Extracellular pH defense against lactic acid in untrained and trained altitude residents

The assumption that buffering at altitude is deteriorated by bicarbonate (bi) reduction was investigated. Extracellular pH defense against lactic acidosis was estimated from changes (Δ) in lactic acid ([La]), [HCO₃ ⁻], pH and PCO₂ in plasma, which equilibrates with interstitial fluid. These quantities were measured in earlobe blood during and after incremental bicycle exercise in 10 untrained (UT) and 11 endurance-trained (TR) highlanders (2,600 m). During exercise the capacity of non-bicarbonate buffers (β _nbi = −Δ[La] · ΔpH⁻¹ − Δ[HCO₃ ⁻] · ΔpH⁻¹) amounted to 40 ± 2 (SEM) and 28 ± 2 mmol l⁻¹ in UT and TR, respectively (P < 0.01). During recovery β _nbi decreased to 20 (UT) and 16 (TR) mmol l⁻¹ (P < 0.001) corresponding to values expected from hemoglobin, dissolved protein and phosphate concentrations related to extracellular fluid (ecf). This was accompanied by a larger decrease of base excess after than during exercise for a given Δ[La]. β _bi amounted to 37–41 mmol l⁻¹ being lower than at sea level. The large exercise β _nbi was mainly caused by increasing concentrations of buffers due to temporary shrinking of ecf. Tr has lower β _nbi in spite of an increased Hb mass mainly because of an expanded ecf compared to UT. In highlanders β _nbi is higher than in lowlanders because of larger Hb mass and reduced ecf and counteracts the decrease in [HCO₃ ⁻]. The amount of bicarbonate is probably reduced by reduction of the ecf at altitude but this is compensated by lower maximal [La] and more effective hyperventilation resulting in attenuated exercise acidosis at exhaustion.     

Extracellular pH defense against lactic acid in normoxia and hypoxia before and after a Himalayan expedition

The extracellular pH defense against the lactic acidosis resulting from exercise can be estimated from the ratios −Δ[La] · ΔpH⁻¹ (where Δ[La] is change in lactic acid concentration and ΔpH is change in pH) and Δ[HCO₃ ⁻] · ΔpH⁻¹ (where Δ[HCO₃ ⁻] is change in bicarbonate concentration) in blood plasma. The difference between −Δ[La] · ΔpH⁻¹ and Δ[HCO₃ ⁻] · ΔpH⁻¹ yields the capacity of available non-bicarbonate buffers (mainly hemoglobin). In turn, Δ[HCO₃ ⁻] · ΔpH⁻¹ can be separated into a pure bicarbonate buffering (as calculated at constant carbon dioxide tension) and a hyperventilation effect. These quantities were measured in 12 mountaineers during incremental exercise tests before, and 7–8 days (group 1) or 11–12 days (group 2) after their return from a Himalayan expedition (2800–7600 m altitude) under conditions of normoxia and acute hypoxia. In normoxia −Δ[La] · ΔpH⁻¹ amounted to [mean (SEM)] 92 (6) mmol · l⁻¹ before altitude, of which 19 (4), 48 (1) and 25 (3) mmol · l⁻¹ were due to hyperventilation, bicarbonate and non-bicarbonate buffering, respectively. After altitude −Δ[La] · ΔpH⁻¹ was increased to 128 (12) mmol · l⁻¹ (P < 0.01) in group 1 and decreased to 72 (5) mmol · l⁻¹ in group 2 (P < 0.05), resulting mainly from apparent large changes of non-bicarbonate buffer capacity, which amounted to 49 (14) mmol · l⁻¹ in group 1 and to 10 (2) mmol · l⁻¹ in group 2. In acute hypoxia the apparent increase in non-bicarbonate buffers of group 1 was even larger [140 (18) mmol · l⁻¹]. Since the hemoglobin mass was only modestly elevated after descent, other factors must play a role. It is proposed here that the transport of La⁻ and H⁺ across cell membranes is differently influenced by high-altitude acclimatization. Accepted: 14 September 2000     

Causes of differences in exercise-induced changes of base excess and blood lactate

It has been concluded from comparisons of base excess (BE) and lactic acid (La) concentration changes in blood during exercise-induced acidosis that more H⁺ than La⁻ leave the muscle and enter interstitial fluid and blood. To examine this, we performed incremental cycle tests in 13 untrained males and measured acid–base status and [La] in arterialized blood, plasma, and red cells until 21 min after exhaustion. The decrease of actual BE (−ΔABE) was 2.2 ± 0.5 (SEM) mmol l⁻¹ larger than the increase of [La]_blood at exhaustion, and the difference rose to 4.8 ± 0.5 mmol l⁻¹ during the first minutes of recovery. The decrease of standard BE (SBE), a measure of mean BE of interstitial fluid (if) and blood, however, was smaller than the increase of [La] in the corresponding volume (Δ[La]_if+blood) during exercise and only slightly larger during recovery. The discrepancy between −ΔABE and Δ[La]_blood mainly results from the Donnan effect hindering the rise of [La]_erythrocyte to equal values like [La]_plasma. The changing Donnan effect during acidosis causes that Cl⁻ from the interstitial fluid enter plasma and erythrocytes in exchange for HCO₃⁻. A corresponding amount of La⁻ remains outside the blood. SBE is not influenced by ion shifts among these compartments and therefore is a rather exact measure of acid movements across tissue cell membranes, but changes have been compared previously to Δ[La]_blood instead to Δ[La]_if+blood. When performing correct comparisons and considering Cl⁻/HCO₃⁻ exchange between erythrocytes and extracellular fluid, neither the use of ΔABE nor of ΔSBE provides evidence for differences in H⁺ and La⁻ transport across the tissue cell membranes.     

