Dynamic maceration of Matricaria chamomilla inflorescences: optimal conditions for flavonoids and antioxidant activity

ABSTRACT The aim of this paper was to study and optimize the dynamic maceration process to obtain Matricaria chamomilla L., Asteraceae, inflorescences extracts with optimum flavonoid content and antioxidant activity using a multivariate approach. Hydroalcoholic extracts were obtained by dynamic maceration in lab scale and the influence of extraction temperature, ratio of plant to solvent, ethanol strength; extraction time and stirring speed on the flavonoid content and antioxidant activity were unveiled using a fractional factorial design. The ethanol strength, ratio of plant to solvent and temperature were the three factors that influenced most the extract properties and were studied by a central composite design. Total flavonoid content and antioxidant activity were affected by the ethanol strength and ranged from 1.49 to 3.95% and 13.3 to 36.2 µg/ml, respectively. The desirability functions resulted in an optimal dynamic maceration condition using 1 h extraction at stirring speed of 900 rpm, ethanol 74.7%, temperature of 69 °C and using 36.8% of plant in solvent (w/v). Under this set of conditions, the extract had total flavonoid content of 4.11 ± 0.07%, in vitro antioxidant activity with IC50 of 18.19 µg/ml and apigenin and apigenin-7-glycoside contents of 2.0 ± 0.1 mg/g and 20.1 ± 0.9 mg/g, respectively. The results showed a low solvent consumption compared to previous works. The model was able to predict extract properties with maximum deviation of 12% and the extraction process developed herein showed to be reliable, efficient and scalable for M. chamomilla inflorescences, enriched with flavonoids, apigenin and apigenin-7-glycoside and high antioxidant activity.

Eccentric contractions affect muscle membrane phospholipid fatty acid composition in rats.

This study investigated if prior eccentric contractions, and thus mechanical strain and muscle damage, exert an effect on the muscle membrane phospholipid fatty acid composition in rats, and whether a possible effect could be attenuated by dietary supplements. Twenty-three rats were randomised to three groups who received chow with added fish oil (n = 8), vitamin C (n = 8) or no supplement (n = 7). After 3 weeks of feeding, calf muscles on one side were stimulated electrically during anaesthesia causing eccentric contractions. Two days later the white gastrocnemius, a part of the stimulated calf muscle, was excised from both legs. In the muscles stimulated to contract eccentrically, compared to the control muscles, the proportion of arachidonic acid, C20:4,n-6 (17.7 +/- 0.6; 16.4 +/- 0.4% of total fatty acids, respectively) and docosapentanoeic acid, C22:5,n-3 (2.9 +/- 0.1 and 2.7 +/- 0.1% of total fatty acids, respectively) was uniformly higher across groups (P < 0.02) with no differences between diet groups. The proportion of long chain polyunsaturates was also significantly higher in the eccentrically contracted (39.9 +/- 0.6% of total fatty acids) compared to the control leg (38.2 +/- 0.6% of total fatty acids; P < 0.01). In contrast no differences were observed in the fatty acid composition of the triacylglycerols stored within the muscle. Thus one severe bout of eccentric contractions modulates the fatty acid composition of the muscle membrane phospholipids when compared to a control leg, and supplemental intake of fish oil or vitamin C did not attenuate this effect.

Chemokines are elevated in plasma after strenuous exercise in humans.

During the last few years much attention has been paid to the chemokines. Chemokine receptors are necessary to render a target permissive for infection by the human immunodeficiency virus (HIV) and high concentrations of chemokines have been shown to protect against the progression of HIV disease towards death. In the present study, we investigated the capability of strenuous exercise to induce elevated plasma concentrations of the chemokines interleukin (IL)-8, macrophage inflammatory protein (MIP)-1 alpha and MIP-1 beta. Eight male athletes completed the Copenhagen Marathon 1997. Blood was sampled before, immediately after the run and every 30 min during a 4 h recovery period. Plasma chemokine concentrations were measured using enzyme-linked immunosorbent assays. The IL-8, MIP-1 alpha and MIP-1 beta concentrations all peaked 0.5 h after the run when they were 6.7-fold, 3.5-fold and 4.1-fold increased, respectively. The elevated concentrations of chemokines in plasma after exercise could have implications for HIV-infected individuals; a possibility that needs further investigation.

