Diffusional constraints on energy metabolism in skeletal muscle

Aerobic metabolic flux depends on the diffusion of high-energy phosphate molecules (e.g., ATP and phosphocreatine) from the mitochondria to cellular ATPases, as well as the diffusion of other molecules (e.g., ADP, Pi) back to the mitochondria. Here, we develop an approach for evaluating the influence of intracellular metabolite diffusion on skeletal muscle aerobic metabolism through the application of the effectiveness factor (η). This parameter provides an intuitive and informative means of quantifying the extent to which diffusion limits metabolic flux. We start with the classical approach assuming an infinite supply of substrate at the fiber boundary, and we expand this model to ultimately include nonlinear boundary and homogeneous reactions. Comparison of the model with experimental data from a wide range of skeletal muscle types reveals that most muscle fibers are not substantially limited by diffusion (η close to unity), but many are on the brink of rather substantial diffusion limitation. This implies that intracellular metabolite diffusion does not dramatically limit aerobic metabolic flux in most fibers, but it likely plays a role in limiting the evolution of muscle fiber design and function.     

Myofibrillar tightly bound calcium in skeletal muscle fibers: a possible role of this cation in titin strands aggregation

In muscle cells, part of the calcium is tightly bound to the N1- and N2-line of the sarcomere but its physiological significance was unknown. In the present work we reported the ability of a recombinant titin fragment spanning titin domains Z9 to I1 to tightly bind calcium ions with a K(d) of 0.049+/-0.004 nM. We further showed that calcium induced a spontaneous aggregation of the titin fragment and that the major aggregate is a tetramer. The implication of these findings on the organization of the six titin strands that emanate from the end of the thick filament within the I-band is discussed.     

The possible role of L-carnitine on the skeletal muscle of ovariectomized rats

Low muscle strength is observed during the peri-and postmenopausal periods, when the secretion of ovarian hormones is drastically reduced. It is also a predictive of adverse health events as well as incident mobility limitation and disability. The objective of the present study is to study the biochemical and the histological changes in the skeletal muscle of premature menopause-induced rats and the possible protective role of L-carnitine. Ovariectomized (OVX) rats were gavaged with L-carnitine (100 mg/kg) daily for 60 days starting from the second post-operative day. Serum levels of estradiol and markers of skeletal muscle damage (creatine kinase and lactic dehydrogenase activities) were determined. Light and electron microscopic study of the quadriceps femoris muscle (QFM) specimens were done. OVX rats showed significant decrease in the serum estradiol level with significant increase the markers for skeletal muscle damage. Histopathological examination of the QFM showed degenerated myofibers, apoptotic changes and compensatory hypertrophy. Degenerated mitochondria, multiple lysosomes and lipid droplets among the damaged myofibrils were also noticed. L-carnitine administration to the OVX rats resulted in non-significant change in the serum estradiol level with significant attenuation of skeletal muscle damage either biochemically or histopathologically. In conclusion, L-carnitine administration recovered muscle degeneration after ovariectomy. This finding suggested that L-carnitine could be recommended in the management of post-menopausal myopathy.     

Effects of muscle activation on fatigue and metabolism in human skeletal muscle.

Increasing stimulation frequency has been shown to increase fatigue but not when the changes in force associated with changes in frequency have been controlled. An effect of frequency, independent of force, may be associated with the metabolic cost resulting from the additional activations. Here, two separate experiments were performed on human medial gastrocnemius muscles. The first experiment (n = 8) was designed to test the effect of the number of pulses on fatigue. The declines in force during two repetitive, 150-train stimulation protocols that produced equal initial forces, one using 80-Hz trains and the other using 100-Hz trains, were compared. Despite a difference of 600 pulses (23.5%), the protocols produced similar rates and amounts of fatigue. In the second experiment, designed to test the effect of the number of pulses on the metabolic cost of contraction, 31P-NMR spectra were collected (n = 6) during two ischemic, eight-train stimulation protocols (80- and 100-Hz) that produced comparable forces despite a difference of 320 pulses (24.8%). No differences were found in the changes in P(i) concentration, phosphocreatine concentration, and intracellular pH or in the ATP turnover produced by the two trains. These results suggest that the effect of stimulation frequency on fatigue is related to the force produced, rather than to the number of activations. In addition, within the range of frequencies tested, increasing total activations did not increase metabolic cost.     