High-intensity exercise decreases muscle buffer capacity via a decrease in protein buffering in human skeletal muscle

We have previously reported an acute decrease in muscle buffer capacity (βm_{in vitro}) following high-intensity exercise. The aim of this study was to identify which muscle buffers are affected by acute exercise and the effects of exercise type and a training intervention on these changes. Whole muscle and non-protein βm_{in vitro} were measured in male endurance athletes (VO_2max = 59.8 ± 5.8 mL kg⁻¹ min⁻¹), and before and after training in male, team-sport athletes (VO_2max = 55.6 ± 5.5 mL kg⁻¹ min⁻¹). Biopsies were obtained at rest and immediately after either time-to-fatigue at 120% VO_2max (endurance athletes) or repeated sprints (team-sport athletes). High-intensity exercise was associated with a significant decrease in βm_{in vitro} in endurance-trained males (146 ± 9 to 138 ± 7 mmol H⁺·kg d.w.⁻¹·pH⁻¹), and in male team-sport athletes both before (139 ± 9 to 131 ± 7 mmol H⁺·kg d.w.⁻¹·pH⁻¹) and after training (152 ± 11 to 142 ± 9 mmol H⁺·kg d.w.⁻¹·pH⁻¹). There were no acute changes in non-protein buffering capacity. There was a significant increase in βm_{in vitro} following training, but this did not alter the post-exercise decrease in βm_{in vitro}. In conclusion, high-intensity exercise decreased βm_{in vitro} independent of exercise type or an interval-training intervention; this was largely explained by a decrease in protein buffering. These findings have important implications when examining training-induced changes in βm_{in vitro}. Resting and post-exercise muscle samples cannot be used interchangeably to determine βm_{in vitro}, and researchers must ensure that post-training measurements of βm_{in vitro} are not influenced by an acute decrease caused by the final training bout.     

Effects of endurance training on intracellular calcium concentration in T lymphocytes

The purpose of the present study was to determine whether 12 months of endurance training reduced [Ca²⁺]i in T helper (CD4⁺) lymphocytes in trained (TR) men compared to untrained (UT). Fourteen trained (Ironman triathletes) and nine untrained (sedentary) men volunteered for the study. The TR group averaged 12 km of swimming, 300 km of cycling and 60 km of running per week during the year. Resting blood samples were taken from TR (VO_2peak 64 ± 2 ml kg⁻¹ min⁻¹) and UT (VO_2peak 42 ± 2 ml kg⁻¹ min⁻¹) subjects every 4 weeks for 52 weeks (October 1, 1999–October 1, 2000). Leukocyte concentration was measured using a full blood count. Unstimulated CD4⁺ lymphocytes were separated and analysed for changes in free ([Ca²⁺]i) and total ([Ca²⁺]t) calcium using flow cytometry. There were no significant differences in leukocyte concentration between UT and TR groups. There were significant differences between TR and UT in [Ca²⁺]i (October B and November), and [Ca²⁺]t (January and March). There were also significant sequential monthly changes in both [Ca²⁺]i and [Ca²⁺]t for TR and UT groups during the study. Significant increases in [Ca²⁺]i and [Ca²⁺]t during summer (January and March) for both TR and UT groups suggest an increase in intracellular signalling during hot weather. [Ca²⁺]i and [Ca²⁺]t were significantly lower in TR lymphocytes during November and March, suggesting that endurance training during warmer months may decrease [Ca²⁺]i through altered intracellular signalling, possibly to maintain lymphocyte function during heat stress.     

