Oxidative metabolism and anaerobic glycolysis during repeated exercise

The purpose of the present study was to examine aerobic and muscle anaerobic energy production during supramaximal repeated exercise. Eight subjects performed three 2-min bouts of cycling (EX1-EX3) at an intensity corresponding to about 125 % of VO2 max separated by 15 min of rest. Ventilatory variables were measured breath by breath during the exercise and a muscle biopsy was taken before and after each exercise bout. Blood samples were collected before and after each cycling period and during the recovery periods. Total work in the first 2 min bout of cycling, EX1, [46.3 +/- 2.1 KJ] was greater than in the second, EX2, (p < 0.01) and in the third, EX3, (p < 0.05). The ATP utilization [4.0 +/- 1.4 mmol x (kg dry weight)(-1), EX1] during the three exercise bouts was the same. The decrement in muscle phosphocreatine (PCr) [46.8 +/- 8.5 mmol x (kg dry weight)(-1), EX1] was also similar for the three exercise bouts. Muscle lactate accumulation was greater (p < 0.05) during EX1 compared to EX2 and EX3. The total oxygen consumption was the same for the three exercise bouts, but when it is corrected for the total work performed, oxygen uptake during EX2 (153 +/- 9 ml x KJ(-1)) and EX3 (150 +/- 9 ml x KJ(-1)) was higher (p < 0.01 and p < 0.05, respectively) than during EX1 (139 +/- 8 ml x KJ(-1)). The present data suggest that oxidative metabolism does not compensate for the reduction of anaerobic glycolysis during repeated fatiguing exercise.     

Impaired muscle Ca2+ and K+ regulation contribute to poor exercise performance post-lung transplantation.

Lung transplant recipients (LTx) exhibit marked peripheral limitations to exercise. We investigated whether skeletal muscle Ca2+ and K+ regulation might be abnormal in eight LTx and eight healthy controls. Peak oxygen consumption and arterialized venous plasma [K+] (where brackets denote concentration) were measured during incremental exercise. Vastus lateralis muscle was biopsied at rest and analyzed for sarcoplasmic reticulum Ca2+ release, Ca2+ uptake, and Ca2+-ATPase activity rates; fiber composition; Na+-K+-ATPase (K+-stimulated 3-O-methylfluorescein phosphatase) activity and content ([3H]ouabain binding sites); as well as for [H+] and H+-buffering capacity. Peak oxygen consumption was 47% less in LTx (P < 0.05). LTx had lower Ca2+ release (34%), Ca2+ uptake (31%), and Ca2+-ATPase activity (25%) than controls (P < 0.05), despite their higher type II fiber proportion (LTx, 75.0 +/- 5.8%; controls, 43.5 +/- 2.1%). Muscle [H+] was elevated in LTx (P < 0.01), but buffering capacity was similar to controls. Muscle 3-O-methylfluorescein phosphatase activity was 31% higher in LTx (P < 0.05), but [3H]ouabain binding content did not differ significantly. However, during exercise, the rise in plasma [K+]-to-work ratio was 2.6-fold greater in LTx (P < 0.05), indicating impaired K+ regulation. Thus grossly subnormal muscle calcium regulation, with impaired potassium regulation, may contribute to poor muscular performance in LTx.     

Short-term training attenuates muscle TCA cycle expansion during exercise in women.

Muscle glycogenolytic flux and lactate accumulation during exercise are lower after 3-7 days of "short-term" aerobic training (STT) in men (e.g., Green HJ, Helyar R, Ball-Burnett M, Kowalchuk N, Symon S, and Farrance B. J Appl Physiol 72: 484-491, 1992). We hypothesized that 5 days of STT would attenuate pyruvate production and the increase in muscle tricarboxylic acid cycle intermediates (TCAI) during exercise, because of reduced flux through the reaction catalyzed by alanine aminotransferase (AAT; pyruvate + glutamate <--> 2-oxoglutarate + alanine). Eight women [22 +/- 1 yr, peak oxygen uptake (Vo2 peak) = 40.3 +/- 4.6 ml. kg-1. min-1] performed seven 45-min bouts of cycle exercise at 70% Vo2 peak over 9 days (1 bout/day; rest only on days 2 and 8). During the first and last bouts, biopsies (vastus lateralis) were obtained at rest and after 5 and 45 min of exercise. Muscle glycogen concentration was approximately 50% higher at rest after STT (493 +/- 38 vs. 330 +/- 20 mmol/kg dry wt; P 相似文献   

