Effects of prior exercise and a low-carbohydrate diet on muscle sarcoplasmic reticulum function during cycling in women.

The effects of exercise and diet on sarcoplasmic reticulum Ca(2+)-cycling properties in female vastus lateralis muscle were investigated in two groups of women following four different conditions. The conditions were 4 days of a low-carbohydrate (Lo CHO) and glycogen-depleting exercise plus a Lo CHO diet (Ex + Lo CHO) (experiment 2) and 4 days of normal CHO (Norm CHO) and glycogen-depleting exercise plus Norm CHO (Ex + Norm CHO) (experiment 1). Peak aerobic power (Vo2peak)) was 38.1 +/- 1.4 (SE); n = 9 and 35.6 +/- 1.4 ml.kg(-1).min(-1); n = 9, respectively. Sarcoplasmic reticulum properties measured in vitro in homogenates (micromol.g protein(-1).min(-1)) indicated exercise-induced reductions (P < 0.05) in maximal Ca(2+)-ATPase activity (0 > 30, 60 min > fatigue), Ca(2+) uptake (0 > 30 > 60 min, fatigue), and Ca(2+) release, both phase 1 (0, 30 > 60 min, fatigue) and phase 2 (0 > 30, 60 min, fatigue; 30 min > fatigue) in Norm CHO. Exercise was without effect in altering the Hill slope (n(H)), defined as the slope of relationship between Ca(2+)-ATPase activity and Ca(2+) concentration. No differences were observed between Norm CHO and Ex+Norm CHO. Compared with Norm CHO, Lo CHO resulted in a lower (P < 0.05) Ca(2+) uptake, phase 1 Ca(2+) release (30 min), and n(H). Ex + Lo CHO resulted in a greater (P < 0.05) Ca(2+) uptake and n(H) compared with Lo CHO. The results demonstrate that Lo CHO alone can disrupt SR Ca(2+) cycling and that, with the exception of Ca(2+) release, a glycogen-depleting session of exercise before Lo CHO can reverse the effects.     

Influence of a low-carbohydrate diet on thermoregulatory responses to prolonged exercise in men

Tympanic temperature (T_ty), mean skin temperature (T_sk) and dynamics of sweating were measured in male subjects, performing prolonged, bicycle-exercise. The exercise was performed under thermoneutral environment after the normal-mixed (C) and low-carbohydrate (LCHO) diet. No difference in changes of T_ty was observed after the C and LCHO diets. Sweating was triggered much faster after the LCHO diet. The delay in onset and inertia time of sweating was shorter after the LCHO diet. It is concluded that LCHO diet affects the thermoregulatory responses to exercise by acceleration of sweating, without noticeable differences in body temperature.     

Manipulation of dietary carbohydrates after prolonged effort modifies muscle sarcoplasmic reticulum responses in exercising males

The hypothesis tested was that disturbances in the sarcoplasmic reticulum (SR) Ca2+-cycling responses to exercise would associate with muscle glycogen reserves. Ten untrained males [peak O2 consumption (VO2 peak) = 3.41 +/- 0.20 (SE) l/min] performed a standardized cycle test (approximately 70% VO2 peak) on two occasions, namely, following 4 days of a high (Hi CHO)- and 4 days of a low (Lo CHO)-carbohydrate diet. Both Hi CHO and Lo CHO were preceded by a session of prolonged exercise designed to deplete muscle glycogen. SR Ca2+ cycling in crude homogenates prepared from vastus lateralis samples indicated higher (P < 0.05) Ca2+ uptake (microM x g protein(-1) x min(-1)) in Hi CHO compared with Lo CHO at 30 min (2.93 +/- 0.10 vs. 2.23 +/- 0.12) and at 67 min (2.77 +/- 0.16 vs. 2.10 +/- 0.12) of exercise, the point of fatigue in Lo CHO. Similar effects (P < 0.05) were noted between conditions for maximal Ca2+-ATPase (microM x g protein(-1) x min(-1)) at 30 min (142 +/- 8.5 vs. 107 +/- 5.0) and at 67 min (130 +/- 4.5 vs. 101 +/- 4.7). Both phase 1 and phase 2 Ca2+ release were 23 and 37% higher (P < 0.05) at 30 min of exercise and 15 and 34% higher (P < 0.05), at 67 min during Hi CHO compared with Lo CHO, respectively. No differences between conditions were observed at rest for any of these SR properties. Total muscle glycogen (mmol glucosyl units/kg dry wt) was higher (P < 0.05) in Hi CHO compared with Lo CHO at rest (+36%), 30 min (+53%), and at 67 min (+44%) of cycling. These results indicate that exercise-induced reductions in SR Ca2+-cycling properties occur earlier in exercise during low glycogen states compared with high glycogen states.     