Insulin action on rates of muscle protein synthesis following eccentric, muscle-damaging contractions.

The purpose of this study was to determine whether eccentric, muscle-damaging contractions affect insulin action on muscle protein synthesis. Male Wistar rats (n = 28) were anaesthetized either once or twice separated by 7 days' rest, and one limb was electrically stimulated to contract eccentrically, while the contralateral limb served as a non-stimulated control. Twenty-four and 48 h after contractions, rates of protein synthesis were assessed in soleus and red or white gastrocnemius muscles during a hindlimb perfusion with or without insulin (20 000 microU mL(-1)). Rates of protein synthesis were not different in non-stimulated muscle, with or without insulin (P > 0.05). In red or white gastrocnemius without insulin, rates of protein synthesis were significantly reduced (P < 0.05) 24 and 48 h after a single session and 48 h after a double session of muscle contractions. However, protein synthesis was normalized with insulin 24 and 48 h after contractions in red, and 48 h after contractions in white gastrocnemius. In soleus muscle, protein synthesis was impaired only 48 h after the second session, but partially restored by insulin (P < 0.05). These results indicate that muscle becomes more sensitive to insulin action on rates of protein synthesis after muscle-damaging contractions.

N-3 polyunsaturated fatty acids do not affect cytokine response to strenuous exercise.

The aim of the present study was to investigate whether fish oil supplementation was able to modulate the acute-phase response to strenuous exercise. Twenty male runners were randomized to receive supplementation (n = 10) with 6.0 g fish oil daily, containing 3.6 g n-3 polyunsaturated fatty acids (PUFA), for 6 wk or to receive no supplementation (n = 10) before participating in The Copenhagen Marathon 1998. Blood samples were collected before the race, immediately after, and 1.5 and 3 h postexercise. The fatty acid composition in blood mononuclear cells (BMNC) differed between the fish oil-supplemented and the control group, showing incorporation of n-3 PUFA and less arachidonic acid in BMNC in the supplemented group. The plasma levels of tumor necrosis factor-alpha, interleukin-6, and transforming growth factor-beta(1) peaked immediately after the run, the increase being 3-, 92-, and 1.1-fold, respectively, compared with resting samples. The level of interleukin-1 receptor antagonist peaked 1.5 h after exercise, with the increase being 87-fold. However, the cytokine levels did not differ among the two groups. Furthermore, supplementation with fish oil did not influence exercise-induced increases in leucocytes and creatine kinase. In conclusion, 6 wk of fish oil supplementation had no influence on the acute-phase response to strenuous exercise.

Muscle contractions induce interleukin-6 mRNA production in rat skeletal muscles.

1. The present study explored the hypothesis that interleukin-6 (IL-6) might be locally produced in response to skeletal muscle contractions and whether the production might reflect the type of muscle contraction performed. Rats were anaesthetized and the calf muscles of one limb were stimulated electrically for concentric or eccentric contractions (4 x 10 contractions with 1 min of rest between the 4 series, 100 Hz). The contralateral muscles served as unstimulated controls. The mRNA levels for IL-6, the glucose transport protein GLUT-4 and beta-actin in the rat muscles (white and red gastrocnemius and soleus) were quantified by quantitative competitive RT-PCR. 2. The IL-6 mRNA level, measured 30 min after the stimulation, increased after both eccentric and concentric contractions and there were no significant differences in IL-6 mRNA levels between the different muscle fibre types. No significant increase in IL-6 mRNA level was seen in the unstimulated contralateral muscle fibres. 3. No increase in GLUT-4 mRNA level was detected, indicating that the increase in IL-6 mRNA level was not due to general changes in transcription. 4. We conclude that IL-6 is locally produced after muscle contraction, with no significant differences between different muscle fibre types. This local production of IL-6 is not due to general changes in transcription, since no changes in the level of GLUT-4 mRNA were found. The fact that increased IL-6 mRNA levels were seen after both concentric and eccentric contractions indicates that the production of IL-6 is not solely due to muscle damage, seen primarily after eccentric exercise.

Marathon running transiently increases c-Jun NH2-terminal kinase and p38 activities in human skeletal muscle.