Phospholipases and phagocytosis: the role of phospholipid-derived second messengers in phagocytosis

Phagocytosis, the process by which leukocytes recognize and destroy invading pathogens, is essential for host defense. The binding of foreign organisms to phagocytic leukocytes initiates a complex signaling cascade which ultimately results in the entrapment and destruction of the pathogen. The signal transduction pathway mediating phagocytosis is the subject of intense investigation and is known to include protein tyrosine kinases, GTP-binding proteins, protein kinase C (PKC), actin polymerization and membrane movement. A rapidly expanding body of evidence suggests that phospholipases play an integral role in phagocytosis by generating essential second messengers. Here we review the data linking activation of phospholipase A2 (PLA2), phospholipase C (PLC) phospholipase D (PLD), and phosphoinositide 3-OH kinase (PI(3)K) to antibody (IgG)-mediated phagocytosis. Evidence is presented that (1) PLA2-derived arachidonic acid (AA) stimulates NADPH oxidase and membrane redistribution during phagocytosis, (2) the inositol-3,4,5-triphosphate (IP3) and diacylglycerol (DAG) products of PLC activate NADPH oxidase and PKC, and (3) sequential activation of PLD and phosphatidic acid phosphohydrolase may provide an alternative pathway for generation of DAG. Additionally, considerable evidence exists that wortmannin, a PI(3)K inhibitor, depresses phagocytosis. This finding is discussed in the context of the extensive effects PI(3)K products have on endocytosis and exocytosis and the potential role of membrane redistribution in phagocytosis. Finally, a model is presented which integrates data obtained from a variety of phagocytic systems and illustrates potential interactions that may exist between phospholipase-derived second messengers and signaling events required for phagocytosis.     

Interrelationship of oxidative metabolism and local perfusion demonstrated by NMR in human skeletal muscle

Toussaint, Jean-François, Kenneth K. Kwong, FidelisM'Kparu, Robert M. Weisskoff, Paul J. LaRaia, and Howard L. Kantor. Interrelationship of oxidative metabolism and local perfusion demonstrated by NMR in human skeletal muscle. J. Appl.Physiol. 81(5): 2221-2228, 1996.Using nuclearmagnetic resonance (NMR), we have examined the relationship ofhigh-energy phosphate metabolism and perfusion in human soleus andgastrocnemius muscles. With ³¹P-NMR spectroscopy, we monitoredphosphocreatine (PCr) decay and recovery in eight normal volunteers andfour heart failure patients performing ischemic plantar flexion. Byusing echo-planar imaging, perfusion was independently measured by alocal [inversion-recovery (T1-flow)] and a regionaltechnique (NMR-plethysmography). After correction for its pHdependence, PCr recovery time constant is 27.5 ± 8.0 s innormal volunteers, with mean flow 118 ± 75 (soleus andgastrocnemius T1-flow) and 30.2 ± 9.7 ml ^· 100 ml^{1 ·} min¹(NMR-plethysmography-flow). We demonstrate a positive correlation between PCr time constant and local perfusion given byy = 50  0.15x(r² = 0.68, P = 0.01) for the 8 normal subjects,and y = 64  0.24x (r² = 0.83, P = 0.0001) for the 12 subjectsrecruited in the study. Regional perfusion techniques also show asignificant but weaker correlation. Using this totally noninvasivemethod, we conclude that aerobic ATP resynthesis is related to themagnitude of perfusion, i.e., O₂availability, and demonstrate that magnetic resonance imaging andmagnetic resonance spectroscopy together can accurately assess musclefunctional status.

