Improving the precision of the accumulated oxygen deficit using VO2-power regression points from below and above the lactate threshold

The accumulated oxygen deficit (AOD) method assumes a linear VO₂-power relationship for exercise intensities increasing from below the lactate threshold (BLT) to above the lactate threshold (ALT). Factors that were likely to effect the linearity of the VO₂-power regression and the precision of the estimated total energy demand (ETED) were investigated. These included the slow component of VO₂ kinetics (SC), a forced resting y-intercept and exercise intensities BLT and ALT. Criteria for linearity and precision included the Pearson correlation coefficient (PCC) of the VO₂-power relationship, the length of the 95% confidence interval (95% CI) of the ETED and the standard error of the predicted value (SEP), respectively. Eight trained male and one trained female triathlete completed the required cycling tests to establish the AOD when pedalling at 80 rev/min. The influence of the SC on the linear extrapolation of the ETED was reduced by measuring VO₂ after three min of exercise. Measuring VO₂ at this time provided a new linear extrapolation method consisting of ten regression points spread evenly from BLT and ALT. This method produced an ETED with increased precision compared to using regression equations developed from intensities BLT with no forced y-intercept value; (95%CI (L), 0.70±0.26 versus 1.85±1.10, P<0.01; SEP(L/Watt), 0.07±0.02 versus 0.28±0.17; P<0.01). Including a forced y-intercept value with five regression points either BLT or ALT increased the precision of estimating the total energy demand to the same level as when using 10 regression points, (5 points BLT + y-intercept versus 5 points ALT + y-intercept versus 10 points; 95%CI(l), 0.61±0.32, 0.87±0.40, 0.70±0.26; SEP(L/Watt), 0.07±0.03, 0.08±0.04, 0.07±0.02; p>0.05). The VO₂-power regression can be designed using a reduced number of regression points... ABSTRACT FROM AUTHOR

Milk lactate determination with a rotating bioreactor based on an electron transfer mediated by osmium complexes incorporating a continuous-flow/stopped-flow system

The high sensitivity that can be attained using a bienzymatic system and mediated by the redox polymer [Os(bpy)₂ClPyCH₂NHpoly(allylamine)] (Os-PAA), has been verified by on-line interfacing of a rotating bioreactor and continuous-flow/stopped-flow/continuous-flow processing. When the hydrogen peroxide formed by LO_x layer reaches the inner layer, the electronic flow between the immobilized peroxidase and the electrode surface produces a current, proportional to lactate concentration. The determination of lactate was possible with a limit of detection of 5 nmol l⁻¹ in the processing of as many as 30 samples per hour. This arrangement allows working in undiluted milk samples with a good stability and reproducibility. Horseradish peroxidase [EC 1.11.1.7] and Os-PAA were covalently immobilized on the glassy carbon electrode surface (upper cell body), lactate oxidase [EC 1.1.3.x] was immobilized on a disk that can be rotated.

Adrenaline increases skeletal muscle glycogenolysis, pyruvate dehydrogenase activation and carbohydrate oxidation during moderate exercise in humans

# 1.
To evaluate the role of adrenaline in regulating carbohydrate metabolism during moderate exercise, 10 moderately trained men completed two 20 min exercise bouts at 58 ± 2 % peak pulmonary oxygen uptake (̇V_o2,peak). On one occasion saline was infused (CON), and on the other adrenaline was infused intravenously for 5 min prior to and throughout exercise (ADR). Glucose kinetics were measured by a primed, continuous infusion of 6,6-[2H]glucose and muscle samples were obtained prior to and at 1 and 20 min of exercise.

# 2.
The infusion of adrenaline elevated (P < 0.01) plasma adrenaline concentrations at rest (pre-infusion, 0.28 ± 0.09; post-infusion, 1.70 ± 0.45 nmol l−1; means ±s.e.m.) and this effect was maintained throughout exercise. Total carbohydrate oxidation increased by 18 % and this effect was due to greater skeletal muscle glycogenolysis (P < 0.05) and pyruvate dehydrogenase (PDH) activation (P < 0.05, treatment effect). Glucose rate of appearance was not different between trials, but the infusion of adrenaline decreased (P < 0.05, treatment effect) skeletal muscle glucose uptake in ADR.

# 3.
During exercise muscle glucose 6-phosphate (G-6-P) (P = 0.055, treatment effect) and lactate (P < 0.05) were elevated in ADR compared with CON and no changes were observed for pyruvate, creatine, phosphocreatine, ATP and the calculated free concentrations of ADP and AMP.