We examined the pattern of activation and deactivation of the stress-activated protein kinase signalling molecules c-Jun NH2-terminal kinase (JNK) and p38 kinase in skeletal muscle in response to prolonged strenuous running exercise in human subjects. Male subjects (n = 14; age 32 +/- 2 years; VO2,max 60 +/- 2 ml kg-1 min-1) completed a 42.2 km marathon (mean race time 3 h 35 min). Muscle biopsies were obtained 10 days prior to the marathon, immediately following the race, and 1, 3 and 5 days after the race. The activation of JNK and p38, including both p38alpha and p38gamma, was measured with immune complex assays. The phosphorylation state of p38 (alpha and gamma) and the upstream regulators of JNK and p38, mitogen-activated protein kinase kinase 4 (MKK4) and mitogen-activated protein kinase kinase 6 (MKK6), were assessed using phosphospecific antibodies. JNK activity increased 7-fold over basal level immediately post-exercise, but decreased back to basal levels 1, 3 and 5 days after the exercise. p38gamma phosphorylation (4-fold) and activity (1.5-fold) increased immediately post-exercise and returned to basal levels at 1, 3 and 5 days following exercise. In contrast, p38alpha phosphorylation and activity did not change over the time course studied. MKK4 and MKK6 phosphorylation increased and decreased in a trend similar to that observed with JNK activity and p38gamma phosphorylation. Prolonged running exercise did not affect JNK, p38alpha, or p38gamma protein expression in the days following the race. This study demonstrates that both JNK and p38 intracellular signalling cascades are robustly, yet transiently increased following prolonged running exercise. The differential activation of the p38 isoforms with exercise in human skeletal muscle indicates that these proteins may have distinct functions in vivo.

Exercise diminishes the activity of acetyl-CoA carboxylase in human muscle.

Studies in rats suggest that increases in fatty acid oxidation in skeletal muscle during exercise are related to the phosphorylation and inhibition of acetyl-CoA carboxylase (ACC), and secondary to this, a decrease in the concentration of malonyl-CoA. Studies in human muscle have not revealed a consistent decrease in the concentration of malonyl-CoA during exercise; however, measurements of ACC activity have not been reported. Thus, whether the same mechanism operates in human muscle in response to physical activity remains uncertain. To investigate this question, ACC was immunoprecipitated from muscle of human volunteers and its activity assayed in the same individual at rest and after one-legged knee-extensor exercise at 60, 85, and 100% of knee extensor VO2max. ACC activity was diminished by 50-75% during exercise with the magnitude of the decrease generally paralleling exercise intensity. Treatment of the immunoprecipitated enzyme with protein phosphatase 2A restored activity to resting values, suggesting the decrease in activity was due to phosphorylation. The measurement of malonyl-CoA in the muscles revealed that its concentration is 1/10 of that in rats, and that it is diminished (12-17%) during the higher-intensity exercises. The respiratory exchange ratio increased with increasing exercise intensity from 0.84 +/- 0.02 at 60% to 0.99 0.04 at 100% VO2max. Calculated rates of whole-body fatty acid oxidation were 121 mg/min at rest and 258 +/- 35, 264 +/- 63, and 174 +/- 76 mg/min at 60, 85, and 100% VO2max, respectively. The results show that ACC activity, and to a lesser extent malonyl-CoA concentration, in human skeletal muscle decrease during exercise. Although these changes may contribute to the increases in fat oxidation from rest to exercise, they do not appear to explain the shift from mixed fuel to predominantly carbohydrate utilization when exercise intensity is increased.

Time pattern of exercise-induced changes in type I collagen turnover after prolonged endurance exercise in humans.