Inositol 1,4,5-trisphosphate-induced Ca2+ release from the sarcoplasmic reticulum and contraction in crustacean muscle

Intracellular applications of a fixed amount (0.2 to 8 nmol) of inositol 1,4,5-trisphosphate (InsP3) over a brief period (2 s) into barnacle muscle fibers induced vigorous contractures. Peak tension attained during the first application depended on [InsP3]: the maximum tension evoked by the injection of 8 nmol was 1.6 kg/cm2. Peak tension during a second application of a high dose of InsP3 (greater than 10 microM) was always smaller than that during the first application. Extracellular Ca2+ could be omitted with no measurable effects on either the amplitude or time course of the contractures evoked by InsP3. Aequorin was used to measure InsP3-evoked Ca2+ release from intracellular stores in minced muscle fibers from lobster and in skinned muscle fibers from barnacle. Provided the sarcoplasmic reticulum was preloaded with Ca2+, application of InsP3 induced a transient Ca2+ release that was [InsP3] dependent. During each transient, [Ca2+] rose rapidly to a peak value (t1/2 less than 5 s) and then slowly returned (t1/2 less than 100 s) to a basal level. Maximum Ca2+ release was obtained at [InsP3] less than 100 microM and amounted to 4 nmol Ca2+/g of muscle, enough to increase [Ca2+]i from 0.1 to 8 microM had the Ca2+ release occurred in the intact fiber. Successive applications of a fixed amount of InsP3 elicited successive transient increases in Ca2+. The effects of [Ca2+] on the incorporation of [3H]inositol into the pools of phosphatidylinositol, phosphatidylinositol 4-phosphate, and phosphatidylinositol 4,5-bisphosphate pools were measured.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of high-intensity intermittent training on lactate and H+ release from human skeletal muscle

The study investigated the effect of training on lactate and H+ release from human skeletal muscle during one-legged knee-extensor exercise. Six subjects were tested after 7-8 wk of training (fifteen 1-min bouts at approximately 150% of thigh maximal O2 uptake per day). Blood samples, blood flow, and muscle biopsies were obtained during and after a 30-W exercise bout and an incremental test to exhaustion of both trained (T) and untrained (UT) legs. Blood flow was 16% higher in the T than in the UT leg. In the 30-W test, venous lactate and lactate release were lower in the T compared with the UT leg. In the incremental test, time to fatigue was 10.6 +/- 0.7 and 8.2 +/- 0.7 min, respectively, in the T and UT legs (P < 0.05). At exhaustion, venous blood lactate was 10.7 +/- 0.4 and 8.0 +/- 0.9 mmol/l in T and UT legs (P < 0.05), respectively, and lactate release was 19.4 +/- 3.6 and 10.6 +/- 2.0 mmol/min (P < 0.05). H+ release at exhaustion was higher in the T than in the UT leg. Muscle lactate content was 59.0 +/- 15.1 and 96.5 +/- 14.5 mmol/kg dry wt in the T and UT legs, and muscle pH was 6.82 +/- 0.05 and 6.69 +/- 0.04 in the T and UT legs (P = 0.06). The membrane contents of the monocarboxylate transporters MCT1 and MCT4 and the Na+/H+ exchanger were 115 +/- 5 (P < 0.05), 111 +/- 11, and 116 +/- 6% (P < 0.05), respectively, in the T compared with the UT leg. The reason for the training-induced increase in peak lactate and H+ release during exercise is a combination of an increased density of the lactate and H+ transporting systems, an improved blood flow and blood flow distribution, and an increased systemic lactate and H+ clearance.     