Red cell hemoglobin,hydrogen ion and electrolyte concentrations during exercise in trained and untrained subjects

Summary Red cell concentrations of hemoglobin (MCHC), H⁺, Na⁺, K⁺, Mg⁺⁺, Cl^– were measured in femoral venous blood of six untrained (UT), six endurance trained (TR) and three semitrained (ST) subjects during graded increasing work (4, 8, 12, 18 and 24 mkp/s, 10–15 min on each step) on a bicycle ergometer. Before exercise no significant differences were detected for the measured variables when comparing UT and TR. During exercise MCHC, [Na⁺], [K⁺] and [Mg⁺⁺] remained constant indicating lack of water shift into the erythrocytes in spite of a marked acidosis (lowest pH_Blood value 7.225). This lack resulted from an elevated extracellular osmolality. [H⁺]_Ery and [Cl^–]_Ery maximally increased by 2.0×10^–8 eq/kg H₂O and 10 meq/l, respectively. The change was markedly greater in UT than in TR at equal load. However, if [H⁺]_Ery and [Cl^–]_Ery were related to pH of whole blood, differences between groups almost disappeared and the ions were distributed as predictable from in vitro experiments (Fitzsimmons and Sendroy, 1961). Behaviour of H⁺ and Cl^– may be of importance for oxygen dissociation under in vivo conditions.Supported by Bundesinstitut für Sportwissenschaften, Köln     

Exercise with hypoventilation induces lower muscle oxygenation and higher blood lactate concentration: role of hypoxia and hypercapnia

Eight men performed three series of 5-min exercise on a cycle ergometer at 65% of normoxic maximal O₂ consumption in four conditions: (1) voluntary hypoventilation (VH) in normoxia (VH_0.21), (2) VH in hyperoxia (inducing hypercapnia) (inspired oxygen fraction [F_IO₂] = 0.29; VH_0.29), (3) normal breathing (NB) in hypoxia (F_IO₂ = 0.157; NB_0.157), (4) NB in normoxia (NB_0.21). Using near-infrared spectroscopy, changes in concentration of oxy-(Δ[O₂Hb]) and deoxyhemoglobin (Δ[HHb]) were measured in the vastus lateralis muscle. Δ[O₂Hb − HHb] and Δ[O₂Hb + HHb] were calculated and used as oxygenation index and change in regional blood volume, respectively. Earlobe blood samples were taken throughout the exercise. Both VH_0.21 and NB_0.157 induced a severe and similar hypoxemia (arterial oxygen saturation [SaO₂] < 88%) whereas SaO₂ remained above 94% and was not different between VH_0.29 and NB_0.21. Arterialized O₂ and CO₂ pressures as well as P50 were higher and pH lower in VH_0.21 than in NB_0.157, and in VH_0.29 than in NB_0.21. Δ[O₂Hb] and Δ[O₂Hb − HHb] were lower and Δ[HHb] higher at the end of each series in both VH_0.21 and NB_0.157 than in NB_0.21 and VH_0.29. There was no difference in Δ[O₂Hb + HHb] between testing conditions. [La] in VH_0.21 was greater than both in NB_0.21 and VH_0.29 but not different from NB_0.157. This study demonstrated that exercise with VH induced a lower tissue oxygenation and a higher [La] than exercise with NB. This was caused by a severe arterial O₂ desaturation induced by both hypoxic and hypercapnic effects.     

pH-regulatory mechanisms in in vitro perfused rectal gland tubules of Squalus acanthias

 Isolated in vitro perfused rectal gland tubules (RGT) were preincubated with the pH-sensitive dye 2′,7′-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF) and pH-regulatory mechanisms were studied. A reduction of bath Cl^– concentration from 269 to 6 mmol/l increased the fluorescence ratio 488/436 [corresponding to cytosolic pH (pH_i)] slightly but significantly (n=10). Depolarization by Ba²⁺ (1 mmol/l) or a bath solution containing 30 mmol/l K⁺ (n=4–6) increased the fluorescence ratio (pH_i). These data suggest that HCO₃ ^– uptake and/or H⁺ extrusion is dependent on Cl^– and/or voltage. A reduction of bath Na⁺ from 278 to 5 mmol/l reduced the ratio significantly (n=3). Addition of trimethylamine (Trima⁺, 20 mmol/l) alkalinized cytosolic pH (n=7). Similarly, addition of NH₄ ⁺ (20 mmol/l) led to an initial alkalinization and a strong acidification when NH₄ ⁺ was removed (n=59). The initial pH_i-recovery rates after NH₄ ⁺ removal were quantified and the responsible H⁺ extrusion and/or HCO₃ ^– import systems were examined. The recovery was almost completely abolished when the extracellular Na⁺ concentration was reduced to 5 mmol/l. In the presence of normal Na⁺, recovery was slower in the absence as compared to the presence of HCO₃ ^– (n=5). It was inhibited by 4,4′-diisothiocyanatostilbene-2,2′-disulphonic acid (DIDS) (0.5 mmol/l, n=11) in the presence of HCO₃ ^– and in the absence of HCO₃ ^– by the Na⁺/H⁺-exchange blocker HOE694 (0.5 mmol/l, n=6). These data suggest that acid extrusion probably occurs by basolateral Na⁺-2HCO₃ ^–/Cl^– exchange in the presence of HCO₃ ^– and by basolateral Na⁺/H⁺ exchange in the absence of HCO₃ ^–. Luminal perfusion with a solution containing a low Cl^– concentration (6 mmol/l) increased the fluorescence ratio (pH_i) (n=5). The ratio (pH_i) was further increased and pH recovery further delayed by basolateral addition of Trima⁺ (20 mmol/l, n=3). These data suggest that the HCO₃ ^–/Cl^– exchanger is present in the luminal membrane. Luminal HCO₃ ^–/Cl^– exchange and basolateral Na⁺-2HCO₃ ^–/Cl^– exchange may work in tandem to secrete HCO₃ ^– and exchange it for luminal Cl^–. Received: 7 January 1998 / Received after revision and accepted: 5 March 1998     