Type I collagen is known to adapt to physical activity, and biomarkers of collagen turnover indicate that synthesis can be influenced by a single intense exercise bout, but the exact time pattern of these latter changes are largely undescribed. In the present study, 17 healthy young males had their plasma concentrations of the carboxyterminal propeptide of type I procollagen (PICP), a marker of collagen formation, and the immunoactive carboxyterminal cross-linked telopeptide (ICTP), a marker of collagen resorption, measured before and immediately postexercise, as well as 1, 2, 3, 4, 5, and 6 days after completion of a marathon run (42 km). Serum concentrations of creatine kinase (S-CK) were measured as an indicator of muscular breakdown in response to the exercise bout. After a transient decrease in collagen formation immediately after exercise (plasma PICP concentration: 176 +/- 17 microg/liter to 156 +/- 9 microg/liter)(P < 0.05), concentrations rose in the days following the marathon, peaked 72 hours after exercise (197 +/- 8 microg/liter)(P < 0.05 versus basal), and returned to basal values similar to those 5 days postexercise (170 +/- 10 microg/liter). Apart from a short increase immediately after exercise, collagen resorption did not change from basal levels throughout the remaining period (P > 0.05). Muscle breakdown was elevated during the days following the exercise and peaked 24 hours after the exercise (S-CK concentration: 3,133 +/- 579 U/liter). The findings in the present study indicate that type I collagen synthesis is accelerated in response to prolonged strenuous exercise, reaching a peak after 3 days and returning to preexercising levels 5 days after the completion of a marathon run.

Glycogen concentration in human skeletal muscle: effect of prolonged insulin and glucose infusion.

To study the upper limit of glycogen storage in human muscle, two healthy male subjects were infused with glucose and insulin for 8 h reaching plasma concentrations of about 21 mM glucose and approximately 2000 microU/ml insulin. Prior to the infusion subjects performed for 1 h one-legged knee-extensor exercise at 75% of their maximum one-legged work capacity in order to lower muscle glycogen stores in one leg. During the 8-h hyperglycemic clamp procedure, glycogen concentrations increased and levelled off at 2- and 5-fold above the pre-infusion levels in the resting and the working leg, respectively. However, the absolute glycogen levels reached in both legs were quite similar, close to 4 g per 100 g wet muscle (about 1000 mumol/g d.w.), independent of prior exercise. Previous studies have shown that glycogen levels, after a bout of glycogen-depleting exercise and subsequent ingestion of a carbohydrate-rich diet for 3 days, can be increased to values around 3-4 g per 100 g wet muscle. It appears that the maximal attainable glycogen concentration in human muscle seems to be close 4 g per 100 g wet muscle. This glycogen level can thus be reached either by a prolonged infusion of supra-physiological concentrations of glucose and insulin or by glycogen-depleting exercise followed by ingestion of a carbohydrate-rich diet.

Pro- and anti-inflammatory cytokine balance in strenuous exercise in humans.

1. The present study investigates to what extent and by which time course prolonged strenuous exercise influences the plasma concentration of pro-inflammatory and inflammation responsive cytokines as well as cytokine inhibitors and anti-inflammatory cytokines. 2. Ten male subjects (median age 27.5 years, range 24-37) completed the Copenhagen Marathon 1997 (median running time 3 : 26 (h : min), range 2 : 40-4 : 20). Blood samples were obtained before, immediately after and then every 30 min in a 4 h post-exercise recovery period. 3. The plasma concentrations of tumour necrosis factor (TNF)alpha, interleukin (IL)-1beta, IL-6, IL-1ra, sTNF-r1, sTNF-r2 and IL-10 were measured by enzyme-linked immunosorbent assay (ELISA). The highest concentration of IL-6 was found immediately after the race, whereas IL-1ra peaked 1 h post exercise (128-fold and 39-fold increase, respectively, as compared with the pre-exercise values). The plasma level of IL-1beta, TNFalpha, sTNF-r1 and sTNF-r2 peaked in the first hour after the exercise (2. 1-, 2.3-, 2.7- and 1.6-fold, respectively). The plasma level of IL-10 showed a 27-fold increase immediately post exercise. 4. In conclusion, strenuous exercise induces an increase in the pro-inflammatory cytokines TNFalpha and IL-1beta and a dramatic increase in the inflammation responsive cytokine IL-6. This is balanced by the release of cytokine inhibitors (IL-1ra, sTNF-r1 and sTNF-r2) and the anti-inflammatory cytokine IL-10. The study suggests that cytokine inhibitors and anti-inflammatory cytokines restrict the magnitude and duration of the inflammatory response to exercise.

Muscle glycogen accumulation after a marathon: roles of fiber type and pro- and macroglycogen.