MR measurements of muscle damage and adaptation after eccentric exercise.

The purposes of this study were, first, to clarify the long-term pattern of T2 relaxation times and muscle volume changes in human skeletal muscle after intense eccentric exercise and, second, to determine whether the T2 response exhibits an adaptation to repeated bouts. Six young adult men performed two bouts of eccentric biceps curls (5 sets of 10 at 110% of the 1-repetition concentric maximum) separated by 8 wk. Blood samples, soreness ratings, and T2-weighted axial fast spin-echo magnetic resonance images of the upper arm were obtained immediately before and after each bout; at 1, 2, 4, 7, 14, 21, and 56 days after bout 1; and at 2, 4, 7 and 14 days after bout 2. Resting muscle T2 [27.6 +/- 0.2 (SE) ms] increased immediately postexercise by 8 +/- 1 ms after both bouts. T2 peaked 7 days after bout 1 at 47 +/- 4 ms and remained elevated by 2.5 ms at 56 days. T2 peaked lower (37 +/- 4 ms) and earlier (2-4 days) after bout 2, suggesting an adaptation of the T2 response. Peak serum creatine kinase values, pain ratings, and flexor muscle swelling were also significantly lower after the second bout (P < 0.05). Total volume of the imaged arm region increased transiently after bout 1 but returned to preexercise values within 2 wk. The exercised flexor compartment swelled by over 40%, but after 2 wk it reverted to a volume 10% smaller than that before exercise and maintained this volume loss through 8 wk, consistent with partial or total destruction of a small subpopulation of muscle fibers.     

Contribution of erythrocytes to the control of the electrolyte changes of exercise

Five healthy males performed four 30-s bouts of maximal isokinetic cycling with 4 min rest between each bout. Arterial and femoral venous blood was sampled during and for 90 min following exercise. During exercise, arterial erythrocyte [K+] increased from 117.0 +/- 6.6 mequiv./L at rest to 124.2 +/- 5.9 mequiv./L after the second exercise bout. Arterial erythrocyte [K+] returned to the resting values during the first 5 min of recovery. No significant change was observed in femoral venous erythrocyte [K+]. Arterial erythrocyte lactate concentration ([Lac-]) increased during exercise from 0.2 +/- 0.1 mequiv./L peaking at 9.5 +/- 1.5 mequiv./L at 5 min of recovery, after which the values returned to control. Femoral venous erythrocyte [Lac-] changed in a similar fashion. Arterial erythrocyte [Cl-] rose during exercise to 76 +/- 3 mequiv./L and returned to resting values (70 +/- 2 mequiv./L) by 25 min recovery. During exercise there was a net flux of Cl- into the erythrocyte. We conclude that erythrocytes are a sink for K+ ions leaving working muscles. Furthermore, erythrocytes function to transport Lac- from working muscle and reduce plasma acidosis by uptake of Cl-. The erythrocyte uptake of K+, Lac-, and Cl- helps to maintain a concentration difference between plasma and muscle, facilitating diffusion of Lac- and K+ from the interstitial space into femoral venous plasma.     

Roles of CO2, O2, and acid in arteriovenous [H+] difference during muscle contractions