Assessment of post-competition peak blood lactate in male and female master swimmers aged 40–79 years and its relationship with swimming performance

The main purpose of this study was to measure the post-competition blood lactate concentration ([La]_b) in master swimmers of both sexes aged between 40 and 79 years in order to relate it to age and swimming performance. One hundred and eight swimmers participating in the World Master Championships were assessed for [La]_b and the average rate of lactate accumulation (La′; mmol l⁻¹ s⁻¹) was calculated. In addition, 77 of them were also tested for anthropometric measures. When the subjects were divided into 10-year age groups, males exhibited higher [La]_b than women (factorial ANOVA, P < 0.01) and a steeper decline with ageing than female subjects. Overall, mean values (SD) of [La]_b were 10.8 (2.8), 10.3 (2.0), 10.3 (1.9), 8.9 (3.2) mmol l⁻¹ in women, and 14.2 (2.5), 12.4 (2.5), 11.0 (1.6), 8.2 (2.0) mmol l⁻¹ in men for, respectively, 40–49, 50–59, 60–69, 70–79 years’ age groups. When, however, [La]_b values were normalised for a “speed index”, which takes into account swimming speed as a percentage of world record, these sex-related differences, although still present, were considerably attenuated. Furthermore, the differences in La′ between males and females were larger in the 40–49 age group (0.34 vs 0.20 mmol l⁻¹ s⁻¹ for 50-m distance) than in the 70–79 age group (0.12 vs 0.14 mmol l⁻¹ s⁻¹ for 50-m distance). Different physiological factors, supported by the considered anthropometric measurements, are suggested to explain the results.     

Effects of bicarbonate ingestion and high intensity exercise on lactate and H(+)-ion distribution in different blood compartments

Lactate (La) and H⁺-ions are unequally distributed in the blood between plasma and red blood cells (RBCs). To our knowledge there is no data concerning the effects of an oral ingestion of bicarbonate (HCO₃ ⁻) on repeated high intensity sprint exercise and La and H⁺ distribution between plasma and RBCs. Since an oral ingestion of HCO₃ ⁻ leads to a higher efflux of La from the working skeletal muscle to the plasma, as it was shown by previous studies, this would lead to a higher gradient of La between plasma and RBCs. Although a higher gradient leads to a higher uptake, it is even more difficult for the RBCs to take up La fast enough, due to the more stressed transport system. Since RBCs function to transport La from the working muscle and help to maintain a concentration difference between plasma and muscle, this potentially increases performance during repeated sprint exercise (e.g. 4 × 30 s). The major goal of the present investigation was to test this hypothesis. 11 male participants ingested either a solution of sodium bicarbonate (NaHCO₃) or placebo (CaCO₃). Thereafter all performed four maximal 30 s sprints with 5 min of passive rest. During the resting periods concentrations of HCO₃ ⁻, La and H⁺ where measured in both blood compartments (plasma and RBCs). There were no significant differences in the La-ratios between plasma and RBCs between both interventions. These results indicate that the La/H⁺ co-transport is not affected by an oral ingestion on NaHCO₃.     

Differential pathways for calcium influx activated by concanavalin A and CD3 stimulation in Jurkat T cells

Sustained increase in [Ca²⁺]_c (Δ[Ca²⁺]_c) is a critical early signal from T-cell receptor (TCR/CD3). In general, Ca²⁺-release activated Ca²⁺ channels (CRAC) are responsible for the Ca²⁺ influx and Δ[Ca²⁺]_c after TCR/CD3 stimulation. However, T cells also express Ca²⁺-permeable nonselective cation channels such as TRPM2 and TRPC. Gd³⁺ is a relatively selective blocker for CRAC at micromolar concentrations. Here, Jurkat T cells were used to investigate the Gd³⁺-resistant Ca²⁺ influx (Δ[Ca²⁺]_c,Gd) induced by concanavalin A (ConA, 1 μg/ml), a widely used mitogenic agent for T cells, or by anti-CD3 Ab (αCD3). αCD3-induced Δ[Ca²⁺]_c was partly (~60%) inhibited by 1 μM Gd³⁺ while thapsigargin-induced Δ[Ca²⁺] was almost completely abolished. ConA-induced Δ[Ca²⁺] was mostly inhibited by 1 μM Gd³⁺ during the early phase (<30 s of ConA application) and became resistant during the late phase (>2 min). Induction of Δ[Ca²⁺]_c,Gd by αCD3 and ConA was inhibited by 2-aminoethoxydiphenyl borate (2-APB) and by N-(p-amylcinnamoyl) anthranilic acid, indicating that TRPM2 and TRPC are involved in this process. Treatment with Pyr-3, a TRPC3-specific inhibitor, potently suppressed Δ[Ca²⁺]_c,Gd by αCD3 (IC₅₀, 0.16 μM). Patch clamp experiments demonstrated that the TRPM2 channels were activated by ConA, and the TRPC-like channels were activated by αCD3. Our present study suggests that TRPM2 and TRPC3 are activated by ConA and TCR/CD3, respectively, in Jurkat T cells and are responsible for the induction of Δ[Ca²⁺]_c,Gd.     

