Mitochondrial ATP Production Rate Research Articles

Effect of short-term training on mitochondrial ATP production rate in human skeletal muscle.

Seven untrained volunteers [3 men, 4 women, 20.1 +/- 2.0 (SD) yr, 66. 0 +/- 11.0 kg, 171 +/- 13 cm] participated in a 10-day cycle exercise training program. Resting muscle samples were obtained from vastus lateralis before and after 5 and 10 days of training. Mitochondrial ATP production rate (MAPR) was assayed in isolated mitochondria by using a bioluminescence technique and referenced to the activity of glutamate dehydrogenase in the muscle sample. MAPR increased 136 and 161% after 10 days of training for the mitochondrial substrate combinations pyruvate + palmitoyl-L-carnitine + alpha-ketoglutarate + malate and palmitoyl-L-carnitine + malate, respectively. Total muscle glutamate dehydrogenase and citrate synthase activity increased 53 and 16%, respectively, after 5 days but did not significantly increase further after 10 days. The results from the present study indicate that MAPR, measured by using the substrate combinations pyruvate + palmitoyl-L-carnitine + alpha-ketoglutarate + malate and palmitoyl-L-carnitine + malate, can rapidly increase in response to endurance training.

Reduced cytosolic acidification during exercise suggests defective glycolytic activity in skeletal muscle of patients with Becker muscular dystrophy. An in vivo 31P magnetic resonance spectroscopy study.

Becker muscular dystrophy is an X-linked disorder due to mutations in the dystrophin gene, resulting in reduced size and/or content of dystrophin. The functional role of this subsarcolemma protein and the biochemical mechanisms leading to muscle necrosis in Becker muscular dystrophy are still unknown. In particular, the role of a bioenergetic deficit is still controversial. In this study, we used 31p magnetic resonance spectroscopy (31p-MRS) to investigate skeletal muscle mitochondrial and glycolytic ATP production in vivo in 14 Becker muscular dystrophy patients. Skeletal muscle glycogenolytic ATP production, measured during the first minute of exercise, was similar in patients and controls. On the other hand, during later phases of exercise, skeletal muscle in Becker muscular dystrophy patients was less acidic than in controls, the cytosolic pH at the end of exercise being significantly higher in Becker muscular dystrophy patients. The rate of proton efflux from muscle fibres of Becker muscular dystrophy patients was similar to that of controls, pointing to a deficit in glycolytic lactate production as a cause of higher end-exercise cytosolic pH in patients. The maximum rate of mitochondrial ATP production was similar in muscle of Becker muscular dystrophy patients and controls. The results of this in vivo 31P-MRS study are consistent with reduced glucose availability in dystrophin-deficient muscles.

Human skeletal muscle pyruvate dehydrogenase kinase activity increases after a low-carbohydrate diet.

To characterize human skeletal muscle enzymatic adaptation to a low-carbohydrate, high-fat, and high-protein diet (LCD), subjects consumed a eucaloric diet consisting of 5% of the total energy intake from carbohydrate, 63% from fat, and 33% from protein for 6 days compared with their normal diet (52% carbohydrate, 33% fat, and 14% protein). Biopsies were taken from the vastus lateralis before and after 3 and 6 days on a LCD. Intact mitochondria were extracted from fresh muscle and analyzed for pyruvate dehydrogenase (PDH) kinase, total PDH, and carnitine palmitoyltransferase I activities and mitochondrial ATP production rate (using carbohydrate and fat substrates). beta-Hydroxyacyl CoA dehydrogenase, active PDH (PDHa), and citrate synthase activities were also measured on whole muscle homogenates. PDH kinase (PDHK) was calculated as the absolute value of the apparent first-order rate constant of the inactivation of PDH in the presence of 0.3 mM Mg2+-ATP. PDHK increased dramatically from 0.10 +/- 0.02 min-1 to 0.35 +/- 0.09 min-1 at 3 days and 0.49 +/- 0. 06 min-1 after 6 days. Resting PDHa activity decreased from 0.63 +/- 0.17 to 0.17 +/- 0.04 mmol. min-1. kg-1 after 6 days on the diet, whereas total PDH activity did not change. Activities for all other enzymes were unaltered by the LCD. In summary, severe deficiency of dietary carbohydrate combined with a twofold increase in dietary fat and protein caused a rapid three- to fivefold increase in PDHK activity in human skeletal muscle. The increased PDHK activity downregulated the amount of PDH in its active form at rest and decreased carbohydrate metabolism. However, an increase in the activities of enzymes involved in fatty acid oxidation did not occur.