Effects of exercise on muscle metabolites and sarcoplasmic reticulum function in ovariectomized rats

The relationships between exercise and metabolites as well as between exercise and sarcoplasmic reticulum function were studied in gastrocnemius muscle of ovariectomized-trained rats. Prolonged moderate-intensity exercise, treadmill up-hill run for 90 min with a 10 degree incline, decreased the muscle glycogen content. Exercise until exhaustion further lowered the glycogen concentration to 13% of the control, together with a significant decrease of ATP and glucose-6-phosphate concentrations. Also, Ag+-induced Ca2+ release, measured in whole muscle homogenate, showed a 30% reduction on exhaustion, while Ca2+ uptake was unaffected by this exercise. ATPase activities, of both homogenate and SR vesicles, and Ca2+ transport in the latter preparation were not altered on exhaustion. It could be concluded from these results that muscular fatigue in ovariectomized rats after aerobic exercise is caused by the change in energy supply and Ca2+ release from the SR, this latter possibly due to metabolites generated by the exercise.     

Metabolic and sarcoplasmic reticulum Ca2+ cycling responses in human muscle 4 days following prolonged exercise

This study investigated the effects of prolonged exercise on muscle sarcoplasmic reticulum (SR) Ca2+ cycling properties and the metabolic responses with and without a session of exercise designed to reduce muscle glycogen reserves while on a normal carbohydrate (CHO) diet. Eight untrained males (VO(2peak) = 3.81 +/- 0.12 L/min, mean +/- SE) performed a standardized cycle-to-fatigue at 55% VO(2peak) while on a normal CHO diet (Norm CHO) and 4 days following prolonged exercise while on a normal CHO diet (Ex+Norm CHO). Compared to rest, exercise in Norm CHO to fatigue resulted in significant reductions (p < 0.05) in Ca2+ uptake (3.17 +/- 0.21 vs. 2.47 +/- 0.12 micromol.(g protein)-1.min-1), maximal Ca2+ ATPase activity (Vmax, 152 +/- 12 vs. 119 +/- 9 micromol.(g protein)-1.min-1) and both phase 1 (15.1 +/- 0.98 vs. 13.1 +/- 0.28 micromol.(g protein)-1.min-1) and phase 2 (6.56 +/- 0.33 vs. 4.91 +/- 0.28 micromol.(g protein)-1.min-1) Ca2+ release in vastus lateralis muscle. No differences were observed between Norm CHO and Ex-Norm CHO in the response of these properties to exercise. Compared with Norm CHO, Ex+Norm CHO resulted in higher (p < 0.05) resting Ca2+ uptake (3.17 +/- 0.21 vs. 3.49 +/- 0.24 micromol.(g protein).min-1 and higher ionophore ratio, defined as the ratio of Vmax measured with and without the Ca2+-ionophore A23187, (2.3 +/- 0.3 vs. 4.4 +/- 0.3 micromol.(g protein).min-1) at fatigue. No differences were observed between conditions in the concentration of muscle glycogen, the high-energy phosphates (ATP and PCr), or metabolites (Pi, Cr, and lactate). Ex+Norm CHO also failed to modify the exercise-induced changes in CHO and fat oxidation. We conclude that prolonged exercise to fatigue performed 4 days following glycogen-depleting exercise while on a normal CHO diet elevates resting Ca2+ uptake and prevents increases in SR membrane permeability to Ca2+ as measured by the ionophore ratio.     

Comparative aspects of cardiac and skeletal muscle sarcoplasmic reticulum.