The effect of intracellular pH on cytosolic Ca2+ in HT29 cells

 The influence of intracellular pH (pH_i) on intracellular Ca²⁺ activity ([Ca²⁺]_i) in HT₂₉ cells was examined microspectrofluorometrically. pH_i was changed by replacing phosphate buffer by the diffusible buffers CO₂/HCO₃ ^–or NH₃/NH₄ ⁺ (pH 7.4). CO₂/HCO₃ ^–buffers at 2,5 or 10% acidified pH_i by 0.1, 0.32 and 0.38 pH units, respectively, and increased [Ca²⁺]_i by 8–15 nmol/l. This effect was independent of the extracellular Ca²⁺ activity and the filling state of thapsigargin-sensitive Ca²⁺ stores. Removing the CO₂/HCO₃ ^–buffer alkalinized pH_i by 0.14 (2%), 0.27 (5%), and 0.38 (10%) units and enhanced [Ca²⁺]_i to a peak value of 20, 65, and 143 nmol/l, respectively. Experiments carried out with Ca²⁺-free solution and with thapsigargin showed that the [Ca²⁺]_i transient was due to release from intracellular pools and stimulated Ca²⁺ entry. NH₃/NH₄ ⁺ (20 mmol/l) induced a transient intracellular alkalinization by 0.6 pHunits and increased [Ca²⁺]_i to a peak (Δ [Ca²⁺]_i = 164 nmol/l). The peak [Ca²⁺]_i increase was not influenced by removal of external Ca²⁺, but the decline to basal [Ca²⁺]_i was faster. Neither the phospholipase C inhibitor U73122 nor the inositol 1,4,5-trisphosphate (InsP ₃) antagonist theophylline had any influence on the NH₃/NH₄ ⁺-stimulated [Ca²⁺]_i increase, whereas carbachol-induced [Ca²⁺]_i transients were reduced by more than 80% and 30%, respectively. InsP ₃ measurements showed no change of InsP ₃ during exposure to NH₃/NH₄ ⁺, whereas carbachol enhanced the InsP ₃ concentration, and this effect was abolished by U73122. The pH_i influence on ”capacitative” Ca²⁺ influx was also examined. An acid pH_i attenuated, and an alkaline pH_i enhanced, carbachol- and thapsigargin-induced [Ca²⁺]_i influx. We conclude that: (1) an alkaline pH_i releases Ca²⁺ from InsP ₃-dependent intracellular stores; (2) the store release is InsP ₃ independent and occurs via an as yet unknown mechanism; (3) the store release stimulates capacitative Ca²⁺ influx; (4) the capacitative Ca²⁺ influx activated by InsP ₃ agonists is decreased by acidic and enhanced by alkaline pH_i. The effects of pH_i on [Ca²⁺]_i should be of relevance under many physiological conditions. Received: 17 June 1996 / Received after revision and accepted: 30 August 1996     

Effect of acute sodium bicarbonate ingestion on excess C02 output during incremental exercise

Summary The effect of bicarbonate ingestion on total excess volume of CO₂ Output (CO₂ excess), due to bicaronate buffering of lactic acid in exercise, was studied in eight healthy male volunteers during incremental exercise on a cycle ergometer performed after ingestion (0.3 g · kg^–1 body mass) of CaCO₃ (control) and NaHCO₃ (alkalosis). The resting arterialized venous blood pH (P<0.05) and bicarbonate concentration ([HCO₃ ^–]_b;P<0.01) were significantly higher in acute metabolic alkalosis [AMA; pH, 7.44 (SD 0.03); [HCO₃ ^–]_b; 29.4 (SD 1.5) mmol·1^-1] than in the control [pH, 7.39 (SD 0.03); [HCO₃ ^–]_b, 25.5 (SD 1.0) mmol·1^–1]. The blood lactate concentrations ([la^–]_b) during exercise below the anaerobic threshold (AT) were not affected by AMA, while significantly higher [la^–]_b at exhaustion [12.29 (SD 1.87) vs 9.57 (SD 2.14) mmol·1^–1,P < 0.05] and at 3 min after exercise [14.41 (SD 1.75) vs 12.26 (SD 1.40) mmol · l^–1,P < 0.05] were found in AMA compared with the control. The CO₂ excess increased significantly from the control [3177 (SD 506) ml] to AMA [3897 (SD 381) ml;P < 0.05]. The CO₂ excess per body mass was found to be significantly correlated with both the increase of [la^–]_b from rest to 3 min after exercise ( [la^–]_b;r=0.926,P < 0.001) and with the decrease of [HCO₃ ^–]_b from rest to 3 min after exercise ( [HCO₃ ^–]_b;_r=0.872,P<0.001), indicating that CO₂ excess per body mass increased linearly with both [la^– _b and [HCO₃ ^–]_b. As a consequence, CO₂ excess per body mass per unit increase of [la^–]_b (CO₂ excess·mass^–1· [la^–]_b) was similar for the two conditions. The present results would suggest that the relationship between CO₂ excess and blood lactate accumulation was unaffected by acute metabolic alkalosis, because the relative contribution of bicarbonate buffering of lactic acid was the same as in the control.     