To determine the origins of the arteriovenous [H+] difference of muscle during contractions, arterial and muscle venous blood sample pairs were taken before and after 0.5, 5.0, and 30.0 min of 4/s isometric twitches of the gastrocnemius-plantaris muscle group of anesthetized dogs. These samples were analyzed for PO2, PCO2, and pH, the concentrations of O2, CO2, K+, Na+, La-, and Cl- in whole blood, and La-, K+, Na+, and Cl- in plasma. Whole blood was hemolyzed and analyzed for PO2, PCO2, and pH. Net O2 uptake, CO2 output, L, K+, Na+, and Cl- were calculated in addition to net output of non-CO2 acid (HA) and strong ion difference ([SID]) and common ion [SID] ([K+] + [Na+] - [Cl-] - [La-]). From these data we partitioned the origins of the arteriovenous [H+] difference via the common PCO2-pH diagram and via a [H+]-PCO2 diagram and determined whether true plasma arteriovenous [H+] differences reflect plasma and cell arteriovenous [H+] differences. The arteriovenous [H+] differences of plasma and hemolyzed blood were the same, showing that true plasma does reflect plasma and cells. K+ showed a small significant but transient output. Na+ was not significant, whereas Cl- showed a significant transient uptake. Lactate output and HA, calculated for dog blood acid-base, showed transient outputs and were the same. At 5.0 min when the arteriovenous difference was largest, CO2 alone would have increased [H+] 15.9 nmol/l whereas desaturation of Hb would have decreased [H+] 4.2 nmol/l and lactate could have raised [H+] 1.0 nmol/l.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inositol 1,4,5-trisphosphate increases myoplasmic [Ca2+] in isolated muscle fibers. Depolarization enhances its effects

Inositol 1,4,5-trisphosphate (InsP3) has been proposed as an intracellular messenger which mobilizes calcium from the sarcoplasmic reticulum, during excitation-contraction coupling in skeletal muscle. We have measured the myoplasmic free calcium concentration ([Ca2+]i) by means of calcium selective microelectrodes in intact fibers isolated from Leptodactylus insularis microinjected with InsP3. In muscle fibers bathed in normal Ringer, the mean resting [Ca2+]i was 0.11 +/- 0.01 microM (M +/- SEM, n = 30). The microinjection of 0.3, 0.5 and 1 microM InsP3 induced transient increments in the [Ca2+]i to 0.35 +/- 0.02 microM (n = 9), to 0.53 +/- 0.03 microM (n = 11) and 0.94 +/- 0.06 microM (n = 10) respectively. Microinjection of 0.3, 0.5 and 1 microM InsP3 in muscle fibers incubated in low Ca2+ solution induced increments in [Ca2+]i similar to those observed in fibers bathed with normal Ringer. The microinjection of 0.3, 0.5 and 1 microM InsP3 in muscle fibers partially depolarized with 10 mM [K+]o induced transient enhancements of the resting [Ca2+]i that were greater than the transients observed in the normally polarized muscle. In partially depolarized fibers microinjected with 0.3, 0.5 and 1 microM InsP3, the [Ca2+]i was changed to 1.45 +/- 0.14 microM (n = 20), to 3.37 +/- 0.34 microM (n = 7) and to 7.43 +/- 0.70 microM (n = 6) respectively. In all partially depolarized fibers these increments in [Ca2+]i were associated with local contraction.(ABSTRACT TRUNCATED AT 250 WORDS)     

Discrepancy in stoichiometry of steady-state muscle [Pi] and [PCr] induced by exercise

Phosphorus nuclear magnetic resonance was used to quantify the relations between metabolic phosphates, intracellular pH, and work rate in forearm muscle of six adult men over a range of work rates from 1.0 to 3.5 W. Three work rates were studied in each of four sessions (either 1.0, 2.0, and 3.0 or 1.5, 2.5, and 3.5 W), with measurements made before and during each bout, thereby permitting the partition of the variance attributable to rest, work-dependent, and time-dependent metabolic functions by regression analysis. There were no time-dependent changes in either [ATP] or intracellular [H+] as assessed during the rest intervals between bouts of exercise. In contrast, the total nuclear magnetic resonance (NMR)-visible phosphorus pool (TVPP) decreased with time, with both phosphocreatine (PCr) and inorganic phosphate (Pi) contributing significantly to TVPP reduction. Muscle [ATP] was unchanged by work at all intensities. Intracellular [H+] increased moderately and proportionately to work rate. [PCr] decreased and [Pi] increased in proportion to work rate, with the work-dependent coefficient for PCr consumption approximately 1.5 times that of Pi production. Neither Pi line width nor motion artifact accounted for the decrease in TVPP, so the reduced Pi accumulation in exercise may represent its sequestering in some NMR-invisible muscle pool and/or loss to the blood. Whatever the process involved, it is proportional to work rate and persists for at least 10-15 min after exercise.     