While differing in numerous physiological and biochemical parameters, mammalian cardiac and skeletal muscles exhibit many common ultrastructural characteristics. General subcellular organization is similar with longitudinal disposition and organization of the myofibrils as well as subcellular organelles such as mitochondria, sarcoplasmic reticulum and transverse tubules. Significant differences are more readily discerned in terms of degree, not only with respect to relative amounts of various organelles, but also in regard to membrane composition. It is these macromolecular variations in membrane components which may, at least in part, provide the basis for differences in overall functional characteristics in the muscles.In cardiac, as well as skeletal muscle, the concentration of Ca²⁺ ions at specific intracellular sites regulates the contractile state of the muscle. The differences in mechanism and sources of Ca²⁺ for contraction in cardiac and skeletal muscle are but a few of the unsolved areas which are now being addressed. We shall focus primarily on research advances involving cardiac and skeletal SR emphasizing the contrasting features related to their functional roles in control of contraction and metabolic events.     

Effects of progressive exercise and hypoxia on human muscle sarcoplasmic reticulum function.

This study examined the effects of progressive exercise to fatigue in normoxia (N) on muscle sarcoplasmic reticulum (SR) Ca(2+) cycling and whether alterations in SR Ca(2+) cycling are related to the blunted peak mechanical power output (PO(peak)) and peak oxygen consumption (Vo(2 peak)) observed during progressive exercise in hypoxia (H). Nine untrained men (20.7 +/- 0.42 yr) performed progressive cycle exercise to fatigue on two occasions, namely during N (inspired oxygen fraction = 0.21) and during H (inspired oxygen fraction = 0.14). Tissue extracted from the vastus lateralis before exercise and at power output corresponding to 50 and 70% of Vo(2 peak) (as determined during N) and at fatigue was used to investigate changes in homogenate SR Ca(2+)-cycling properties. Exercise in H compared with N resulted in a 19 and 21% lower (P < 0.05) PO(peak) and Vo(2 peak), respectively. During progressive exercise in N, Ca(2+)-ATPase kinetics, as determined by maximal activity, the Hill coefficient, and the Ca(2+) concentration at one-half maximal activity were not altered. However, reductions with exercise in N were noted in Ca(2+) uptake (before exercise = 357 +/- 29 micromol x min(-1) x g protein(-1); at fatigue = 306 +/- 26 micromol x min(-1) x g protein(-1); P < 0.05) when measured at free Ca(2+) concentration of 2 microM and in phase 2 Ca(2+) release (before exercise = 716 +/- 33 micromol x min(-1) x g protein(-1); at fatigue = 500 +/- 53 micromol x min(-1) x g protein(-1); P < 0.05) when measured in vitro in whole muscle homogenates. No differences were noted between N and H conditions at comparable power output or at fatigue. It is concluded that, although structural changes in SR Ca(2+)-cycling proteins may explain fatigue during progressive exercise in N, they cannot explain the lower PO(peak) and Vo(2 peak) observed during H.     

Human muscle sarcoplasmic reticulum function during submaximal exercise in normoxia and hypoxia.

In this study, the response of the sarcoplasmic reticulum (SR) to prolonged exercise, performed in normoxia (inspired O(2) fraction = 0.21) and hypoxia (inspired O(2) fraction = 0.14) was studied in homogenates prepared from the vastus lateralis muscle in 10 untrained men (peak O(2) consumption = 3.09 +/- 0.25 l/min). In normoxia, performed at 48 +/- 2.2% peak O(2) consumption, maximal Ca(2+)-dependent ATPase activity was reduced by approximately 25% at 30 min of exercise compared with rest (168 +/- 10 vs. 126 +/- 8 micromol.g protein(-1) x min(-1)), with no further reductions observed at 90 min (129 +/- 6 micromol x g protein(-1) x min(-1)). No changes were observed in the Hill coefficient or in the Ca(2+) concentration at half-maximal activity. The reduction in maximal Ca(2+)-dependent ATPase activity at 30 min of exercise was accompanied by oxalate-dependent reductions (P < 0.05) in Ca(2+) uptake by approximately 20% (370 +/- 22 vs. 298 +/- 25 micromol x g protein(-1) x min(-1)). Ca(2+) release, induced by 4-chloro-m-cresol and assessed into fast and slow phases, was decreased (P < 0.05) by approximately 16 and approximately 32%, respectively, by 90 min of exercise. No differences were found between normoxia and hypoxia for any of the SR properties examined. It is concluded that the disturbances induced in SR Ca(2+) cycling with prolonged moderate-intensity exercise in human muscle during normoxia are not modified when the exercise is performed in hypoxia.     

