 ml day(-1), ∼26 mmol NO3(-)) in young active males (n = 10) who had muscle biopsies taken before and after supplementation for assessments of mitochondrial bioenergetics. Subjects performed 20 min of cycling (10 min at 50% and 70% V̇O2 peak) 48 h before 'Pre' (baseline) and 'Post' (day 5 of supplementation) biopsies. Whole body V̇O2 decreased (P < 0.05) by ∼3% at 70% V̇O2 peak following supplementation. Mitochondrial respiration in permeabilized muscle fibres showed no change in leak respiration, the content of proteins associated with uncoupling (UCP3, ANT1, ANT2), maximal substrate-supported respiration, or ADP sensitivity (apparent Km). In addition, isolated subsarcolemmal and intermyofibrillar mitochondria showed unaltered assessments of mitochondrial efficiency, including ADP consumed/oxygen consumed (P/O ratio), respiratory control ratios and membrane potential determined fluorometrically using Safranine-O. In contrast, rates of mitochondrial hydrogen peroxide (H2O2) emission were increased following BRJ. Therefore, in contrast to sodium nitrate, BRJ supplementation does not alter key parameters of mitochondrial efficiency. This occurred despite a decrease in exercise V̇O2, suggesting that the ergogenic effects of BRJ ingestion are not due to a change in mitochondrial coupling or efficiency. It remains to be determined if increased mitochondrial H2O2 contributes to this response.

Omega-3 supplementation alters mitochondrial membrane composition and respiration kinetics in human skeletal muscle.

Studies have shown increased incorporation of omega-3 fatty acids into whole skeletal muscle following supplementation, although little has been done to investigate the potential impact on the fatty acid composition of mitochondrial membranes and the functional consequences on mitochondrial bioenergetics. Therefore, we supplemented young healthy male subjects (n = 18) with fish oils [2 g eicosapentaenoic acid (EPA) and 1 g docosahexanoic acid (DHA) per day] for 12 weeks and skeletal muscle biopsies were taken prior to (Pre) and following (Post) supplementation for the analysis of mitochondrial membrane phospholipid composition and various assessments of mitochondrial bioenergetics. Total EPA and DHA content in mitochondrial membranes increased (P < 0.05) ∼450 and ∼320%, respectively, and displaced some omega-6 species in several phospholipid populations. Mitochondrial respiration, determined in permeabilized muscle fibres, demonstrated no change in maximal substrate-supported respiration, or in the sensitivity (apparent Km) and maximal capacity for pyruvate-supported respiration. In contrast, mitochondrial responses during ADP titrations demonstrated an enhanced ADP sensitivity (decreased apparent Km) that was independent of the creatine kinase shuttle. As the content of ANT1, ANT2, and subunits of the electron transport chain were unaltered by supplementation, these data suggest that prolonged omega-3 intake improves ADP kinetics in human skeletal muscle mitochondria through alterations in membrane structure and/or post-translational modification of ATP synthase and ANT isoforms. Omega-3 supplementation also increased the capacity for mitochondrial reactive oxygen species emission without altering the content of oxidative products, suggesting the absence of oxidative damage. The current data strongly emphasize a role for omega-3s in reorganizing the composition of mitochondrial membranes while promoting improvements in ADP sensitivity.

Effect of age and gender on sweat lactate and ammonia concentrations during exercise in the heat.