Muscle glycogen remains subnormal several days after muscle damaging exercise. The aims of this study were to investigate how muscle acid-soluble macroglycogen (MG) and acid-insoluble proglycogen (PG) pools are restored after a competitive marathon and also to determine whether glycogen accumulates differently in the various muscle fiber types. Six well-trained marathon runners participated in the study, and muscle biopsies were obtained from the vastus lateralis of the quadriceps muscle before, immediately after, and 1, 2, and 7 days (days 1, 2, and 7, respectively) after the marathon. During the race, 56 +/- 3.8% of muscle glycogen was utilized, and a greater fraction of MG (72 +/- 3.7%) was utilized compared with PG (34 +/- 6.5%). On day 2, muscle glycogen and MG values remained lower than prerace values, despite a carbohydrate-rich diet, but they had both returned to prerace levels on day 7. The PG concentration was lower on day 1 compared with before the race, whereas there were no significant differences between the prerace PG concentration and the concentrations on days 2 and 7. On day 2 the glycogen concentration was particularly low in the type I fibers, indicating that local processes are important for the accumulation pattern. We conclude that a greater fraction of human muscle MG than of PG is utilized during a marathon and that accumulation of MG is particularly delayed after the prolonged exercise bout. Furthermore, factors produced locally appear important for the glycogen accumulation pattern.

Competitive sustained exercise in humans, lymphokine activated killer cell activity, and glutamine--an intervention study.

This study examined whether oral glutamine supplementation abolishes some of the exercise-induced changes in lymphocyte functions following long-term intense exercise. A group of 16 marathon runners participating in The Copenhagen Marathon 1996 were placed randomly in either a placebo (n = 7) or a glutamine receiving group (n = 9). Each subject received four doses of either placebo or glutamine (100 mg x kg(-1)) administered at 0, 30, 60, and 90-min post-race. In the placebo group the plasma glutamine concentrations were lower than pre-race values during the post-exercise period [mean 647 (SEM 32) compared to 470 (SEM 22) micromol x 1(-1) 90-min post-race, P < 0.05] whereas glutamine supplementation maintained the plasma glutamine concentration (at approximately 750 micromol x 1(-1)). Glutamine supplementation in vivo had no effect on the lymphokine activated killer (LAK) cell activity, the proliferative responses or the exercise-induced changes in concentrations or percentages of any of the leucocyte subpopulations examined. Glutamine addition in in vitro studies enhanced the proliferative response in both groups. These data would suggest that decreased plasma glutamine concentrations post-exercise are not responsible for exercise-induced decrease in LAK activity and that the influence of glutamine in vitro is not dependent on the plasma glutamine concentration at the time of sampling.

Training effects on muscle glucose transport during exercise.

Muscle glucose uptake is increased during exercise compared to rest. In general, muscle glucose uptake increases with increasing exercise intensity and duration. Whereas the arterio-venous concentration difference only increases 2-4-fold during exercise compared with rest the increase in muscle perfusion in 10-20 times and therefore quantitatively very important. During exercise the surface membrane glucose transport capacity increases in skeletal muscle primarily due to an increase in surface membrane GLUT4 protein content. Endurance training decreases muscle glucose uptake during exercise at a given absolute submaximal work-load despite a large increase in muscle GLUT4 protein content. We have shown that this decrease in glucose uptake at least in part is due to a blunted exercise-induced increase in sarcolemmal glucose transport capacity secondary to a blunted increase in sarcolemmal GLUT4 transporter number. Thus, endurance training leads to a marked reduction of the fraction of muscle GLUT4 that is translocated during a given submaximal exercise bout. Whether this is true also during exercise at higher intensities remains to be seen.

Evidence that interleukin-6 is produced in human skeletal muscle during prolonged running.