# 4.
The data demonstrate that elevated plasma adrenaline levels during moderate exercise in untrained men increase skeletal muscle glycogen breakdown and PDH activation, which results in greater carbohydrate oxidation. The greater muscle glycogenolysis appears to be due to increased glycogen phosphorylase transformation whilst the increased PDH activity cannot be readily explained. Finally, the decreased glucose uptake observed during exercise in ADR is likely to be due to the increased intracellular G-6-P and a subsequent decrease in glucose phosphorylation.

Cycling at 120 when compared to 80 rev/min increases the accumulated oxygen deficit but does not affect the precision of its calculation

The aim of the present study was to determine the influence of pedal rate on the precision and quantification of the accumulated oxygen deficit (AOD). Eight trained male triathletes completed a lactate threshold test, VO2 peak test, 10 x 3 min submaximal exercise bouts and a high-intensity exercise bout, all performed at 80 and 120 rev/min. For both pedal rates the intensities for the sub-maximal and high-intensity tests were relative to the lactate threshold and VO2 peak work rates. The VO2-power regressions were calculated using 5 intensities from above the lactate threshold combined with a y intercept value with VO2 measured after 3 min of exercise. For the 120 compared to the 80 rev/min tests, the lactate threshold work rate (255±13 versus 276±47 Watts) (p<0.01) and VO2 peak work rate (352±17 versus 382±20, Watts) (p<0.05) were lower at 120 rev/m. Conversely, the VO2 peak and the VO2 measured during the exhaustive exercise were the same for both pedal rates (p>0.05). Using linear regression modelling the slope of the VO2-power regression (0.0112 versus 0.010 L/Watt) (p<0.01), the estimated total energy demand (ETED) (5.13±0.75 versus 4.89±0.88 L/min) and the AOD (4.27±0.94 versus 3.66±1.25 L) (p<0.05) were greater at 120 rev/m. However, the 95% confidence interval for the ETED and the standard error of the predicted value were the same for both pedal rates (p>0.05). Our results demonstrate that pedal rate effects the size but not the precision of the calculated AOD and should therefore be considered when developing an AOD protocol.

The influence of pacing during 6-minute supra-maximal cycle ergometer performance

The aim of this study was to evaluate the influence of pacing on performance, oxygen uptake (V̇O₂), oxygen deficit and blood lactate accumulation during a 6-minute cycle ergometer test. Six recreational cyclists completed three 6-minute cycling tests using fast-start, even-pacing and slow-fast pacing conditions. Cycle ergometer performance was measured as the mean power output produced for each cycling test. Energy system contribution during each cycling trial was estimated using a modified accumulated oxygen deficit (AOD) method. Blood lactate concentration was analysed from blood sampled using a catheter in a forearm vein prior to exercise, at 2 minutes, 4 minutes and 6 minutes during exercise, and at 2 minutes, 5 minutes and 10 minutes post-exercise. There was no significant difference between the pacing conditions for mean power output (P=0.09), peak V̇O₂ (P=0.92), total V̇O₂ (P=0.76), AOD (P=0.91), the time-course of V̇O₂ (P=0.22) or blood lactate accumulation (P=0.07). There was, however, a significant difference between the three pacing conditions in the oxygen deficit measured over time (P=0.02). These changes in the time-course of oxygen deficit during cycling trials did not, however, significantly affect the mean power output produced by each pacing condition.

The effects of short-term sprint training on MCT expression in moderately endurance-trained runners

The purpose of this study was to assess the effects of short-term sprint training on transient changes in monocarboxylate lactate transporter 1 (MCT1) and MCT4 protein and mRNA content. Seven moderately endurance-trained runners (mean ± SE; age 27.7±2.9 years, body mass 81.1±5.9 kg, VO₂ max 58.1±2.0 ml kg⁻¹ min⁻¹) completed a VO₂ max and a supramaximal running test to exhaustion (RTE) before and after a 6-week period of sprint training. The sprint training was progressive and consisted of 18 sessions of near maximal short duration (5–15 s) sprints to compliment the athlete’s endurance training. Prior to the training period there was a significant (P<0.05) increase in MCT1, but not MCT4 protein, 2 h after the RTE. This occurred without any change in corresponding mRNA levels. After the training period, there was a significant increase in MCT1 protein but no significant change in the MCT4 isoform. Both MCT1 and MCT4 mRNA was significantly lower at rest and 2 h post-RTE after the completion of the training period. After the training period, there was a significant increase in the time to exhaustion and distance covered during the RTE. This study demonstrates that sprint training of this length and type results in an upregulation of MCT1 protein, but not MCT4 content. Additionally, this study shows conflicting adaptations in MCT1 and MCT4 protein and mRNA levels following training, which may indicate post-transcriptional regulation of MCT expression in human muscle.