Muscle power and metabolism in maximal intermittent exercise

Muscle power and the associated metabolic changes in muscle were investigated in eight male human subjects who performed four 30-s bouts of maximal isokinetic cycling at 100 rpm, with 4-min recovery intervals. In the first bout peak power and total work were (mean +/- SE) 1,626 +/- 102 W and 20.83 +/- 1.18 kJ, respectively; muscle glycogen decreased by 18.2 mmol/kg wet wt, lactate increased to 28.9 +/- 2.7 mmol/kg, and there were up to 10-fold increases in glycolytic intermediates. External power and work decreased by 20% in both the second and third exercise periods, but no further change occurred in the fourth bout. Muscle glycogen decreased by an additional 14.8 mmol/kg after the second exercise and thereafter remained constant. Muscle adenosine triphosphate (ATP) was reduced by 40% from resting after each exercise period; creatine phosphate (CP) decreased successively to less than 5% of resting; in the recovery periods ATP and CP increased to 76 and 95% of initial resting levels, respectively. Venous plasma glycerol increased linearly to 485% of resting; free fatty acids did not change. Changes in muscle glycogen, lactate, and glycolytic intermediates suggested rate limitation at phosphofructokinase during the first and second exercise periods, and phosphorylase in the third and fourth exercise periods. Despite minimal glycolytic flux in the third and fourth exercise periods, subjects generated 1,000 W peak power and sustained 400 W for 30 s, 60% of the values recorded in the first exercise period.(ABSTRACT TRUNCATED AT 250 WORDS)     

A vagally mediated response to metabolic acidosis in anesthetized rabbits

Pentobarbital sodium-anesthetized rabbits received 10-min infusions of acetic, lactic, or propionic acid delivered via a catheter to the right atrium at a rate of 1 mmol/min (n = 14). Arterial [H+] increased by 35.8 +/- 7.6 (SD) nmol/l, a decrease in pH of 0.27 +/- 0.04. By the end of the infusion period respiratory frequency (f), tidal volume (VT), and minute ventilation (V) had increased by 15.5 +/- 6.2 breaths/min, 7.3 +/- 2.7 ml, and 0.86 +/- 0.34 l/min, respectively. Arterial PCO2 (PaCO2) increased initially, but isocapnia was established during the latter half of the infusion (delta PaCO2 = 0.4 +/- 2.0 Torr). Bilateral cervical vagotomy eliminated the f response to acid infusions (n = 9, delta f = 0.6 +/- 2.4 breaths/min). The increase in VT (12.6 +/- 3.1 ml) was greater, but that in V (0.39 +/- 0.11 l/min) was less than in intact animals (P less than 0.05). PaCO2 remained elevated throughout the infusion (delta PaCO2 = 5.5 +/- 2.6 Torr), resulting in a greater rise in arterial [H+] (delta[H+]a = 53.6 +/- 6.6 nmol/l, delta pHa = -0.37 +/- 0.04). It is concluded that vagal afferents play a role in the f response to acute metabolic acidosis in rabbits.     