1. This study was performed to test the hypothesis that inflammatory cytokines are produced in skeletal muscle in response to prolonged intense exercise. Muscle biopsies and blood samples were collected from runners before, immediately after, and 2 h after a marathon race. 2. The concentration of interleukin (IL)-6 protein in plasma increased from 1.5 +/- 0.7 to 94.4 +/- 12.6 pg ml-1 immediately post-exercise and to 22.1 +/- 3.8 pg ml-1 2 h post-exercise. IL-1 receptor antagonist (IL-1ra) protein in plasma increased from 123 +/- 23 to 2795 +/- 551 pg ml-1, and increased further to 4119 +/- 527 pg ml-1 2 h post-exercise. 3. The comparative polymerase chain reaction technique was used to evaluate mRNA for IL-6, IL-1ra, IL-1beta and tumour necrosis factor (TNF)-alpha in skeletal muscle and blood mononuclear cells (BMNC) (n = 8). Before exercise, mRNA for IL-6 could not be detected either in muscle or in BMNC, and was only detectable in muscle biopsies (5 out of 8) after exercise. Increased amounts of mRNA for IL-1ra were found in two muscle biopsies and five BMNC samples, and increased amounts of IL-1beta mRNA were found in one muscle and four BMNC samples after exercise. TNF-alpha mRNA was not detected in any samples. 4. This study suggests that exercise-induced destruction of muscle fibres in skeletal muscles may trigger local production of IL-6, which stimulates the production of IL-1ra from circulating BMNC.

Exercise metabolism in human skeletal muscle exposed to prior eccentric exercise.

1. The effects of unaccustomed eccentric exercise on exercise metabolism during a subsequent bout of graded concentric exercise were investigated in seven healthy male subjects. Arterial and bilateral femoral venous catheters were inserted 2 days after eccentric exercise of one thigh (eccentric thigh) and blood samples were taken before and during graded two-legged concentric knee-extensor exercise. Muscle biopsies were obtained from the eccentric and control vastus lateralis before (rest) and after (post) the concentric exercise bout. 2. Maximal knee-extensor concentric exercise capacity was decreased by an average of 23 % (P < 0.05) in the eccentric compared with the control thigh. 3. The resting muscle glycogen content was lower in the eccentric thigh than in the control thigh (402 +/- 30 mmol (kg dry wt)-1 vs. 515 +/- 26 mmol (kg dry wt)-1, means +/- s.e.m., P < 0.05), and following the two-legged concentric exercise this difference substantially increased (190 +/- 46 mmol (kg dry wt)-1 vs. 379 +/- 58 mmol (kg dry wt)-1, P < 0.05) despite identical power and duration of exercise with the two thighs. 4. There was no measurable difference in glucose uptake between the eccentric and control thigh before or during the graded two-legged concentric exercise. Lactate release was higher from the eccentric thigh at rest and, just before termination of the exercise bout, release of lactate decreased from this thigh (suggesting decreased glycogenolysis), whereas no decrease was found from the contralateral control thigh. Lower glycerol release from the eccentric thigh during the first, lighter part of the exercise (P < 0.05) suggested impaired triacylglycerol breakdown. 5. At rest, sarcolemmal GLUT4 glucose transporter content and glucose transport were similar in the two thighs, and concentric exercise increased sarcolemmal GLUT4 content and glucose transport capacity similarly in the two thighs. 6. It is concluded that in muscle exposed to prior eccentric contractions, exercise at a given power output requires a higher relative workload than in undamaged muscle. This increases utilization of the decreased muscle glycogen stores, contributing to decreased endurance.

Effect of prior eccentric contractions on lactate/H+ transport in rat skeletal muscle.

The effect of prior eccentric contractions on skeletal muscle lactate/H+ transport was investigated in rats. Lactate transport was measured in sarcolemmal giant vesicles obtained from soleus and red (RG) and white gastrocnemii (WG) muscles 2 days after intense eccentric contractions (ECC) and from the corresponding contralateral control (CON) muscles. The physiochemical buffer capacity was determined in the three muscle types from both ECC and CON legs. Furthermore, the effect of prior eccentric contractions on release and muscle content of lactate and H+ during and after supramaximal stimulation was examined using the perfused rat hindlimb preparation. The lactate transport rate was lower (P < 0.05) in vesicles obtained from ECC-WG (29%) and ECC-RG (13%) than in vesicles from the CON muscles. The physiochemical buffer capacity was reduced (P < 0.05) in ECC-WG (13%) and ECC-RG (9%) compared with the corresponding CON muscles. There were only marginal effects on the soleus muscle. Muscle lactate concentrations and release of lactate during recovery from intense isometric contractions were lower (P < 0.05) in ECC than in CON hindlimbs, indicating decreased anaerobic glycogenolysis. In conclusion, the sarcolemmal lactate/H+ transport capacity and the physiochemical buffer capacity were reduced in prior eccentrically stimulated WG and RG in rats, suggesting that muscle pH regulation may be impaired after unaccustomed eccentric exercise. In addition, the data indicate that the glycogenolytic potential is decreased in muscles exposed to prior eccentric contractions.