     

The role of lipid second messengers in aldosterone synthesis and secretion

Second messengers are small rapidly diffusing molecules or ions that relay signals between receptors and effector proteins to produce a physiological effect. Lipid messengers constitute one of the four major classes of second messengers. The hydrolysis of two main classes of lipids, glycerophospholipids and sphingolipids, generate parallel profiles of lipid second messengers: phosphatidic acid (PA), diacylglycerol (DAG), and lysophosphatidic acid versus ceramide, ceramide-1-phosphate, sphingosine, and sphingosine-1-phosphate, respectively. In this review, we examine the mechanisms by which these lipid second messengers modulate aldosterone production at multiple levels. Aldosterone is a mineralocorticoid hormone responsible for maintaining fluid volume, electrolyte balance, and blood pressure homeostasis. Primary aldosteronism is a frequent endocrine cause of secondary hypertension. A thorough understanding of the signaling events regulating aldosterone biosynthesis may lead to the identification of novel therapeutic targets. The cumulative evidence in this literature emphasizes the critical roles of PA, DAG, and sphingolipid metabolites in aldosterone synthesis and secretion. However, it also highlights the gaps in our knowledge, such as the preference for phospholipase D-generated PA or DAG, as well as the need for further investigation to elucidate the precise mechanisms by which these lipid second messengers regulate optimal aldosterone production.     

Effects of corticosteroid on the transport and metabolism of glutamine in rat skeletal muscle

Intramuscular glutamine falls with injury and disease in circumstances associated with increases in blood corticosteroids. We have investigated the effects of corticosteroid administration (0.44 mg/kg dexamethasone daily for 8 days, 200 g female rats) on intramuscular glutamine and Na⁺, muscle glutamine metabolism and sarcolemmal glutamine transport in the perfused hindlimb. After dexamethasone treatment intramuscular glutamine fell by 45% and Na⁺ rose by 25% (the respective muscle/plasma distribution ratios changed from 8.6 to 4.5 and 0.12 to 0.15); glutamine synthetase and glutaminase activities were unchanged at 475 ± 75 and 60 ± 19 nmol/g muscle per min. Glutamine output by the hindlimb of anaesthetized rats was increased from 31 to 85 nmol/g per min. Sarcolemmal glutamine transport was studied by paired-tracer dilution in the perfused hindlimb: the maximal capacity (V_max) for glutamine transport into muscle (by Na⁺-glutamine symport) fell from 1058 ± 310 to 395 ± 110 nmol/g muscle per min after dexamethasone treatment, accompanied by a decrease in the K_m (from 8.1 ± 1.9 to 2.1 ± 0.4 mM glutamine). At physiological plasma glutamine concentration (0.75 mM) dexamethasone appeared to cause a proportional increase in sarcolemmal glutamine efflux over influx. Addition of dexamethasone (200 nM) to the perfusate of control rat hindlimbs caused acute changes in V_max and K_m of glutamine transport similar to those resulting from 8-day dexamethasone treatment. The reduction in muscle glutamine concentration after dexamethasone treatment may be primarily due to a reduction in the driving force for intramuscular glutamine accumulation, i.e., in the Na⁺ electrochemical gradient. The prolonged increase in muscle glutamine output after dexamethasone treatment (which occurs despite a reduction in the size of the intramuscular glutamine pool) appears to be due to a combination of (a) accelerated sarcolemmal glutamine efflux and (b) increased intramuscular synthesis of glutamine.     

Effects of raising muscle glycogen synthesis rate on skeletal muscle ATP turnover rate in type 2 diabetes

