E. Hultman

Glycogen, Glycolytic Intermediates and High-Energy Phosphates Determined in Biopsy Samples of Musculus Quadriceps Femoris of Man at Rest. Methods and Variance of Values

Methods are described for the determination of glycogen, glycolytic intermediates, and high-energy phosphates in muscle biopsy samples. Initial freezedrying of samples and extraction of metabolites with relatively weak acid are preferred. Normal values in muscle are similar to those found by other workers. Variation in muscle content of ATP, ATP + ADP + AMP, phosphorylcreatine (PC), creatine (Cr), PC + Cr, and glycogen, between legs, between sites on the same muscle, or as a result of error introduced during analysis, was small compared with the between-individuals variance. The importance of the different sources of variance on taking a biopsy is discussed.

The time course of phosphorylcreatine resynthesis during recovery of the quadriceps muscle in man.

SummaryThe time course of phosphorylcreatine (PC) resynthesis in the human m. quadriceps femoris was studied during recovery from exhaustive dynamic exercise and from isometric contraction sustained to fatigue. The immediate postexercise muscle PC content after either form of exercise was 15–16% of the resting muscle content. The time course of PC resynthesis during recovery was biphasic exhibiting a fast and a slow recovery component. The half-time for the fast component was 21–22 s but this accounted for a smaller fraction of the total PC restored during recovery from the isometric contraction than after the dynamic exercise. The half-time for the slow component was in each case more than 170 s. After 2 and 4 min recovery the total amounts of PC resynthesized after the isometric exercise were significantly lower than from the dynamic exercise.Occlusion of the circulation to the quadriceps completely abolished the resynthesis of PC. Restoration of resynthesis occurred only after release of occlusion.

Resynthesis of creatine phosphate in human muscle after exercise in relation to intramuscular pH and availability of oxygen

After exhaustive exercise the muscular store of creatine phosphate (CP) is almost completely depleted. The resynthesis of CP during recovery normally occurs rapidly, but is totally inhibited if the local circulation to the muscle is occluded. The limiting factor for CP resynthesis which could be a low intramuscular pH or availability of oxygen has been investigated in the present study. Biopsies from musculis quadriceps femoris of man were analyzed for pH, ATP, ADP, CP, creatine, lactate and pyruvate. It was shown that resynthesis of CP only occurs when the blood supply to the muscle is intact. From this it was concluded that the creatine kinase reaction is at a steady state or at equilibrium during the period of recovery. The influence of oxygen on the resynthesis of CP was investigated by incubating muscle samples taken after a fatiguing isometric contraction in atmospheres of oxygen and nitrogen, respectively. During 15 min incubation in oxygen CP was resynthesized from a starting value of 4% to 68% of the normal value at rest. No resynthesis was observed when parallel muscle samples were incubated for the same time in nitrogen. It is suggested that the initial fast phase of CP resynthesis is limited by the availability of oxygen whereas the subsequent slow phase is limited by the hydrogen ion transport out from the muscle.

Clinical Physiology: The Effect of Circulatory Occlusion on Isometric Exercise Capacity and Energy Metabolism of the Quadriceps Muscle in Man

Occlusion of the circulation to the quadriceps muscle for 20 min resulted in decreases in the muscle ATP and phosphorylcreatine (PC) contents of 1 and 32 per cent, respectively, and increases in ADP and AMP of 7 and 37 per cent. Decrease in PC was statistically significant after 4 min of occlusion, suggesting that the local intramuscular oxygen store was sufficiently depleted at this time as to be limiting to normal mitochondrial function. Pyruvate and lactate concentrations in muscle and the lactate to pyruvate ratio were significantly increased after 15 min of occlusion but not before. The calculated local oxygen store in the muscle was 2.0 mmol O2 - (kg dry muscle)-1. Local oxygen store depletion after 4 and 10 min of occlusion was estimated to be 40-50 and 90-100 per cent complete. Increasing time of pre-exercise occlusion resulted in decreased isometric endurance capacity. The observed decreases in endurance, however, were far greater than could be accounted for by any parallel decrease in the local muscle energy stores.

Energy Rich Phosphagens in Dynamic and Static Work

The understanding of muscle metabolism and its regulation in situ has been greatly advanced in recent years by the analysis of the levels of metabolic intermediates under differing conditions. Only recently has it been possible to perform similar studies in mail. In the present studies muscle samples have been obtained using the needle biopsy technique (1, 10, 11) and the levels of a number of metabolites in these have been determined by enzymatic micro-methods.