No effect of mild heat stress on the regulation of carbohydrate metabolism at the onset of exercise

To investigate the influence of heat stress on the regulation of skeletal muscle carbohydrate metabolism, six active, but not specifically trained, men performed 5 min of cycling at a power output eliciting 70% maximal O(2) uptake in either 20 degrees C (Con) or 40 degrees C (Heat) after 20 min of passive exposure to either environmental condition. Although muscle temperature (T(mu)) was similar at rest when comparing trials, 20 min of passive exposure and 5 min of exercise increased (P < 0.05) T(mu) in Heat compared with Con (37.5 +/- 0.1 vs. 36.9 +/- 0.1 degrees C at 5 min for Heat and Con, respectively). Rectal temperature and plasma epinephrine were not different at rest, preexercise, or 5 min of exercise between trials. Although intramuscular glycogen phosphorylase and pyruvate dehydrogenase activity increased (P < 0.05) at the onset of exercise, there were no differences in the activities of these regulatory enzymes when comparing Heat with Con. Accordingly, glycogen use in the first 5 min of exercise was not different when comparing Heat with Con. Similarly, no differences in intramuscular concentrations of glucose 6-phosphate, lactate, pyruvate, acetyl-CoA, creatine, phosphocreatine, or ATP were observed at any time point when comparing Heat with Con. These results demonstrate that, whereas mild heat stress results in a small difference in contracting T(mu), it does not alter the activities of the key regulatory enzymes for carbohydrate metabolism or glycogen use at the onset of exercise, when plasma epinephrine levels are unaltered.

Effect of carbohydrate ingestion on metabolism during running and cycling

The effects of carbohydrate or water ingestion on metabolism were investigated in seven male subjects during two running and two cycling trials lasting 60 min at individual lactate threshold using indirect calorimetry, U-¹⁴C-labeled tracer-derived measures of the rates of oxidation of plasma glucose, and direct determination of mixed muscle glycogen content from the vastus lateralis before and after exercise. Subjects ingested 8 ml/kg body mass of either a 6.4% carbohydrate-electrolyte solution (CHO) or water 10 min before exercise and an additional 2 ml/kg body mass of the same fluid after 20 and 40 min of exercise. Plasma glucose oxidation was greater with CHO than with water during both running (65 ± 20 vs. 42 ± 16 g/h; P < 0.01) and cycling (57 ± 16 vs. 35 ± 12 g/h; P < 0.01). Accordingly, the contribution from plasma glucose oxidation to total carbohydrate oxidation was greater during both running (33 ± 4 vs. 23 ± 3%; P < 0.01) and cycling (36 ± 5 vs. 22 ± 3%; P < 0.01) with CHO ingestion. However, muscle glycogen utilization was not reduced by the ingestion of CHO compared with water during either running (112 ± 32 vs. 141 ± 34 mmol/kg dry mass) or cycling (227 ± 36 vs. 216 ± 39 mmol/kg dry mass). We conclude that, compared with water, 1) the ingestion of carbohydrate during running and cycling enhanced the contribution of plasma glucose oxidation to total carbohydrate oxidation but 2) did not attenuate mixed muscle glycogen utilization during 1 h of continuous submaximal exercise at individual lactate threshold.

Effect of glycerol-induced hyperhydration on thermoregulation and metabolism during exercise in the heat

This study examined the effect of glycerol ingestion on fluid homeostasis, thermoregulation, and metabolism during rest and exercise. Six endurance-trained men ingested either 1 g glycerol in 20 ml H₂O.kg^-1 body weight (bw) (GLY) or 20 ml H₂O.kg^-1bw (CON) in a randomized double-blind fashion, 120 min prior to undertaking 90 min of steady state cycle exercise (SS) at 98 % of lactate threshold in dry heat (35 degrees C, 30 % RH), with ingestion of CHO-electrolyte beverage (6 % CHO) at 15-min intervals. A 15-min cycle, where performance was quantified in kJ, followed (PC). Pre-exercise urine volume was lower in GLY than CON (1119 ± 97 vs. 1503 ± 146 ml· 120 min^-1; p < .05). Heart rate was lower (p < .05) throughout SS in GLY, while forearm blood flow was higher (17.1 ± 1.5 vs. 13.7 ± 3.0 ml.100 g tissue·min^-1; p < .05) and rectal  temperature lower (38.7 ± 0.1 vs. 39.1 ± 0.1 ° C; p < .05) in GLY late in SS. Despite these changes, skin and muscle temperatures and circulating catecholamines were not different between trials. Accordingly, no differences were observed in muscle glycogenolysis, lactate accumulation, adenine nucleotide, and phosphocreatine degradation or inosine 5'-monophosphate accumulation when comparing GLY with CON. Of note, the work performed during PC was 5 % greater in GLY (252 ± 10 vs. 240 ± 9 kJ; p < .05). These results demonstrate that glycerol, when ingested with a bolus of water 2 hours prior to exercise, results in fluid retention, which is capable of reducing cardiovascular strain and enhancing thermoregulation. Furthermore, this practice increases exercise performance in the heat by mechanisms other than alterations in muscle metabolism.