The dependence of sweat composition and acidity on sweating rate (SR) suggests that the lower SR in children compared to adults may be accompanied by a higher level of sweat lactate (Lac-) and ammonia (NH3) and a lower sweat pH. Four groups (15 girls, 18 boys, 8 women, 8 men) cycled in the heat (42 degrees C, 20% relative humidity) at 50% VO2max for two 20-min bouts with a 10-min rest before bout 1 and between bouts. Sweat was collected into plastic bags attached to the subject's lower back. During bout 1, sweat from girls and boys had higher Lac- concentrations (23.6 +/- 1.2 and 21.2 +/- 1.7 mM; P < 0.05) than sweat from women and men (18.2 +/- 1.9 and 14.8 +/- 1.6 mM, respectively), but Lac- was weakly associated with SR (P > 0.05; r = -0.27). Sweat Lac- concentration dropped during exercise bout 2, reaching similar levels among all groups (overall mean = 13.7 +/- 0.4 mM). Children had a higher sweat NH3 than adults during bout 1 (girls = 4.2 +/- 0.4, boys = 4.6 +/- 0.6, women = 2.7 +/- 0.2, and men = 3.0 +/- 0.2 mM; P < 0.05). This difference persisted through bout 2 only in females. On average, children's sweat pH was lower than that of adults (mean +/- SEM, girls = 5.4 +/- 0.2, boys = 5.0 +/- 0.1, women = 6.2 +/- 0.5, and men = 6.2 +/- 0.4 for bout 1, and girls = 5.4 +/- 0.2, boys = 6.5 +/- 0.5, women = 5.2 +/- 0.2, and men = 6.9 +/- 0.4 for bout 2). This may have favored NH3 transport from plasma to sweat as accounted for by a significant correlation between sweat NH3 and H+ (r = 0.56). Blood pH increased from rest (mean +/- SEM; 7.3 +/- 0.02) to the end of exercise (7.4 +/- 0.01) without differences among groups. These results, however, are representative of sweat induced by moderate exercise in the absence of acidosis.

AMP-activated protein kinase is not down-regulated in human skeletal muscle of obese females.

Obesity in humans is associated with lipid accumulation in skeletal muscle, insulin and leptin resistance, and type 2 diabetes. AMP-activated protein kinase (AMPK) is an important regulator of fatty acid (FA) metabolism in skeletal muscle. To address the hypothesis that lipid accumulation in skeletal muscle of obese subjects may be due to down-regulation of AMPK, we measured mRNA and protein levels of AMPK isoforms, AMPKalpha1 and -alpha2 activity, AMPK kinase activity, acetyl-coenzyme A carboxylase (ACCbeta) expression and phosphorylation, and FA metabolism in biopsies of rectus abdominus muscle from lean and obese women. We also examined the effect of 5-aminoimidazole-4-carboxamide riboside (AICAR) on AMPK activity and the effects of AICAR and leptin on FA metabolism. Skeletal muscle of obese subjects had increased total FA uptake and triglyceride esterification, and leptin failed to stimulate FA oxidation. However, AMPK mRNA and protein expression, AMPKalpha1 and -alpha2 activities, AMPK kinase activity, ACCbeta phosphorylation, and FA oxidation were similar in lean and obese subjects. Moreover, AICAR increased AMPKalpha2 activity, ACCbeta phosphorylation, and palmitate oxidation to a similar degree in muscle from lean and obese subjects. We conclude that the abnormal lipid metabolism and leptin resistance of skeletal muscle of obese subjects is not due to down-regulation of AMPK. In addition, the similar stimulation by AICAR of AMPK in skeletal muscle of lean and obese subjects suggests that direct pharmacological activation of AMPK may be a therapeutic approach for stimulating FA oxidation in the treatment of human obesity.

Effect of high-fat and high-carbohydrate diets on pulmonary O2 uptake kinetics during the transition to moderate-intensity exercise.