Mitochondrial dysfunction has been implicated in the pathogenesis of type 2 diabetes. We hypothesized that any impairment in insulin-stimulated muscle ATP production could merely reflect the lower rates of muscle glucose uptake and glycogen synthesis, rather than cause it. If this is correct, muscle ATP turnover rates in type 2 diabetes could be increased if glycogen synthesis rates were normalized by the mass-action effect of hyperglycemia. Isoglycemic- and hyperglycemic-hyperinsulinemic clamps were performed on type 2 diabetic subjects and matched controls, with muscle ATP turnover and glycogen synthesis rates measured using (31)P- and (13)C-magnetic resonance spectroscopy, respectively. In diabetic subjects, hyperglycemia increased muscle glycogen synthesis rates to the level observed in controls at isoglycemia [from 19 ± 9 to 41 ± 12 μmol·l(-1)·min(-1) (P = 0.012) vs. 40 ± 7 μmol·l(-1)·min(-1) in controls]. This was accompanied by a modest increase in muscle ATP turnover rates (7.1 ± 0.5 vs. 8.6 ± 0.7 μmol·l(-1)·min(-1), P = 0.04). In controls, hyperglycemia brought about a 2.5-fold increase in glycogen synthesis rates (100 ± 24 vs. 40 ± 7 μmol·l(-1)·min(-1), P = 0.028) and a 23% increase in ATP turnover rates (8.1 ± 0.9 vs. 10.0 ± 0.9 μmol·l(-1)·min(-1), P = 0.025) from basal state. Muscle ATP turnover rates correlated positively with glycogen synthesis rates (r(s) = 0.46, P = 0.005). Changing the rate of muscle glucose metabolism in type 2 diabetic subjects alters demand for ATP synthesis at rest. In type 2 diabetes, skeletal muscle ATP turnover rates reflect the rate of glucose uptake and glycogen synthesis, rather than any primary mitochondrial defect.     

Relationship between erythrocyte aggregate size and flow rate in skeletal muscle venules

In previous studies we showed that intravenous infusion of Dextran 500 in the rat causes blunting of the velocity profile of red blood cells in venules at low shear rates. To determine whether this blunting is associated with the formation of red blood cell aggregates, we measured the length and width of particles in the venular flow stream at systemic hematocrits up to 20% with a high-speed video camera and a new image analysis technique. Data were obtained at various shear rates under normal (nonaggregating) conditions as well as after infusion of Dextran 500. Under normal conditions, particle length (parallel to the vessel axis) was 6.5 +/- 2.7 microm and width (perpendicular to the axis) was 6.1 +/- 1.7 microm, in agreement with published dimensions of individual red blood cells for this species. After Dextran 500 infusion, particle length and width increased significantly to 8.7 +/- 5.1 and 10.4 +/- 4.4 microm, respectively. Particle dimensions were greater in the central region of the flow stream for both normal and dextran-treated blood and increased at low flow rates with dextran-treated blood. This study provides direct confirmation of aggregate formation at low shear in venules with high-molecular-weight dextran as well as an estimate of aggregate size and range.     

有氧运动对衰老大鼠骨骼肌线粒体能量代谢的影响

目的：探讨有氧运动对衰老大鼠骨骼肌线粒体能量代谢的影响。方法：将20只12月龄的雌性Wistar大鼠随机分为老年安静组（AC,n=10）及老年运动组（AE,n=10）,另取10只2月龄的雌性Wistar大鼠为青年安静组（YC,n=10）;安静组大鼠进行正常饲养,运动组大鼠进行坡度为5°,速度为15.2 m/min,第1天运动15 min、第2天运动30 min、从第3天开始每天运动45 min,每周6 d,共12周。12周后所有大鼠断头处死,取腓肠肌样本,差速离心法提取线粒体,测定SOD和GSH-Px活性、MDA含量、三羧酸循环限速酶（CS、ICD和α-KGDHC）活性及呼吸链酶复合体（RCCⅠ~Ⅳ）活性。结果：①与YC组相比,AC组骨骼肌线粒体SOD活性和MDA含量显著增加（P<0.05）,CS和α-KGDHC活性均显著降低（P<0.05）,RCCⅠ、RCCⅡ和RCCⅣ活性均显著下降（P<0.05）,RCCⅢ活性显著升高（P<0.05）;AE组骨骼肌线粒体SOD、GSH-Px活性和MDA含量均显著增加（P<0.01）,CS、ICD和α-KGDHC活性均显著升高（P<0.01）,RCCⅠ~Ⅳ活性均显著升高（P<0.01）。②与AC组相比,AE组骨骼肌线粒体SOD、GSH-Px活性均显著升高（P<0.05）,MDA含量显著下降（P<0.05）,CS、ICD、α-KGDHC和RCCⅠ~Ⅳ活性均显著升高（P<0.01）。结论：有氧运动可以提高老年大鼠骨骼肌线粒体抗氧化能力,降低脂质过氧化水平,提高三羧酸循环及呼吸链功能,促进线粒体能量代谢,延缓衰老过程中线粒体的退行性变化。     