Muscle metabolites and performance during high-intensity, intermittent exercise

Six men werestudied during four 30-s "all-out" exercise bouts on anair-braked cycle ergometer. The first three exercise bouts wereseparated by 4 min of passive recovery; after the third bout, subjectsrested for 4 min, exercised for 30 min at 30-35% peakO₂ consumption, and rested for afurther 60 min before completing the fourth exercise bout. Peak powerand total work were reduced (P < 0.05) during bout 3 [765 ± 60 (SE) W; 15.8 ± 1.0 kJ] compared withbout 1 (1,168 ± 55 W, 23.8 ± 1.2 kJ), but no difference in exercise performance was observed betweenbouts 1 and4 (1,094 ± 64 W, 23.2 ± 1.4 kJ). Before bout 3, muscle ATP,creatine phosphate (CP), glycogen, pH, and sarcoplasmic reticulum (SR)Ca²⁺ uptake were reduced, whilemuscle lactate and inosine 5'-monophosphate wereincreased. Muscle ATP and glycogen before bout4 remained lower than values beforebout 1 (P < 0.05), but there were no differences in muscle inosine 5'-monophosphate, lactate, pH, and SR Ca²⁺ uptake. Muscle CP levelsbefore bout 4 had increased aboveresting levels. Consistent with the decline in muscle ATP wereincreases in hypoxanthine and inosine before bouts3 and 4. The decline in exercise performance does not appear to be related to a reduction inmuscle glycogen. Instead, it may be caused by reduced CP availability, increased H⁺ concentration,impairment in SR function, or some other fatigue-inducing agent.

     

Use of a microfluorometric method for measuring free calcium in the cytoplasm of isolated cultured rat cardiomyocytes

Dual wavelength microfluorometry was used to measure the cytoplasmic free calcium concentration [( Ca2+]in) in single cultured cells from ventricular myocytes of neonatal rats loaded with the indicator fura-2. At 2.5 nmol/l extracellular Ca2+ in the resting cells [Ca2+]in was between 80 and 110 nmol/l. Sometimes, spontaneous low-frequency (approximately 0.1 Hz) [Ca2+]in oscillations were observed. High-potassium depolarization led to a Ca2+-antagonists-sensitive rise of [Ca2+]in. Both caffeine++ (5-10 mmol/l) and thymol (lmmol/l) initialized transient increase of [Ca2+]in. Mechanisms of [Ca2+]in homeostasis in heart muscle cells were discussed.     

Epinephrine-induced changes in muscle carbohydrate metabolism during exercise in male subjects

Epinephrine increases glycogenolysis in resting skeletal muscle, but less is known about the effects of epinephrine on exercising muscle. To study this, epinephrine was given intraarterially to one leg during two-legged cycle exercise in nine healthy males. The epinephrine-stimulated (EPI) and non-stimulated (C) legs were compared with regard to glycogen, glucose, glucose 6-phosphate (G6P), alpha-glycerophosphate (alpha-GP), and lactate contents in muscle biopsies taken before and after the 45-min submaximal exercise, as well as brachial arterial-femoral venous (a-fv) differences for epinephrine, norepinephrine, lactate, glucose, and O2 during exercise. During exercise the arterial plasma epinephrine concentration was 4.8 +/- 0.8 nmol/l and the femoral venous epinephrine concentrations were 10.3 +/- 2.1 and 3.9 +/- 0.6 nmol/l, respectively, in the EPI and C leg. During exercise the a-fv difference for lactate was greater (-0.41 +/- 0.14 vs. -0.21 +/- 0.14 mmol/l; P less than 0.001), and the a-fv difference for glucose was smaller (0.07 +/- 0.12 vs. 0.24 +/- 0.12 mmol/l; P less than 0.01) in the EPI than in the C leg, but the a-fv differences for O2 were similar. Muscle glycogen depletion (137 +/- 63 vs. 99 +/- 43 mmol/kg dry muscle; P less than 0.1) and the muscle concentrations of glucose (P less than 0.05), alpha-GP (P less than 0.1), G6P (P greater than 0.1), and lactate (P greater than 0.1) tended to be higher in the EPI than the C leg after exercise. These findings suggest that physiological concentrations of epinephrine may enhance muscle glycogenolysis during submaximal exercise in male subjects.     

Enhanced plasma IL-6 and IL-1ra responses to repeated vs. single bouts of prolonged cycling in elite athletes.