Mitochondrial pyruvate dehydrogenase (PDH) regulates the delivery of carbohydrate-derived substrate to the mitochondrial tricarboxylic acid cycle and electron transport chain. PDH activity at rest and its activation during exercise is attenuated following high-fat (HFAT) compared with high-carbohydrate (HCHO) diets. Given the reliance on carbohydrate-derived substrate early in transitions to exercise, this study examined the effects of HFAT and HCHO on phase II pulmonary O2 uptake (V̇o2 p) kinetics during transitions into the moderate-intensity (MOD) exercise domain. Eight active adult men underwent dietary manipulations consisting of 6 days of HFAT (73% fat, 22% protein, 5% carbohydrate) followed immediately by 6 days of HCHO (10% fat, 10% protein, 80% carbohydrate); each dietary phase was preceded by a glycogen depletion protocol. Participants performed three MOD transitions from a 20 W cycling baseline to work rate equivalent to 80% of estimated lactate threshold on days 5 and 6 of each diet. Steady-state V̇o2 p was greater (P < 0.05), and respiratory exchange ratio and carbohydrate oxidation rates were lower (P < 0.05) during HFAT. The phase II V̇o2 p time constant (τV̇o2 p) [HFAT 40 ± 16, HCHO 32 ± 19 s (mean ± SD)] and V̇o2 p gain (HFAT 10.3 ± 0.8, HCHO 9.4 ± 0.7 ml·min(-1·)W(-1)) were greater (P < 0.05) in HFAT. The overall adjustment (effective time constant) of muscle deoxygenation (Δ[HHb]) was not different between diets (HFAT 24 ± 4 s, HCHO 23 ± 4 s), which coupled with a slower τV̇o2 p, indicates a slowed microvascular blood flow response. These results suggest that the slower V̇o2 p kinetics associated with HFAT are consistent with inhibition and slower activation of PDH, a lower rate of pyruvate production, and/or attenuated microvascular blood flow and O2 delivery.

O2 uptake kinetics, pyruvate dehydrogenase activity, and muscle deoxygenation in young and older adults during the transition to moderate-intensity exercise.

The adaptation of pulmonary O(2) uptake (Vo(2)(p)) kinetics is slowed in older compared with young adults during the transition to moderate-intensity exercise. In this study, we examined the relationship between Vo(2)(p) kinetics and mitochondrial pyruvate dehydrogenase (PDH) activity in young (n = 7) and older (n = 6) adults. Subjects performed cycle exercise to a work rate corresponding to approximately 90% of estimated lactate threshold. Phase 2 Vo(2)(p) kinetics were slower (P < 0.05) in older (tau = 40 +/- 17 s) compared with young (tau = 21 +/- 6 s) adults. Relative phosphocreatine (PCr) breakdown was greater (P < 0.05) at 30 s in older compared with young adults. Absolute PCr breakdown at 6 min was greater (P < 0.05) in older compared with young adults. In young adults, PDH activity increased (P < 0.05) from baseline to 30 s, with no further change observed at 6 min. In older adults, PDH activity during baseline exercise was similar to that seen in young adults. During the exercise transition, PDH activity did not increase (P > 0.05) at 30 s of exercise but was elevated (P < 0.05) after 6 min. The change in deoxyhemoglobin (HHb) was greater for a given Vo(2)(p) in older adults, and there was a similar time course of HHb accompanying the slower Vo(2)(p) kinetics in the older adults, suggesting a slower adaptation of bulk O(2) delivery in older adults. In conclusion, the slower adaptation of Vo(2)(p) in older adults is likely a result of both an increased metabolic inertia and lower O(2) availability.

The role of adipokines as regulators of skeletal muscle fatty acid metabolism and insulin sensitivity.

Several adipose-derived cytokines (adipokines) have been suggested to act as a link between accumulated fat mass and altered insulin sensitivity. Resistin and tumour necrosis factor-alpha (TNF-alpha) have been implicated in impairing insulin sensitivity in rodents; conversely, two other adipokines, leptin and adiponectin, increase insulin sensitivity in lean and obese rodents. Currently, there is considerable focus on the concept that lipid accumulation in skeletal muscle leads to the development of insulin resistance. Adiponectin and leptin have each been demonstrated to increase rates of fatty acid (FA) oxidation and decrease muscle lipid content, which may in part be the underlying mechanism to their insulin sensitizing effect. These effects on FA metabolism appear to be mediated in part through the activation of AMP-activated protein kinase. Evidence derived from animal and human studies suggests that the ability of leptin and adiponectin to stimulate FA oxidation in muscle is impaired in the obese condition. Thus, leptin and adiponectin resistance may be an initiating factor in the accumulation of intramuscular lipids, such as diacylglycerol and ceramide, and the ensuing development of insulin resistance. Lifestyle factors such as diet and exercise are able to restore the sensitivity of muscle to leptin. The actual physiological roles of resistin and TNF-alpha in altering muscle lipid metabolism are more controversial, but each has been shown to directly impair insulin signalling and consequently, insulin stimulated glucose uptake in muscle. However, the possibility that resistin and TNF-alpha reduces insulin sensitivity in muscle by directly impairing FA metabolism in this tissue leading to lipid accumulation, has been virtually unexamined. Thus, the contribution of various adipokines to the development of insulin resistance is complex and not fully understood. Finally, the effects of these adipokines on metabolism and insulin sensitivity are generally studied in isolation, making it difficult to predict the interactive effects and the net impact on insulin sensitivity.