Pressure effects on sarcoplasmic reticulum

The irreversible effects of pressure (1-2000 atm) upon the enzymatic activity and structure of the Ca2+-ATPase of sarcoplasmic reticulum were investigated. Sarcoplasmic reticulum vesicles suspended in a medium of 0.1 M KCl, 10 mM imidazole, pH 7.0, 5 mM MgCl2, and 0.5 mM EGTA irreversibly lose their Ca2+ transport and Ca2+-stimulated ATPase activities on exposure to pressures of 800-2000 atmospheres. The pressure-induced inactivation of Ca2+-ATPase is accompanied by inhibition of the formation of phosphorylated enzyme intermediate, an increase in the passive Ca2+ permeability of the membrane, and structural changes in the Ca2+-ATPase as shown by disruption of Ca2+-ATPase membrane crystals, increased susceptibility to tryptic digestion, unmasking of SH groups, and loss of the conformational responses to Ca2+ and vanadate. The sensitivity to pressure is influenced by enzyme conformation. Ca2+ or vanadate + EGTA protect the Ca2+-ATPase against pressure-induced inactivation, implying a greater stability of the enzyme in the E1 and E2 states than in the conformational equilibrium that prevails at low [Ca2+] in the absence of vanadate. Protection against pressure inactivation was also observed in the presence of sucrose, glycerol, ethylene glycol and 1 M KCl, suggesting that water density modifying groups significantly affect the stability of Ca2+-ATPase under pressure.     

Pyruvate overrides inhibition of PDH during exercise after a low-carbohydrate diet

The effects of carbohydrate deprivation on the regulation of pyruvate dehydrogenase (PDH) were studied at rest and during moderate-intensity exercise. An inhibitory effect of a chronic low-carbohydrate diet (LCD) on the active form of PDH (PDHa) mediated by a stable increase in PDH kinase (PDHK) activity has recently been reported (Peters SJ, Howlett RA, St. Amand TA, Heigenhauser GJF, and Spriet LL. Am J Physiol Endocrinol Metab 275: E980-E986, 1998.). In the present study, seven males cycled at 65% maximal O(2) uptake for 30 min after a 6-day LCD. Exercise was repeated 1 wk later after a mixed diet (MD). Muscle biopsies were sampled from the vastus lateralis at rest and at 2 and 30 min of exercise. At rest, PDHa activity (0.18 +/- 0.04 vs. 0.63 +/- 0.18 mmol x min(-1) x kg wet wt(-1)), muscle glycogen content (310.2 +/- 36.9 vs. 563.9 +/- 32.6 mmol/kg dry wt), and muscle lactate content (2.6 +/- 0.3 vs. 4.2 +/- 0.6 mmol/kg dry wt) were significantly lower after the LCD. Resting muscle acetyl-CoA (10.8 +/- 1.9 vs. 7.4 +/- 0.8 micromol/kg dry wt) and acetylcarnitine (5.3 +/- 1.4 vs. 1.6 +/- 0.3 mmol/kg dry wt) contents were significantly elevated after the LCD. During exercise, PDHa, glycogenolytic rate (LCD 5.8 +/- 0.4 vs. MD 6.9 +/- 0.2 mmol x min(-1) x kg dry wt(-1)), and muscle concentrations of acetylcarnitine, pyruvate, and lactate increased to the same extent in both conditions. The results of the present study suggest that inhibition of resting PDH by elevated PDHK activity after a LCD may be overridden by the availability of muscle pyruvate during exercise.     