The impact of repeated bouts of exercise on plasma levels of interleukin (IL)-6 and IL-1 receptor antagonist (IL-1ra) was examined. Nine well-trained men participated in four different 24-h trials: Long [two bouts of exercise, at 0800-0915 and afternoon exercise (Ex-A), separated by 6 h]; Short (two bouts, at 1100-1215 and Ex-A, separated by 3 h); One (single bout performed at the same Ex-A as second bout in prior trials); and Rest (no exercise). All exercise bouts were performed on a cycle ergometer at 75% of maximal O(2) uptake and lasted 75 min. Peak IL-6 observed at the end of Ex-A was significantly higher in Short (8.8 +/- 1.3 pg/ml) than One (5.2 +/- 0.7 pg/ml) but not compared with Long (5.9 +/- 1.2 pg/ml). Peak IL-1ra observed 1 h postexercise was significantly higher in Short (1,774 +/- 373 pg/ml) than One (302 +/- 53 pg/ml) but not compared with Long (1,276 +/- 451 pg/ml). We conclude that, when a second bout of endurance exercise is performed after only 3 h of recovery, IL-6 and IL-1ra responses are elevated. This may be linked to muscle glycogen depletion.     

Sarcoplasmic ionic calcium concentration in neuroleptic malignant syndrome

The neuroleptic malignant syndrome (NMS) is an uncommon but serious adverse effect of antipsychotic medication. Similarities in the clinical picture, and muscle alterations, between NMS and susceptibility to malignant hyperthermia (MH) suggest common mechanisms underlying both disorders. Sarcoplasmic ionic calcium concentration ([Ca2+]i) was measured by means of Ca2+ selective microelectrodes in intact intercostal muscle fibers isolated from NMS patients and from subjects with no evidence of neuromuscular disease, who served as controls. The mean resting membrane potential and [Ca2+]i were -84 +/- 0.4 mV and 0.11 +/- 0.01 microM (mean +/- SEM) in the control subjects, while they were -84 +/- 0.6 mV and 0.51 +/- 0.02 microM in NMS muscle fibers. Only the difference in [Ca2+]i is significant (P less than 0.001). The incubation of control and NMS muscle bundles in dantrolene (10(-6) M) induced a reduction of [Ca2+]i to 0.06 +/- 0.01 microM and 0.20 +/- 0.04 microM respectively. These results show an alteration in sarcoplasmic ionic [Ca2+] in NMS muscle fibers, suggesting that a dysfunction in skeletal muscle plays some role in the pathogenesis of NMS.     

Preconditioning improves function and recovery of single muscle fibers during severe hypoxia and reoxygenation

Reperfusion following prolonged ischemia induces cellular damage in whole skeletal muscle models. Ischemic preconditioning attenuates the deleterious effects. We tested whether individual skeletal muscle fibers would be similarly affected by severe hypoxia and reoxygenation (H/R) in the absence of extracellular factors and whether cellular damage could be alleviated by hypoxic preconditioning. Force and free cytosolic Ca2+ ([Ca2+]c) were monitored in Xenopus single muscle fibers (n = 24) contracting tetanically at 0.2 Hz during 5 min of severe hypoxia and 5 min of reoxygenation. Twelve cells were preconditioned by a shorter bout of H/R 1 h before the experimental trial. In preconditioned cells, force relative to initial maximal values (P/P(o)) and relative peak [Ca2+]c fell (P < 0.05) during 5 min of hypoxia and recovered during reoxygenation. In contrast, P/P(o) and relative peak [Ca2+]c fell more during hypoxia (P < 0.05) and recovered less during reoxygenation (P < 0.05) in control cells. The ratio of force to [Ca2+]c was significantly higher in the preconditioned cells during severe hypoxia, suggesting that changes in [Ca2+]c were not solely responsible for the loss in force. We conclude that 1) isolated skeletal muscle fibers contracting in the absence of extracellular factors are susceptible to H/R injury associated with changes in Ca2+ handling; and 2) hypoxic preconditioning improves contractility, Ca2+ handling, and cell recovery during subsequent hypoxic insult.     

Prolonged exercise to fatigue in humans impairs skeletal muscle Na+-K+-ATPase activity, sarcoplasmic reticulum Ca2+ release, and Ca2+ uptake.