The fatty acid transporter FAT/CD36 is upregulated in subcutaneous and visceral adipose tissues in human obesity and type 2 diabetes.

BACKGROUND: Long-chain fatty acids (LCFAs) cross the plasma membrane via a protein-mediated mechanism involving one or more LCFA-binding proteins. Among these, FAT/CD36 has been identified as key LCFA transporter in the heart and skeletal muscle, where it is regulated acutely and chronically by insulin. In skeletal muscle, FAT/CD36 expression and/or subcellular distribution is altered in obesity and type 2 diabetes. There is limited information as to whether the expression of this protein is also altered in subcutaneous and/or visceral adipose tissue depots in human obesity or type 2 diabetes. OBJECTIVES: To compare (a) the expression of FAT/CD36 in subcutaneous and visceral adipose tissue depots in lean, overweight, and obese individuals and in type 2 diabetics, (b) to determine whether the protein expression of FAT/CD36 in these depots is associated with the severity of insulin resistance (type 2 diabetes>obese>overweight/lean) and (c) whether FAT/CD36 protein expression in these adipose tissue depots is associated with alterations in circulating substrates and hormones. SUBJECTS: Subjects who were undergoing abdominal surgery and who were lean (n=10; three men, seven women), overweight (n=10; three men, seven women) or obese (n=7; one man, six women), or who had been diagnosed with type 2 diabetes (n=5; one man, four women) participated in this study. MEASUREMENTS: Subcutaneous and visceral adipose tissue samples, as well as blood samples, were obtained from the subjects while under general anesthesia. Adipose tissue samples were analyzed for FAT/CD36 using Western blotting. Serum samples were analyzed for glucose, insulin, FFA and leptin. BMI was also calculated. RESULTS: Subcutaneous adipose tissue FAT/CD36 expression was upregulated by +58, +76 and +150% in overweight, obese and type 2 diabetics, respectively. Relative to subcutaneous adipose tissue, visceral adipose tissue FAT/CD36 expression was upregulated in lean (+52%) and overweight subjects (+30%). In contrast, in obese subjects and type 2 diabetics, no difference in FAT/CD36 protein expression was observed between their subcutaneous and visceral adipose tissue depots (P>0.05). The subcutaneous adipose tissue FAT/CD36 expression (R=0.85) and the visceral adipose tissue FAT/CD36 expression (R=0.77) were associated with alteration in BMI and circulating glucose and insulin. CONCLUSIONS: Subcutaneous adipose tissue FAT/CD36 expression is upregulated in obesity and type 2 diabetes. As FAT/CD36 expression is not different in lean, overweight and obese subjects, and was only increased in type 2 diabetics, it appears that visceral adipose tissue FAT/CD36 may respond in a less dynamic manner to metabolic disturbances than subcutaneous adipose tissue FAT/CD36.

Prior heavy exercise elevates pyruvate dehydrogenase activity and speeds O2 uptake kinetics during subsequent moderate-intensity exercise in healthy young adults.