The effect of a low-carbohydrate diet on performance, hormonal and metabolic responses to a 30-s bout of supramaximal exercise

The aim of this study was to find out whether a low-carbohydrate diet (L-CHO) affects: (1) the capacity for all-out anaerobic exercise, and (2) hormonal and metabolic responses to this type of exercise. To this purpose, eight healthy subjects underwent a 30-s bicycle Wingate test preceded by either 3 days of a controlled mixed diet (130 kJ/kg of body mass daily, 50% carbohydrate, 30% fat, 20% protein) or 3 days of an isoenergetic L-CHO diet (up to 5% carbohydrate, 50% fat, 45% protein) in a randomized order. Before and during 1 h after the exercise venous blood samples were taken for measurement of blood lactate (LA), β-hydroxybutyrate (β-HB), glucose, adrenaline (A), noradrenaline (NA) and insulin levels. Oxygen consumption (V˙O₂) was also determined. It was found that the L-CHO diet diminished the mean power output during the 30-s exercise bout [533 (7) W vs 581 (7) W, P < 0.05] without changing the maximal power attained during the first or second 5-s interval of the exercise. In comparison with the data obtained after the consumption of a mixed diet, after the consumption of a L-CHO diet resting plasma concentrations of β-HB [2.38 (0.18) vs 0.23 (0.01) mmol · l⁻¹, P < 0.001] and NA [4.81 (0.68) vs 2.2 (0.31) nmol · l⁻¹, P < 0.05] were higher, while glucose [4.6 (0.1) vs 5.7 (0.2) mmol · l⁻¹, P < 0.05] and insulin concentrations [11.9 (0.9) vs 21.8 (1.8) mU · l⁻¹] were lower. The 1-h post-exercise excess of V˙O₂ [9.1 (0.25) vs 10.6 (0.25) l, P < 0.05], and blood LA measured 3 min after the exercise [9.5 (0.4) vs 10.6 (0.5) mmol · l⁻¹, P < 0.05] were lower following the L-CHO treatment, whilst plasma NA and A concentrations reached higher values [2.24 (0.40) vs 1.21 (0.13) nmol · l⁻¹ and 14.30 (1.41) vs 8.20 (1.31) nmol · l⁻¹, P < 0.01, respectively]. In subjects on the L-CHO diet, the plasma β-HB concentration decreased quickly after exercise, attaining ≈30% of the pre-exercise value within 60 min, while insulin and glucose levels were elevated. The main conclusions of this study are: (1) a L-CHO diet is detrimental to anaerobic work capacity, possibly because of a reduced muscle glycogen store and decreased rate of glycolysis; (2) reduced carbohydrate intake for 3 days enhances activity of the sympathoadrenal system at rest and after exercise. Accepted: 31 January 1997     

Effects of continuous low-carbohydrate diet after long-term exercise on GLUT-4 protein content in rat skeletal muscle.

Stimulation of AMPK and decreased glycogen levels in skeletal muscle have a deep involvement in enhanced insulin action and GLUT-4 protein content after exercise training. The present study examined the chronic effects of a continuous low-carbohydrate diet after long-term exercise on GLUT-4 protein content, glycogen content, AMPK, and insulin signaling in skeletal muscle. Rats were divided randomly into four groups: normal chow diet sedentary (N-Sed), low carbohydrate diet sedentary (L-Sed), normal chow diet exercise (N-Ex), and low carbohydrate diet exercise (L-Ex) groups. Rats in the exercise groups (N-Ex and L-Ex) were exercised by swimming for 6 hours/day in two 3-hour bouts separated by 45 minutes of rest. The 10-day exercise training resulted in a significant increase in the GLUT-4 protein content (p<0.01). Additionally, the GLUT-4 protein content in L-Ex rats was increased by 29% above that in N-Ex rats (p<0.01). Finally, the glycogen content in skeletal muscle of L-Ex rats was decreased compared with that of N-Ex rats. Taken together, we suggest that the maintenance of glycogen depletion after exercise by continuous low carbohydrate diet results in the increment of the GLUT-4 protein content in skeletal muscle.     

The effects of chlorbutol on skeletal muscle sarcoplasmic reticulum function in porcine malignant hyperpyrexia

1. Chlorbutol, a muscle relaxant, inhibits the in vitro muscle hypercontractility which is characteristic of the anaesthetic complication, malignant hyperpyrexia (MH). 2. Studies on isolated sarcoplasmic reticulum vesicles have shown that this effect of chlorbutol in MH is not due to a modification of Ca2+-transport mechanisms.     