Effect of sodium bicarbonate on muscle metabolism during intense endurance cycling

Introduction: Sodium bicarbonate (NaHCO₃) ingestion has been shown to increase both muscle glycogenolysis and glycolysis during brief submaximal exercise. These changes may be detrimental to performance during more prolonged, exhaustive exercise. This study examined the effect of NaHCO₃ ingestion on muscle metabolism and performance during intense endurance exercise of ~60 min in seven endurance-trained men. Methods: Subjects ingested 0.3 g·kg^-1 body mass of either NaHCO₃ or CaCO₃ (CON) 2 h before performing 30 min of cycling exercise at 77 ± 1% [latin capital V with dot above]O_2peak followed by completion of 469 ± 21 kJ as quickly as possible (~30 min, ~80% [latin capital V with dot above]O_2peak). Results: Immediately before, and throughout exercise, arterialized-venous plasma HCO₃- concentrations were higher (P < 0.05) whereas plasma and muscle H⁺ concentrations were lower (P < 0.05) in NaHCO₃ compared with CON. Blood lactate concentrations were higher (P < 0.05) during exercise in NaHCO₃, but there was no difference between trials in muscle glycogen utilization or muscle lactate content during exercise. Reductions in PCr and ATP and increases in muscle Cr during exercise were also unaffected by NaHCO₃ ingestion. Accordingly, exercise performance time was not different between treatments. Conclusion: NaHCO₃ ingestion resulted in a small muscle alkalosis but had no effect on muscle metabolism or intense endurance exercise performance in well-trained men.

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 O2 uptake (V̇O2,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 μmol (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.

AMP-activated protein kinase activates transcription of the UCP3 and HKII genes in rat skeletal muscle

AMP-activated protein kinase (AMPK) has recently emerged as a key signaling protein in skeletal muscle, coordinating the activation of both glucose and fatty acid metabolism in response to increased cellular energy demand. To determine whether AMPK signaling may also regulate gene transcription in muscle, rats were given a single subcutaneous injection (1 mg/g) of the AMP analog 5-aminoimidazole-4-carboxamide-1-ß-D-ribonucleoside (AICAR). AICAR injection activated (P < 0.05) AMPK-α₂ (~2.5-fold) and transcription of the uncoupling protein-3 (UCP3, ~4-fold) and hexokinase II (HKII, ~10-fold) genes in both red and white skeletal muscle. However, AICAR injection also elicited (P < 0.05) an acute drop (60%) in blood glucose and a sustained (2-h) increase in blood lactate, prompting concern regarding the specificity of AICAR on transcription. To maximize AMPK activation in muscle while minimizing potential systemic counterregulatory responses, a single-leg arterial infusion technique was employed in fully conscious rats. Relative to saline-infused controls, single-leg arterial infusion of AICAR (0.125, 0.5, and 2.5 µg · g^-1 · min^-1 for 60 min) induced a dose-dependent increase (2- to 4-fold, P < 0.05) in UCP3 and HKII transcription in both red and white skeletal muscle. Importantly, AICAR infusion activated transcription only in muscle from the infused leg and had no effect on blood glucose or lactate levels. These data provide evidence that AMPK signaling is linked to the transcriptional regulation of select metabolic genes in skeletal muscle.

Effect of AICAR treatment on gycogen metabolism in skeletal muscle

AMP-activated protein kinase (AMPK) is proposed to stimulate fat and carbohydrate catabolism to maintain cellular energy status. Recent studies demonstrate that pharmacologic activation of AMPK and mutations in the enzyme are associated with elevated muscle glycogen content in vivo. Our purpose was to determine the mechanism for increased muscle glycogen associated with AMPK activity in vivo. AMPK activity and glycogen metabolism were studied in red and white gastrocnemius muscles from rats treated with 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) in vivo, and also in muscles incubated with AICAR in vitro. In vivo AICAR treatment reduced blood glucose and increased blood lactate compared with basal values. AICAR increased muscle α2 AMPK activity, glycogen, and glucose-6-phosphate concentrations. Glycogen synthase activity was increased in the red gastrocnemius but was decreased in the white gastrocnemius. Glycogen phosphorylase activity increased in both muscles, with an inhibition initially observed in the red gastrocnemius. In vitro incubation with AICAR activated α2 AMPK but had no effect on either glycogen synthase or glycogen phosphorylase. These results suggest that AICAR treatment does not promote glycogen accumulation in skeletal muscle in vivo by altering glycogen synthase and glycogen phosphorylase. Rather, the increased glycogen is due to the well-known effects of AICAR to increase glucose uptake.