Prior eccentric contractions impair maximal insulin action on muscle glucose uptake in the conscious rat.

Our aim was to examine the effect of prior eccentric contractions on insulin action locally in muscle in the intact conscious rat. Anesthetized rats performed one-leg eccentric contractions through the use of calf muscle electrical stimulation followed by stretch of the active muscles. Two days later, basal and euglycemic clamp studies were conducted with the rats in the awake fasted state. Muscle glucose metabolism was estimated from 2-[14C(U)]deoxy-D-glucose and D-[3-3H] glucose administration, and comparisons were made between the eccentrically stimulated and nonstimulated (control) calf muscles. At midphysiological insulin levels, effects of prior eccentric exercise on muscle glucose uptake were not statistically significant. Maximal insulin stimulation revealed reduced incremental glucose uptake above basal (P < 0.05 in the red gastrocnemius; P < 0.1 in the white gastrocnemius and soleus) and impaired net glycogen synthesis in all eccentrically stimulated muscles (P < 0.05). We conclude that prior eccentric contractions impair maximal insulin action (responsiveness) on local muscle glucose uptake and glycogen synthesis in the conscious rat.

Impaired muscle glycogen resynthesis after a marathon is not caused by decreased muscle GLUT-4 content.

Our purpose was to investigate whether the slow rate of muscle glycogen resynthesis after a competitive marathon is associated with a decrease in the total muscle content of the muscle glucose transporter (GLUT-4). Seven well-trained marathon runners participated in the study, and muscle biopsies were obtained from the lateral head of the gastrocnemius muscle before, immediately after, and 1, 2, and 7 days after the marathon, as were venous blood samples. Muscle GLUT-4 content was unaltered over the experimental period. Muscle glycogen concentration was 758 +/- 53 mmol/kg dry weight before the marathon and decreased to 148 +/- 39 mmol/kg dry weight immediately afterward. Despite a carbohydrate-rich diet (containing at least 7 g carbohydrate.kg body mass-1.day-1), the muscle glycogen concentration remained 30% lower than before-race values 2 days after the race, whereas it had returned to before-race levels 7 days after the race. We conclude that the total GLUT-4 protein content is unaltered in the lateral gastrocnemius after a competitive marathon and that the slow recovery of muscle glycogen after the race apparently involves factors other than changes in the total content of this protein.

Decreased insulin action on muscle glucose transport after eccentric contractions in rats.

We have recently shown that eccentric contractions (Ecc) of rat calf muscles cause muscle damage and decreased glycogen and glucose transporter GLUT-4 protein content in the white (WG) and red gastrocnemius (RG) but not in the soleus (S) (S. Asp, S. Kristiansen, and E. A. Richter. J. Appl. Physiol. 79: 1338-1345, 1995). To study whether these changes affect insulin action, hindlimbs were perfused at three different insulin concentrations (0, 200, and 20,000 microU/ml) 2 days after one-legged eccentric contractions of the calf muscles. Compared with control, basal glucose transport was slightly higher (P < 0.05) in Ecc-WG and -RG, whereas it was lower (P < 0.05) at both submaximal and maximal insulin concentrations in the Ecc-WG and at maximal concentrations in the Ecc-RG. In the Ecc-S, the glucose transport was unchanged in hindquarters perfused in the absence or presence of a submaximal stimulating concentration of insulin, whereas it was slightly (P < 0.05) higher during maximal insulin stimulation compared with control S. At the end of perfusion the glycogen concentrations were lower in both Ecc-gastrocnemius muscles compared with control muscles at all insulin concentrations. Fractional velocity of glycogen synthase increased similarly with increasing insulin concentrations in Ecc- and control WG and RG. We conclude that insulin action on glucose transport but not glycogen synthase activity is impaired in perfused muscle exposed to prior eccentric contractions.