Normal in vivo skeletal muscle oxidative metabolism in sporadic inclusion body myositis assessed by 31P-magnetic resonance spectroscopy.

Sporadic inclusion body myositis (s-IBM) is a chronic inflammatory myopathy of unknown pathogenesis. The common findings of ragged red fibres, cytochrome c oxidase-negative fibres and multiple mitochondrial DNA deletions in the muscle of patients with s-IBM have suggested that a deficit of energy metabolism may be of pathogenic relevance. To test this hypothesis we used 31P magnetic resonance spectroscopy to assess in vivo skeletal muscle mitochondrial function in the calf muscles of 12 patients with definite s-IBM. Eleven patients showed multiple mitochondrial DNA deletions in skeletal muscle and 67% showed ragged red fibres and/or cytochrome c oxidase-negative fibres. T1-weighted MR images showed increased signal intensity in the calf muscle of all patients except one. The involvement of calf muscle was confirmed by 31P magnetic resonance spectroscopy of resting muscle, which disclosed abnormalities in metabolite ratios in all patients. However, muscle oxidative metabolism assessed during recovery from exercise was normal in patients with s-IBM, as maximum rates of mitochondrial ATP production and post-exercise ADP recovery rates were within the normal range in all cases. We conclude that muscle mitochondrial abnormalities are a secondary process and unlikely to play a significant role in the pathogenesis of s-IBM.

In vivo skeletal muscle mitochondrial function in Leber's hereditary optic neuropathy assessed by 31P magnetic resonance spectroscopy.

We used 31P magnetic resonance spectroscopy (31P-MRS) to assess in vivo skeletal muscle mitochondrial function in 10 Leber's hereditary optic neuropathy patients/carriers with a mitochondrial DNA (mtDNA) mutation at one of three nucleotide positions, 11,778, 14,484, and 3,460. We studied one affected patient for each mutation and two unaffected carriers with the 11,778 or 3,460 mutation and three carriers with 14,484. All subjects were homoplasmic except the two 3,460 carriers, who showed 80% and 15% of mutated mtDNA. 31P-MRS at rest disclosed some abnormalities in all subjects. In particular, the phosphorylation potential was below the normal range in all cases. During recovery from exercise, the maximum rate of mitochondrial ATP production (Vmax) was reduced to 27% of normal in the 11,778 mutation and to 53% in the 14,484 mutation patient/carrier groups. Mitochondrial Vmax was within the normal range in all subjects with the 3,460 mutation but correlated inversely with the percentage of mutated mtDNA. This in vivo study shows that the 11,778 mutation causes a mitochondrial impairment more severe than the 14,484 and that the 3,460 mutation results in only a mild depression of muscle mitochondrial function.