Peripheral markers of central fatigue in trained and untrained during uncompensable heat stress

The development of fatigue is more pronounced in the heat than thermoneutral environments; however, it is unclear whether biomarkers of central fatigue are consistent with the higher core temperature (T _c) tolerated by endurance trained (TR) versus untrained (UT) during exertional heat stress (EHS). The purpose of this study was to examine the indicators of central fatigue during EHS in TR versus UT. Twelve TR and 11 UT males (mean ± SE [(V)\dot]\textO _{2 \textpeak} \dot{V}{\text{O}}_{{ 2 {\text{peak}}}}  = 70 ± 2 and 50 ± 1 mL kg LBM⁻¹ min⁻¹, respectively) walked on a treadmill to exhaustion (EXH) in 40°C (dry) wearing protective clothing. Venous blood was obtained at PRE and 0.5°C T _c increments from 38 to 40°C/EXH. Free tryptophan (f-TRP) decreased dramatically at 39.5°C for the TR. Branch chain amino acids decreased with T _c and were greater for UT than TR at EXH. Tyrosine and phenylalanine remained unchanged. Serum S100β was undetectable (<5 pg mL⁻¹). Albumin was greater for the UT from PRE to 39.0°C and at EXH. Prolactin (PRL) responded to relative thermal strain with similar EXH values despite higher T _c tolerated for TR (39.7 ± 0.09°C) than UT (39.0 ± 0.09°C). The high EXH PRL values for both groups support its use as a biomarker of the serotonin and dopamine interplay within the brain during the development of central fatigue.     

Effect of alkalinization of cytosolic pH by amines on intracellular Ca2+ activity in HT29 cells

The effect of secondary, tertiary and quaternary methyl- and ethylamines on intracellular pH (pH_i) and intracellular Ca²⁺ activity ([Ca²⁺]_i) of HT₂₉ cells was investigated microspectrofluorimetrically using pH- and Ca²⁺- sensitive fluorescent indicators, [i.e. 2′,7′-biscarboxyethyl-5(6)-carboxyfluorescein (BCECF) and fura-2 respectively]. Membrane voltage (V _m) was studied by the patch-clamp technique. Secondary and tertiary amines led to a rapid and stable concentration-dependent alkalinization which was independent of their pK _a value. Trimethylamine (20 mmol/l) increased pH_i by 0.78 ± 0.03 pH units (n = 9) and pH remained stable for the application time. Removal led to an undershoot of pH_i and a slow and incomplete recovery: pH_i stayed 0.26 ± 0.06 pH units more acid than the resting value. The quaternary amines, tetramethyl- and tetraethylamine were without influence on pH_i. All tested secondary and tertiary amines (dimethyl-, diethyl-, trimethyl-, and triethyl-amine) induced a [Ca²⁺]_i transient which reached a peak value within 10–25 s and then slowly declined to a [Ca²⁺]_i plateau. The initial Δ[Ca²⁺]_i induced by trimethylamine (20 mmol/l) was 160 ± 15 nmol/l (n = 17). The [Ca²⁺]_i peak was independent of the Ca²⁺ activity in the bath solution, but the [Ca²⁺]_i plateau was significantly lower under Ca²⁺-free conditions and could be immediately interrupted by application of CO₂ (10%; n = 6), a manoeuvre to acidify pH_i in HT₂₉ cells. Emptying of the carbachol- or neurotensin-sensitive intracellular Ca²⁺ stores completely abolished this [Ca²⁺]_i transient. Tetramethylamine led to higher [Ca²⁺]_i changes than the other amines tested and only this transient could be completely blocked by atropine (10⁻⁶ mol/l). Trimethylamine (20 mmol/l) hyperpolarized V _m by 22.5 ± 3.7 mV (n = 16) and increased the whole-cell conductance by 2.3 ± 0.5 nS (n = 16). We conclude that secondary and tertiary amines induce stable alkaline pH_i changes, release Ca²⁺ from intracellular, inositol-1,4,5-trisphosphate-sensitive Ca²⁺ stores and increase Ca²⁺ influx into HT₂₉ cells. The latter may be related to both the store depletion and the hyperpolarization. Received: 11 September 1995/Received after revision and accepted: 18 December 1995     