Influence of intensity of food restriction on skeletal muscle mitochondrial energy metabolism in rats

Variable durations of food restriction (FR; lasting weeks to years) and variable FR intensities are applied to animals in life span-prolonging studies. A reduction in mitochondrial proton leak is suggested as a putative mechanism linking such diet interventions and aging retardation. Early mechanisms of mitochondrial metabolic adaptation induced by FR remain unclear. We investigated the influence of different degrees of FR over 3 days on mitochondrial proton leak and mitochondrial energy metabolism in rat hindlimb skeletal muscle. Animals underwent 25, 50, and 75% and total FR compared with control rats. Proton leak kinetics and mitochondrial functions were investigated in two mitochondrial subpopulations, intermyofibrillar (IMF) and subsarcolemmal (SSM) mitochondria. Regardless of the degree of restriction, skeletal muscle mass was not affected by 3 days of FR. Mitochondrial basal proton conductance was significantly decreased in 50% restricted rats in both mitochondrial subpopulations (46 and 40% for IMF and SSM, respectively) but was unaffected in other groups compared with controls. State 3 and uncoupled state 3 respiration rates were decreased in SSM mitochondria only for 50% restricted rats when pyruvate + malate was used as substrate (-34.5 and -38.9% compared with controls, P < 0.05). IMF mitochondria respiratory rates remained unchanged. Three days of FR, particularly at 50% FR, were sufficient to lower mitochondria energetic metabolism in both mitochondrial populations. Our study highlights an early step in mitochondrial adaptation to FR and the influence of the severity of restriction on this adaptation. This step may be involved in an aging-retardation process.     

The study of energy metabolism in denervated skeletal muscle with 31P-NMR

1. The denervated frog sartorius muscle showed a decrease in the energy store more than that in the control. 2. In the caffeine contractures, both the denervated and the innervated muscles showed similar sequential changes in the relative concentration of phosphocreatine (PCr) to beta-adenosine triphosphate (beta-ATP) and inorganic phosphate (Pi) to beta-ATP. Instead, the intracellular pH value of the denervated muscle was lower than that of the control. 3. It is suggested that phosphate metabolism of the denervated muscle during contracture shows little difference from that of the control, nevertheless, the buffering capacity is decreased in the early stage of atrophy.     

In vivo modular control analysis of energy metabolism in contracting skeletal muscle

We used (31)P MRS (magnetic resonance spectroscopy) measurements of energetic intermediates [ATP, P(i) and PCr (phosphocreatine)] in combination with the analytical tools of metabolic control analysis to study in vivo energy metabolism in the contracting skeletal muscle of anaesthetized rats over a broad range of workload. According to our recent MoCA (modular control analysis) used to describe regulatory mechanisms in beating heart, we defined the energetic system of muscle contraction as two modules (PCr-Producer and PCr-Consumer) connected by the energetic intermediates. Hypoxia and electrical stimulation were used in this in vivo study as reasonably selective modulations of Producer and Consumer respectively. As quantified by elasticity coefficients, the sensitivities of each module to PCr determine the control of steady-state contractile activity and metabolite concentrations. The magnitude of the elasticity of the producer was high (4.3+/-0.6) at low workloads and decreased 5-fold (to 0.9+/-0.2) at high workloads. By contrast, the elasticity of the consumer remained low (0.5-1.2) over the range of metabolic rates studied. The control exerted by each module over contraction was calculated from these elasticities. The control of contraction was found on the consumer at low workloads and then swung to the producer, due to the workload-dependent decrease in the elasticity of producer. The workload-dependent elasticity and control pattern of energy production in muscle is a major difference from heart. Since module rate and elasticity depend on the concentrations of substrates and products, the absence of homoeostasis of the energetic intermediates in muscle, by contrast with heart, is probably the origin of the workload-dependent elasticity of the producer module.