Post-mortem electrical stimulation of muscle and its effects on sarcoplasmic reticulum adenosine triphosphatase.

Sarcoplasmic reticulum (SR) was isolated from control muscles and from muscles which had been subjected to short-term post-mortem electrical stimulation. Both preparations had similar protein compositions but the SR from electrically stimulated muscle had a lower 'extra' ATPase activity. The ability of the SR preparations from electrically stimulated muscles to accumulate Ca2+ was about the same as the controls. There was, therefore, an apparently greater efficiency of Ca2+ transport in the isolated vesicles, the reason for which is not known, but an alteration in the 'leakiness' of the membrane may be involved. Purified ATPase isolated from control and stimulated SR contained, in addition to the ATPase protein, a polypeptide of molecular weight about 30 000. The purified ATPase vesicles from electrically stimulated muscle had a reduced activity as measured by ATP splitting activity, phosphoenzyme formation from either inorganic orthophosphate (Pi) or ATP, or by an ATP in equilibrium Pi exchange reaction. These reduced activities probably result from an alteration in the binding affinities of the ATPase for ATP and Pi. The low affinity site for calcium binding was not affected by electrical stimulation. Purified ATPase vesicles from stimulated muscle were more susceptible to proteolytic attack, suggesting that the conformation of the protein or its association with the membrane lipids had been altered.     

Effect of a low-carbohydrate diet on plasma and sweat ammonia concentrations during prolonged nonexhausting exercise

The purpose of this investigation was to examine the effect of low body glycogen stores on plasma ammonia concentration and sweat ammonia excretion during prolonged, nonexhausting exercise of moderate intensity. On two occasions seven healthy untrained men pedalled on a cycle ergometer for 60 min at 50% of their predetermined maximal O₂ uptakes ( _max) firstly, following 3 days on a normal mixed diet (N-diet) (60% carbohydrates, 25% fat and 15% protein) and secondly, following 3 days on a low-carbohydrate diet (LC-diet) (less than 5% carbohydrates, 50% fat and 45% protein) of equal energy content. Blood was collected from the antecubital vein immediately before, at 30th and at 60th min of exercise. Sweat was collected from the hypogastric region using gauze pads. It was shown that plasma ammonia concentrations after the LC-diet were higher than after the N-diet at both the 30th and 60th min of exercise. Sweat ammonia concentration and total ammonia loss through the sweat were also higher after the LC-diet. The higher ammonia concentrations in plasma and sweat after the LC-diet would seem to indicate an increased ammonia production, which may be related to reduced initial carbohydrate stores.     

Effects of diet on the function of sarcoplasmic reticulum.

We have examined the effect of diet on the phospholipid composition of the sarcoplasmic reticulum of rabbit muscle. Enriching the diet with corn or fish oil results in significant changes in the fatty acyl chain composition of the various phospholipid classes, with relatively little change in the relative contents of the phospholipids. These alterations in composition have no significant effect on the ATPase activity of vesicles of sarcoplasmic reticulum or on the pattern of Ca2+ uptake and release.     

Characterization of sarcoplasmic reticulum in skinned muscle cultures

The plasma membranes of chick or rat skeletal muscles, grown in cell culture, were made permeable with saponin in a solution lacking calcium. The cells were then supplied with a medium resembling the cytosol and the ATP-dependent Ca2+ sequestration was performed. Based on the low concentration of free Ca2+ in the medium (below 5 microM), the presence of mitochondrial inhibitors and the effect of drugs that interfere with sarcoplasmic reticulum (SR) function, we assume that the measured Ca2+ accumulation expresses SR function on the saponin-treated myotubes. The development of the SR in muscle cultures is augmented as myogenesis proceeds and depends on its occurrence. Whereas creatine kinase activity is elevated immediately following cell fusion, there is a delay of at least 1 day between myoblast fusion and the increase in Ca2+ accumulation in the SR. Thyroxine or triiodothyronine caused an inhibition of Ca2+ accumulation in rat or chick muscle cultures. This inhibition could explain some of the muscle abnormalities caused by excess of thyroid hormones. A comparison was made between a white-type (fast) and heterogeneous muscle, differentiated in cell culture. There was no significant difference in SR function, indicating the important role of innervation in specifying the properties of muscle fiber types.