MITOCHONDRIAL ATP PRODUCTION RATE (MAPR) IN HUMAN SKELETAL MUSCLE AFTER SHORT-TERM TRAINING 1552

The present study employed a novel technique that directly measures mitochondrial ATP production rate (MAPR) in muscle samples to examine the effect of 10 days of exercise training on mitochondrial oxidative capacity. Healthy, untrained volunteers (3 males, 4 females, 20 ± 2 yrs, 66± 11 kg, mean ± SD) participated in a 10 day training program which consisted of one training session per day, either 60 min of cycle exercise at 75% ˙VO2 peak, or 6 × 5 min exercise bouts at 95%˙VO2 peak, alternating over the 10 days. Muscle samples were obtained before training, and after 5 and 10 days of training. MAPR was determined in isolated mitochondria using a bioluminescence technique, referenced to the activity of glutamate dehydrogenase (GDH). MAPR for the mitochondrial substrate combinations pyruvate + palmitoyl-L-camitine +α-ketoglutarate + malate (PPKM) and palmitoyl-L-camitine + malate (PM) increased (*P<0.05) 41%* and 19% after 5 days and 144%* and 133%* after 10 days. MAPR for both substrates increased (P<0.05) from day 5 to day 10. Total GDH activity increased 53%* after 5 days, but there was no further increase after 10 days.˙VO2 peak increased 9% after 10 days of training (2.85 ± 0.83 vs. 3.10 ± 0.89 l.min-1, P<0.05). The results from the present study indicate that MAPR measured using the substrates PPKM and PM is increased after 10 days of training. Furthermore, the absence of changes total GDH activity during the last 5 days of training suggests that the increase in MAPR with short-term training may be due to mechanisms in addition to greater mitochondrial mass.

Quantitative analysis of skeletal muscle bioenergetics and proton efflux in migraine and cluster headache

Phosphorus MR spectroscopy ( 31P-MRS) was used to quantify skeletal muscle bioenergetics and proton efflux in 63 patients with migraine (23 with migraine without aura, MwoA, 22 with migraine with aura, MwA, and 18 with prolonged aura or smoke, CM) and in 14 patients with cluster headache (CH), all in an attack-free period. At rest mitochondrial function was abnormal only in CM, as shown by a low phosphocreatine (PCr) concentration. At the end of a mixed glycolytic/aerobic exercised all three migraine groups showed a significantly smaller decrease of cytosolic pH compared to controls with a similar end-exercise PCr breakdown, while end-exercise pH was normal in cluster headache patients. The normal rate of proton efflux in all headache groups suggests that the reduced end-exercise acidification was due to a reduction of glycolytic flux in migraine patients. The maximum rate of mitochondrial ATP production ( Q max), calculated from the rate of post-exercise PCr recovery and the end-exercise [ADP], was low in cluster headache patients as well as in migraine patients except MwoA. In migraine the degree of the mitochondrial impairement, that apparently is associated with a reduced glycolytic flux, is related to the severity of the clinical phenotype. © 1997 Eseevier Science B.V. All rights reserved.

Prior heat stress enhances survival of renal epithelial cells after ATP depletion.

The 72-kDa heat stress protein (HSP-72) is an inducible cytoprotectant protein. Although transient renal ischemia in vivo induces HSP-72, it is not known whether prior heat stress protects renal epithelial cells from injury mediated by ATP depletion. To evaluate this hypothesis, opossum kidney (OK) cells were exposed to sodium cyanide and 2-deoxy-D-glucose in the absence of medium glucose, a maneuver that reduced cell ATP content to < 10% of the control value within 10 min and decreased cell survival. One day after 2 h of ATP depletion, OK cells previously exposed to heat stress (to induce accumulation of HSP-72) exhibited marked improvement in survival (a > 4-fold increase in total DNA), less uptake of vital dye, and less release of lactate dehydrogenase (LDH) than cells subjected to ATP depletion alone (23.0 +/- 1.6 vs. 34.1 +/- 1.2% of total LDH, respectively). Enhanced clonogenicity post-heat stress was completely prevented by cycloheximide and positively correlated with the steady-state content of HSP-72. In the recovery period after ATP depletion, cell ATP content, maximum mitochondrial ATP production rate, and total LDH activity were all significantly higher in cells with abundant HSP-72. Although the protective effects associated with heat stress are likely to be multifactoral, preserved cell metabolism and higher ATP content could enhance cellular repair processes after ATP depletion.