The adaptation of pulmonary oxygen uptake (.VO2) during the transition to moderate-intensity exercise (Mod) is faster following a prior bout of heavy-intensity exercise. In the present study we examined the activation of pyruvate dehydrogenase (PDHa) during Mod both with and without prior heavy-intensity exercise. Subjects (n = 9) performed a Mod(1)-heavy-intensity-Mod(2) exercise protocol preceded by 20 W baseline. Breath-by-breath .VO2 kinetics and near-infrared spectroscopy-derived muscle oxygenation were measured continuously, and muscle biopsy samples were taken at specific times during the transition to Mod. In Mod(1), PDHa increased from baseline (1.08 +/- 0.2 mmol min(-1) (kg wet wt)(-1)) to 30 s (2.05 +/- 0.2 mmol min(-1) (kg wet wt)(-1)), with no additional change at 6 min exercise (2.07 +/- 0.3 mmol min(-1) (kg wet wt)(-1)). In Mod(2), PDHa was already elevated at baseline (1.88 +/- 0.3 mmol min(-1) (kg wet wt)(-1)) and was greater than in Mod(1), and did not change at 30 s (1.96 +/- 0.2 mmol min(-1) (kg wet wt)(-1)) but increased at 6 min exercise (2.70 +/- 0.3 mmol min(-1) (kg wet wt)(-1)). The time constant of .VO2 was lower in Mod(2) (19 +/- 2 s) than Mod(1) (24 +/- 3 s). Phosphocreatine (PCr) breakdown from baseline to 30 s was greater (P < 0.05) in Mod(1) (13.6 +/- 6.7 mmol (kg dry wt)(-1)) than Mod(2) (6.5 +/- 6.2 mmol (kg dry wt)(-1)) but total PCr breakdown was similar between conditions (Mod(1), 14.8 +/- 7.4 mmol (kg dry wt)(-1); Mod(2), 20.1 +/- 8.0 mmol (kg dry wt)(-1)). Both oxyhaemoglobin and total haemoglobin were elevated prior to and throughout Mod(2) compared with Mod(1). In conclusion, the greater PDHa at baseline prior to Mod(2) compared with Mod(1) may have contributed in part to the faster .VO2 kinetics in Mod(2). That oxyhaemoglobin and total haemoglobin were elevated prior to Mod(2) suggests that greater muscle perfusion may also have contributed to the observed faster .VO2 kinetics. These findings are consistent with metabolic inertia, via delayed activation of PDH, in part limiting the adaptation of pulmonary .VO2 and muscle O2 consumption during the normal transition to exercise.

Exercise and recovery metabolism in the Pacific spiny dogfish (Squalus acanthias).

We examined the effects of exhaustive exercise and post-exercise recovery on white muscle substrate depletion and metabolite distribution between white muscle and blood plasma in the Pacific spiny dogfish, both in vivo and in an electrically stimulated perfused tail-trunk preparation. Measurements of arterial-venous lactate, total ammonia, beta-hydroxybutyrate, glucose, and L-alanine concentrations in the perfused tail-trunk assessed white muscle metabolite fluxes. Exhaustive exercise was fuelled primarily by creatine phosphate hydrolysis and glycolysis as indicated by 62, 71, and 85% decreases in ATP, creatine phosphate, and glycogen, respectively. White muscle lactate production during exercise caused a sustained increase (approximately 12 h post-exercise) in plasma lactate load and a short-lived increase (approximately 4 h post-exercise) in plasma metabolic acid load during recovery. Exhaustive exercise and recovery did not affect arterial PO2, PCO2, or PNH3 but the metabolic acidosis caused a decrease in arterial HCO3- immediately after exercise and during the first 8 h recovery. During recovery, lactate was retained in the white muscle at higher concentrations than in the plasma despite increased lactate efflux from the muscle. Pyruvate dehydrogenase activity was very low in dogfish white muscle at rest and during recovery (0.53 +/- 0.15 nmol g wet tissue(-1) min(-1); n=40) indicating that lactate oxidation is not the major fate of lactate during post-exercise recovery. The lack of change in white muscle free-carnitine and variable changes in short-chain fatty acyl-carnitine suggest that dogfish white muscle does not rely on lipid oxidation to fuel exhaustive exercise or recovery. These findings support the notion that extrahepatic tissues cannot utilize fatty acids as an oxidative fuel. Furthermore, our data strongly suggest that ketone body oxidation is important in fuelling recovery metabolism in dogfish white muscle and at least 20% of the ATP required for recovery could be supplied by uptake and oxidation of beta-hydroxybutyrate from the plasma.