Cultured ruminal epithelial cells express a large-conductance channel permeable to chloride, bicarbonate, and acetate

The absorption of short-chain fatty acids (SCFA) from the rumen requires efficient mechanisms for both apical uptake and basolateral extrusion. Previous studies suggest that the rumen expresses a basolateral chloride conductance that might be permeable to SCFA. In order to characterize this conductance in more detail, isolated cultured ruminal epithelial cells were studied with the patch-clamp technique, revealing a whole-cell conductance with p(Cl⁻) ≈ p(NO₃ ⁻) > p(HCO₃ ⁻) > p(acetate⁻) > p(gluconate⁻). Currents could be blocked by diisothiocyanato-stilbene-2,2′-disulfonic acid (1 mmol l⁻¹ > 100 μmol l⁻¹), 5-nitro-2-(3-phenylpropyl-amino)benzoic acid (50 μmol l⁻¹), niflumic acid (100 μmol l⁻¹), and p-chloromercuribenzoate (1 mmol l⁻¹). Single-channel conductance was 350 ± 7 pS for chloride and 142 ± 7 pS for acetate. Open probability could be fitted with a three-state gating model. We propose a role for this channel in mediating the permeation of chloride, bicarbonate, and acetate across the basolateral membrane of the ruminal epithelium.     

In vivo 4-androstene-3,17-dione and 4-androstene-3β,17β-diol supplementation in young men

To determine if known androgenic hormone precursors for testosterone in the androgen pathway would be readily transformed to testosterone, eight male subjects [mean age 23.8 (SEM 3) years, bodymass 83.1 (SEM 8.7) kg, height 175.6 (SEM 8.5) cm] underwent a randomized, double-blind, cross-over, placebo-controlled oral treatment with 200 mg of 4-androstene-3,17-dione (Δ4), 4-androstene-3β,17β-diol (Δ4Diol), and placebo (PL). The periods of study were separated by 7 days of washout. Blood was drawn at baseline and subsequently every 30 min for 90 min after treatment. Analysis revealed mean area-under-the-curve (AUC) serum Δ4 concentrations to be higher during Δ4 treatment [2177 (SEM 100) nmol · l⁻¹] than Δ4Diol [900 (SEM 96) nmol · l⁻¹] or PL [484 (SEM 82) nmol · l⁻¹; P < 0.0001]. The Δ4 treatment also revealed a significant effect on total testosterone with a mean AUC [1632.5 (SEM 121) nmol · l⁻¹] that was greater than PL [1418.5 (SEM 131) nmol · l⁻¹; P < 0.05] but not significantly different from those observed after Δ4Diol treatment [1602.9 (SEM 119) nmol · l⁻¹; P = 0.77]. Free testosterone concentrations followed a similar pattern where mean AUC for the Δ4 treatment [6114.0 (SEM 600) pmol · l⁻¹] was greater than after PL [4974.6 (SEM 565) pmol · l⁻¹; P < 0.06] but not significantly different from those observed after Δ4Diol [5632.0 (SEM 389) pmol · l⁻¹; P = 0.48]. The appearance and apparent conversion to total and free testosterone over 90 min was stronger for the Δ4 treatment (r = 0.91, P < 0.045) than for Δ4Diol treatment (r = 0.69, NS) and negatively correlated for PL (r = −0.90, P < 0.02). These results would suggest that Δ4, and perhaps Δ4Diol, taken by month are capable of producing in vivo increases in testosterone concentrations in apparently healthy young men as has already been observed in women after treatment with Δ4. Accepted: 26 August 1999     

Inspiratory resistive loading after all-out exercise improves subsequent performance

We have previously shown that post-exercise inspiratory resistive loading (IRL) reduces blood lactate ([Lac_b⁻]). In this study, we tested the hypothesis that IRL during recovery could improve subsequent exercise performance. Eight healthy men underwent, on different days, two sequential 30-s, cycle ergometer Wingate tests. During the 10-min recovery period from test 1, subjects breathed freely or through an inspiratory resistance (15 cm H₂O) with passive leg recovery. Arterialized [Lac_b⁻] values, perceptual scores (Borg), cardiac output by impedance cardiography (QT), and changes in the deoxygenation status of the M. vastus lateralis by near-infrared spectroscopy (ΔHHb), were recorded. [Lac_b⁻] was significantly reduced after 4 min of recovery with IRL (peak [Lac_b⁻] 12.5 ± 2.3 mmol l⁻¹ with free-breathing vs. 9.8 ± 1.5 mmol l⁻¹ with IRL). Effort perception was reduced during late recovery with IRL compared with free-breathing. Cardiac work was increased with IRL, since heart rate and QT were elevated during late recovery. Peripheral muscle reoxygenation, however, was significantly impaired with IRL, suggesting that post-exercise convective O₂ delivery to the lower limbs was reduced. Importantly, IRL had a dual effect on subsequent performance, i.e., improvement in peak and mean power, but increased fatigue index (P < 0.05). Our data demonstrate that IRL after a Wingate test reduces post-exercise effort perception and improves peak power on subsequent all-out maximal-intensity exercise.     