Locomotor muscle of high-performance fishes: What do comparisons of tunas with ectothermic sister taxa reveal?

Abstract Many studies of high-performance fishes focus on tunas because they are morphologically adapted for efficient swimming, they migrate long distances and accelerate rapidly, and they successfully exploit the epipelagic marine environment. Tunas have also received attention because, by conserving metabolically generated heat, they elevate the temperature of certain tissues above ambient water temperature; that is, they are endothermic. However, few studies have used phylogenetically based comparisons to determine what is unique about tunas and to elucidate evolutionary trends. This paper compares tunas with their closest relatives, ectothermic fishes of the family Scombridae, and examines characteristics of the myotomal musculature that are related to swimming performance. White muscle anaerobic capacity is significantly greater in tunas than in ectothermic scombrid fishes, indicating that energy generation for bursts may be greater in tunas. Because tunas have high capacities to buffer metabolic acids within the white muscle and blood, and to recover from exhaustive exercise rapidly, they may be able to burst repeatedly. The white muscle of tunas also has a higher aerobic capacity than that of ectothermic scombrids, which may indicate that at least some tuna white muscle fibers are recruited at sustainable speeds or it may be related to rapid lactate turnover or growth rates. In contrast, many intracellular characteristics of the red myotomal muscle do not differ significantly between tunas and ectothermic scombrid fishes. Many of these, including mitochondrial ATP production rate and muscle contraction speed, are temperature dependent and are higher at the higher in vivo red muscle temperatures of tunas. Thus, endothermy may result in elevated aerobic performance levels in tunas, and their high blood hemoglobin concentrations, muscle myoglobin contents, and red muscle capillary densities would ensure adequate oxygen delivery to the active, warm red muscle fibers. However, direct tests of this hypothesis are needed. Much more remains to be learned about tunas and ectothermic scombrids so that more complete phylogenetic analyses can be used to understand the adaptive strategies that have resulted in tunas' success as active epipelagic predators.

Mitochondrial ATP production rate in 55 to 73-year-old men: effect of endurance training.

The effect of 6-week endurance training on mitochondrial ATP production rate was investigated in 14 elderly men. Mean age, body weight and height were 63 +/- 6 yr, 75.6 +/- 9.2 kg and 174 +/- 4 cm, respectively. Subjects trained on a Monark cycle ergometer at 79 +/- 8% of their maximal heart rate for 1 h day-1, 4 days week-1. Muscle samples were obtained at rest, before and after endurance training, by a needle biopsy technique and used for determination of mitochondrial ATP production rate in isolated mitochondria and enzyme assays. Endurance training resulted in a significant increase in maximal oxygen uptake (L min-1) (P < 0.01). Citrate synthase activity, a mitochondrial marker enzyme, and hexokinase activity increased significantly (both P < 0.01) in response to training while 3-hydroxyacyl-CoA dehydrogenase and carnitine palmitoyltransferase I activities remained statistically unchanged. A higher mitochondrial ATP production rate was observed after endurance training with the substrate combinations pyruvate+palmitoyl-L-carnitine+L-glutamate+malate (P < 0.01), L-glutamate (P < 0.001), pyruvate+malate (P < 0.05) and palmitoyl-L-carnitine+malate (P < 0.01). The largest increase was obtained with L-glutamate (170%). Significant correlations were observed between the percent increase in citrate synthase activity and those of mitochondrial ATP production rates. It was concluded that the increased mitochondrial ATP production rate of aged human skeletal muscle with training seems mainly to occur through an increased mitochondrial content, and in a way similar to those observed in young men.