Carbohydrate ingestion reduces skeletal muscle acetylcarnitine availability but has no effect on substrate phosphorylation at the onset of exercise in man.

This study investigated the effect of reduced acetylcarnitine availability on oxidative metabolism during the transition from rest to steady-state exercise. Eight male subjects completed two randomised exercise trials at 68 % of the peak rate of O(2) uptake (V((O(2)),peak)). On one occasion subjects ingested 1 g (kg body mass)(-1) glucose 75 min prior to exercise (CHO), whereas the other trial acted as a control (CON). Muscle samples were obtained pre- and 75 min post-ingestion, and following 1 and 10 min of exercise. Plasma glucose and insulin were elevated (P < 0.05), and plasma free fatty acids (FFA) were lower at the onset of exercise in CHO. Acetylcarnitine (CON, 4.8 +/- 1.8; CHO, 1.5 +/- 0.9 mmol (kg dry mass (d.m.))(-1), P < 0.05) and acetyl CoA (CON, 13.2 +/- 2.3; CHO, 6.3 +/- 0.6 micromol (kg d.m.)(-1), P < 0.05) were lower at rest, whereas pyruvate dehydrogenase activation (PDHa) was greater in CHO compared with CON (CON, 0.78 +/- 0.07; CHO, 1.44 +/- 0.19 mmol min(-1) (kg wet mass (w.m.))(-1)). Respiratory exchange ratio (RER) was significantly elevated during exercise in CHO. The acetyl groups increased at similar rates at the onset of exercise (1 min) and there was no difference in substrate phosphorylation as determined from lactate accumulation and phosphocreatine degradation between trials. Subsequently, oxidative metabolism during the transition from rest to steady-state exercise was not affected by prior carbohydrate ingestion. Although exercise resulted in the rapid activation of PDH in both trials, PDHa was greater at 1 min in CHO (CON, 2.36 +/- 0.22; CHO, 2.91 +/- 0.18 mmol min(-1) (kg w.m.)(-1)). No differences in muscle metabolite levels and PDHa were observed after 10 min of moderate exercise between trials. In summary, at rest, carbohydrate ingestion induced multiple metabolic changes which included decreased acetylcarnitine availability and small increases in PDHa. The prior changes in PDHa and acetylcarnitine availability had no effect on substrate phosphorylation and oxidative metabolism at the onset of exercise. These data suggest that acetylcarnitine availability is unlikely to be the site of metabolic inertia during the transition from rest to steady-state moderate intensity exercise.

Effects of reduced free fatty acid availability on skeletal muscle PDH activation during aerobic exercise. Pyruvate dehydrogenase.

This study investigated the effect of reduced free fatty acid (FFA) availability on pyruvate dehydrogenase activation (PDHa) and carbohydrate metabolism during moderate aerobic exercise. Eight active male subjects cycled for 40 min at 55% Vo(2 peak) on two occasions. During one trial, subjects ingested 20 mg/kg body mass of the antilipolytic drug nicotinic acid (NA) during the hour before exercise to reduce FFA. Nothing was ingested in the control trial (CON). Blood and expired gas measurements were obtained throughout the trials, and muscle biopsy samples were obtained immediately before exercise and at 5, 20, and 40 min of exercise. Plasma FFA were lower in the NA trial (0.13 +/- 0.01 vs. 0.48 +/- 0.03 mM, P < 0.05), and the respiratory exchange ratio (RER) was increased with NA (0.93 +/- 0.01 vs. 0.89 +/- 0.01, P < 0.05), resulting in a 14.5 +/- 1.8% increase in carbohydrate oxidation compared with CON. PDHa increased rapidly in both trials at exercise onset but was approximately 15% higher (P < 0.05) throughout exercise in the NA trial (2.44 +/- 0.19 and 2.07 +/- 0.12 mmol x kg wet muscle(-1) x min(-1) for NA and CON at 40 min). Muscle glycogenolysis was 15.3 +/- 9.6% greater in the NA trial vs. the CON trial but did not reach statistical significance. Glucose 6-phosphate contents were elevated (P < 0.05) in the NA trial at 30 and 40 min of exercise, but pyruvate and lactate contents were unaffected. These data demonstrate that the reduction of exogenous FFA availability increased the activation of PDH and carbohydrate oxidation during moderate aerobic exercise in men. The increased activation of PDH was not explained by changes in muscle pyruvate or the ATP/ADP ratio but may be related to a decrease in the NADH/NAD(+) ratio or an epinephrine-induced increase in calcium concentration.