Exercise-induced changes in blood levels of alpha-tocopherol

Levels of α-tocopherol (αT) in plasma and red blood cells (RBC) are assumed to be modulated by exercise. The mechanisms involved remain to be established. We examined the influence of different running bouts on the content of αT in RBC (αT_RBC), the concentration in plasma (αT_plasma), and their relationship with lipolysis, as indicated by changes (Δ) in plasma glycerol concentration ([glycerol]). Eleven healthy runners [mean (SD) age 35 (9) years, height 177.3 (7.6) cm, body mass 69.6 (9.4) kg, and peak oxygen consumption, , 57.8 (4.8) ml^.kg^–1.min^–1] performed an incremental treadmill test [duration 17 (2) min, peak velocity, v _peak 4.8 (0.4) m^.s^–1], a training run [173 (12) min, 57 (4)% v _peak] and a marathon [197 (24) min, 75 (5)% v _peak]. Before (pre) and after (post) each run, haematological and lipid parameters, αT_RBC and αT_Plasma were determined. Haemoconcentration was observed after each run. Δ[glycerol] was +0.10 (0.10) mmol^.l^–1, +0.40 (0.14) mmol^.l^–1 and +0.51 (0.15) mmol^.l^–1 in the treadmill test, training run and marathon, respectively. When corrected for haemoconcentration, values of αT_plasma decreased [–5.4 (7.5)%, P<0.05] in the treadmill test, were unchanged [+0.7 (8.7)%] in the training run and increased [+7.8 (8.3)%, P<0.05] in the marathon. αT_RBC decreased [pre vs post: 22.7 (3.2) nmol^.g haemoglobin^–1 (nmol^.g Hb^–1) vs 18.9 (3.8) nmol^.g Hb^–1, P<0.05] in the treadmill test and was not significantly changed in either the training run [20.8 (1.9) nmol^.g Hb^–1 vs 19.1 (3.0) nmol^.g Hb^–1] or the marathon [21.6 (2.9) nmol^.g Hb^–1 vs 23.4 (2.7) nmol^.g Hb^–1]. ΔαT_RBC and ΔαT_plasma were positively related to Δ[glycerol]. The reduction in αT_RBC and αT_plasma after short-lasting heavy exercise indicates the consumption of αT, whereas the association between ΔαT and Δ[glycerol] suggests mobilisation of αT, especially in long-lasting exercises. However, although αT appears to be influenced by exercise, the results suggest a well-balanced regulation of αT during exercise resulting in small, and only in part, significant ΔαT in blood. Electronic Publication     

Role of physiological HCO3 –buffer on intracellular pH and histamine release in rat peritoneal mast cells

 The purpose of this study was to examine how intracellular pH (pH_i) regulation and histamine release are affected by HCO₃ ^–in rat peritoneal mast cells. The pH_i was measured using the pH-sensitive dye 2′, 7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). We observed a pH_i of 6.88±0.012 (n=24) in resting mast cells exposed to a HEPES buffer (pH 7.4), but a sustained drop of 0.21 pH units to 6.67±0.015 (n=23) when we exposed the mast cells to a HEPES/HCO₃ ^–buffer equilibrated at all time with 5% CO₂ (pH 7.4). This fall in pH_i is inhibited by the carbonic anhydrase inhibitor dichlorphenamide and is Na⁺-independent, indicating the involvement of Na⁺-independent Cl^–/HCO₃ ^–exchange activity. Furthermore removal of external Cl^–in the presence but not in the absence of HCO₃ ^–reversed the Cl^–/HCO₃ ^–exchange and induced an alkaline load. The recovery from this alkaline load was dependent on external Cl^–but independent of Na⁺. Both the alkalinization and the recovery were inhibited by the anion transport inhibitor 4,4′-diisothiocyanatostilbene-2,2′-disulphonic acid (DIDS). In addition, ³⁶Cl^–uptake measurements confirm the presence of a Cl^–/HCO₃ ^–exchanger. Histamine release stimulated by antigen and compound 48/80 was substantially reduced in the presence of HEPES/ HCO₃ ^–buffer (pH_o 7.4, pH_i 6.66). Histamine release was increased, however, when pH_i was clamped to 6.66 in HCO₃ ^–-free media (pH_o 6.9). We conclude that: (1) Na⁺-independent Cl^–/HCO₃ ^–exchange determines steady-state pH_i in rat peritoneal mast cells; and (2) the reduction in histamine release observed in the presence of HCO₃ ^–is not due to its effect on pH_i per se, but rather on other changes in ion transport. Received: 29 January 1998 / Received after revision and accepted: 3 April 1998