Glycolytic intermediates in human muscle after isometric contraction

Isometric contraction of the quadriceps muscle sustained to fatigue with a force of 66% of the maximum voluntary contraction force resulted in a mean glycogen utilization of 80.4 (S.D. 58.4) mmol glucosyl units/kg dry muscle (d.m.) and an accumulation of glycolytic intermediates and glucose corresponding to 82.9 (S.D. 17.5) mmol glucosyl units/kg d.m. Accumulation of hexose phosphates (principally glucose 6-phosphate) accounted for 35.4% (S.D. 4.1) of the total increase and lactate for 59.3% (S.D. 2.8). During a 4 min recovery period glucose 6-[hosphate content showed a linear decrease with a half time of 2.0 min and lactate decreased exponentially with a half time of 2.5 min. The rate of lactate disappearance from the muscle was approximately 4 times as fast as that observed previously after maximal bicycle exercise. This was probably due to a lower lactate concentration in blood after isometric contraction resulting in a larger muscle-blood gradient for lactate. Muscle content of free glucose was increased after contraction and increased further during recovery. It is concluded that the glucose increase is confined to the intracellular pool and is an effect of hexokinase inhibition by accumulated glucose 6-phosphate. Occlusion, of the local circulation after the contraction inhibited the recovery processes for lactate and glucose 6-phosphate.

Enzyme activities in hepatic venous blood under strenuous physical exercise

SummaryIn order to study the influence of physical exercise on liver function, experiments were performed in healthy volunteers subjected to controlled major physical effort. Blood samples obtained by catheterization of hepatic vein during exercise were analyzed for activities of LD (thermostabile and thermolabile isoenzymes), AIAT, SDH, ICD, AP and CPK, and these activities were compared with corresponding values in arterial blood samples taken simultaneously. Hepatic blood flow and oxygen saturation of hepatic venous blood were measured. Physical exercise resulted both in diminished hepatic blood flow and a fall in hepatic venous oxygen saturation. These changes were accompanied by release of liver specific enzymes, indicating that exhausting exercise may induce an “increased hepatocyte membrane permeability” in man.

Isometric Exercise - Factors Influencing Endurance and Fatigue

In two healthy male subjects we have made some preliminary observations of the factors influencing endurance and fatigue during successive sustained contractions at two thirds the intensity of a maximal voluntary contraction of the Quadriceps Muscle. In all experiments subjects sat in an adjustable chair (5), measurements being made with a strain gauge attached to the ankle when the knee joint was flexed to 90°. Fig. 1 shows the relationship between the duration of successive contractions and the interval between contractions, both under normal conditions and when the circulation to the leg was occluded using a sphygmomanometer cuff inflated to 250 mm Hg, immediately prior to starting the series. Each subject was allowed at least 24 hours recovery between each experiment. In the absence of circulatory occlusion more than seven holds were possible, the duration of successive holds being significantly related (p =0.016) to the recovery interval. With circulatory occlusion no more than three holds could be sustained, the duration of the second and third holds being unrelated to the recovery interval. As observed by othere (3, 4) the duration of the first hold was not significantly reduced by prior circulatory occlusion suggesting that muscle blood flow was practically arrested at this relative load.

The effect of Cobalt on mitochondrial ATP-production in the rat myocardium and skeletal muscle

Cobalt has been shown to accumulate in the myocardium of uraemic patients and has been suggested as a myocardial toxin inhibiting mitochondrial respiration. In order to study the cellular effects of cobalt exposure three groups of rats (n = 12 per group) were fed a diet containing 12% protein without supplementation or with 20 mg and 40 mg CoSO4 7 H2O/kg body weight/day respectively. After 8 weeks the hearts and soleus muscles were removed. Cobalt in tissues and in four cell fractions were analysed with neutron-activation analysis (ng/g wet weight and ng/mg protein respectively). Mitochondrial respiration was analysed as ATP-production rate using pyruvate + malate and palmitoyl-carnitine + malate as substrate. The ATP-production from pyruvate + malate was unchanged in both heart and skeletal muscle in the exposed animals. With palmitate as substrate, the heart muscle showed a slightly lower ATP-production rate (p less than 0.05) after the 20 mg cobalt dose, but the rate was unchanged in the group with higher cobalt intake. No changes in ATP-production rate from palmitate was observed in soleus muscle. The microsomal (100,000 g) fraction in the myocardial cells contained significantly higher cobalt concentrations compared to the mitochondrial fraction in both the unexposed (1.4 ng/mg protein vs 0.19, p less than 0.05) and exposed rats (53.4 ng/mg protein vs 13.2, p less than 0.005). In conclusion, cobalt showed a large accumulation in myocardial cells, without significant effects on mitochondrial ATP-formation rate from oxidation of pyruvate or palmitate and with the highest cobalt content contained in the microsomal (100,000 g) fraction.