Prolonged exhaustive submaximal exercise in humans induces marked metabolic changes, but little is known about effects on muscle Na+-K+-ATPase activity and sarcoplasmic reticulum Ca2+ regulation. We therefore investigated whether these processes were impaired during cycling exercise at 74.3 +/- 1.2% maximal O2 uptake (mean +/- SE) continued until fatigue in eight healthy subjects (maximal O2 uptake of 3.93 +/- 0.69 l/min). A vastus lateralis muscle biopsy was taken at rest, at 10 and 45 min of exercise, and at fatigue. Muscle was analyzed for in vitro Na+-K+-ATPase activity [maximal K+-stimulated 3-O-methylfluorescein phosphatase (3-O-MFPase) activity], Na+-K+-ATPase content ([3H]ouabain binding sites), sarcoplasmic reticulum Ca2+ release rate induced by 4 chloro-m-cresol, and Ca2+ uptake rate. Cycling time to fatigue was 72.18 +/- 6.46 min. Muscle 3-O-MFPase activity (nmol.min(-1).g protein(-1)) fell from rest by 6.6 +/- 2.1% at 10 min (P <0.05), by 10.7 +/- 2.3% at 45 min (P <0.01), and by 12.6 +/- 1.6% at fatigue (P <0.01), whereas 3[H]ouabain binding site content was unchanged. Ca2+ release (mmol.min(-1).g protein(-1)) declined from rest by 10.0 +/- 3.8% at 45 min (P <0.05) and by 17.9 +/- 4.1% at fatigue (P < 0.01), whereas Ca2+ uptake rate fell from rest by 23.8 +/- 12.2% at fatigue (P=0.05). However, the decline in muscle 3-O-MFPase activity, Ca2+ uptake, and Ca2+ release were variable and not significantly correlated with time to fatigue. Thus prolonged exhaustive exercise impaired each of the maximal in vitro Na+-K+-ATPase activity, Ca2+ release, and Ca2+ uptake rates. This suggests that acutely downregulated muscle Na+, K+, and Ca2+ transport processes may be important factors in fatigue during prolonged exercise in humans.     

Intracellular Ca2+ requirement for activation of the Na+/H+ exchanger in vascular smooth muscle cells

The role for intracellular Ca2+ in modulating activity of the Na+/H+ exchanger was studied in cultured vascular smooth muscle cells. Na+/H+ exchange was activated by four distinct stimuli: 1) phorbol 12-myristate 13-acetate, 2) thrombin, 3) cell shrinkage, and 4) intracellular acid loading. [Ca2+]i was independently varied between 40 and 200 nM by varying the bathing Ca2+ from 10 nM to 5.0 mM. Thrombin-induced intracellular Ca2+ transients were blocked with bis(2-amino-5-methylphenoxy)ethane-N,N,N',N'-tetraacetic acid tetraacetoxymethyl ester (MAPTAM). In the absence of stimulators of Na+/H+ exchange, varying [Ca2+]i above or below the basal level of 140 nM did not activate Na+/H+ exchange spontaneously. However, varying [Ca2+]i did affect stimulus-induced activation of Na+/H+ exchange. Activation of the exchanger by phorbol 12-myristate 13-acetate was blunted by reduced intracellular Ca2+ (half-maximal activity at 50-90 nM [Ca2+]i), consistent with a Ca2+ requirement for protein kinase C (Ca2+/phospholipid-dependent enzyme). Activation of the exchanger by thrombin in protein kinase C-depleted cells was also sensitive to reduced intracellular Ca2+ (half-maximal activity at 90-140 nM [Ca2+]i) and was increased 40% by raising [Ca2+]i to 200 nM. Activation of the exchanger by cell shrinkage or intracellular acid loads was not significantly affected over the range of [Ca2+]i tested. Thus, altered [Ca2+]i does not itself affect Na+/H+ exchange activity in vascular smooth muscle but instead modulates activation of the transporter by particular stimuli.