Effects of respiratory alkalosis on human skeletal muscle metabolism at the onset of submaximal exercise.

The purpose of this study was to examine the effects of respiratory alkalosis on human skeletal muscle metabolism at rest and during submaximal exercise. Subjects exercised on two occasions for 15 min at 55 % of their maximal oxygen uptake while either hyperventilating (R-Alk) or breathing normally (Con). 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 R-Alk. In the first minute of exercise, there was a delayed activation of pyruvate dehydrogenase (PDH) in R-Alk compared with Con, resulting in a reduced rate of pyruvate oxidation. Also, glycogenolysis was higher in R-Alk compared with Con, which was attributed to a higher availability of the monoprotonated form of inorganic phosphate (P(i)), resulting in an elevated rate of pyruvate production. The mismatch between pyruvate production and its oxidation resulted in net lactate accumulation. These effects were not seen after 15 min of exercise, with no further differences in muscle metabolism between conditions. The results from the present study suggest that respiratory alkalosis may play an important role in lactate accumulation during the transition from rest to exercise in acute hypoxic conditions, but that other factors mediate lactate accumulation during steady-state exercise.

Hormone-sensitive lipase activity and triacylglycerol hydrolysis are decreased in rat soleus muscle by cyclopiazonic acid.

Cyclopiazonic acid (CPA) is a sarcoplasmic reticulum Ca2+-ATPase inhibitor that increases intracellular calcium. The role of CPA in regulating the oxidation and esterification of palmitate, the hydrolysis of intramuscular lipids, and the activation of hormone-sensitive lipase (HSL) was examined in isolated rat soleus muscles at rest. CPA (40 micro M) was added to the incubation medium to levels that resulted in subcontraction increases in muscle tension, and lipid metabolism was monitored using the previously described pulse-chase procedure. CPA did not alter the cellular energy state, as reflected by similar muscle contents of ATP, phosphocreatine, free AMP, and free ADP. CPA increased total palmitate uptake into soleus muscle (11%, P < 0.05) and was without effect on palmitate oxidation. This resulted in greater esterification of exogenous palmitate into the triacylglycerol (18%, P < 0.05) and phospholipid (89%, P < 0.05) pools. CPA decreased (P < 0.05) intramuscular lipid hydrolysis, and this occurred as a result of reduced HSL activity (20%, P < 0.05). Incubation of muscles with 3 mM caffeine, which is also known to increase Ca2+ without affecting the cellular energy state, reduced HSL activity (24%, P < 0.05). KN-93, a calcium/calmodulin-dependent kinase inhibitor (CaMKII), blocked the effects of CPA and caffeine, and HSL activity returned to preincubation values. The results of the present study demonstrate that CPA simultaneously decreases intramuscular triacylglycerol (IMTG) hydrolysis and promotes lipid storage in isolated, intact soleus muscle. The decreased IMTG hydrolysis is likely mediated by reduced HSL activity, possibly via the CaMKII pathway. These responses are not consistent with the increased hydrolysis and decreased esterification observed in contracting muscle when substrate availability and the hormonal milieu are tightly controlled. It is possible that more powerful signals or a higher [Ca2+] may override the lipid-storage effect of the CPA-mediated effects during muscular contractions.