Altered properties of mitochondrial ATP-synthase in patients with a T → G mutation in the ATPase 6 (subunit a) gene at position 8993 of mtDNA

A family is described with a T → G mutation at position 8993 of mtDNA. This mutation is located in the ATPase 6 gene of mtDNA which encodes subunit a of the ATP-synthase complex (F 1F 0-ATPase). Clinically, the patients showed severe infantile lactate acidosis and encephalomyopathy in a form that was different from the classical Leigh syndrome. In 3 affected boys, ranging in age from 3 months to 8 years, the mutation was found in 95–99% of the mtDNA population. The clinical symptoms correlated with the mtDNA heteroplasmy and in the healthy mother 50% of the mtDNA was mutated. The rate of mitochondrial ATP production by cultured skin fibroblasts containing 99% of mutated ml was about 2-fold lower than that in normal fibroblasts. Native electrophoresis of the mitochondrial enzyme complexes revealed instability of the F 1F 0-ATPase in all the tissues of the patient that were investigated (heart, muscle, kidney, liver). Only a small portion of the ATP-synthase complex was present in the complete, intact form (620 kDa). Incomplete forms of the enzyme were present as subcomplexes with approx. molecular weights of 460, 390 and 150 kDa, respectively, which differed in the content of F 1 and F 0 subunits. Immunochemical analysis of the subunits of the F 1F 0-ATPase further revealed a markedly decreased content of the F 0 subunit b in mitochondria from muscle and heart, and an increased content of the F 0 subunit c in muscle mitochondria respectively. These results indicate that in this family the T → G point mutation at position 8993 in the mitochondrial ATPase 6 gene is accompanied by structural instability and altered assembly of the enzyme complex, that are both most likely due to changes in the properties of subunit a of the membrane sector part of the ATP-synthase.

Adaptation of mitochondrial ATP production in human skeletal muscle to endurance training and detraining.

The adaptation of mitochondrial ATP production rate (MAPR) to training and detraining was evaluated in nine healthy men. Muscle samples (approximately 60 mg) were obtained before and after 6 wk of endurance training and after 3 wk of detraining. MAPR was measured in isolated mitochondria by a bioluminometric method. In addition, the activities of mitochondrial and glycolytic enzymes were determined in skeletal muscle. In response to training, MAPR increased by 70%, with a substrate combination of pyruvate + palmitoyl-L-carnitine + alpha-ketoglutarate + malate, by 50% with only pyruvate + malate, and by 92% with palmitoyl-L-carnitine + malate. With detraining MAPR decreased by 12-28% from the posttraining rate (although not significantly for all substrates). No differences were found when MAPR was related to the protein content in the mitochondrial fraction. The largest increase in mitochondrial enzyme activities induced by training was observed for cytochrome-c oxidase (78%), whereas succinate cytochrome c reductase showed only an 18% increase. The activity of citrate synthase increased by 40% and of glutamate dehydrogenase by 45%. Corresponding changes in maximal O2 uptake were a 9.6% increase by training and a 6.0% reversion after detraining. In conclusion, both MAPR and mitochondrial enzyme activities are shown to increase with endurance training and to decrease with detraining.

ATP production in isolated muscle mitochondria from haemodialysis patients: effects of correction of anaemia with erythropoietin.

1. The ATP production rate in isolated skeletal muscle mitochondria was measured with a bioluminescence method, before and during erythropoietin treatment, in 21 anaemic haemodialysis patients. In addition, the concentrations of ATP, phosphocreatine and total creatine and the ratio of alkali-soluble protein to DNA were determined in skeletal muscle. Maximal oxygen uptake and maximal exercise capacity were determined on a bicycle ergometer. 2. The results unexpectedly showed a 35% higher mitochondrial ATP production rate in the patients before erythropoietin treatment than in sedentary control subjects. On the other hand, mitochondrial density, as measured by the activity of the matrix enzyme glutamate dehydrogenase, was the same in the patients as in the sedentary control group. After 1 year on maintenance erythropoietin treatment, the ATP production rate per kg of muscle decreased in five out of seven patients and reached the same level as in the sedentary control subjects. The ratio between ATP production rate and glutamate dehydrogenase activity was on average 40% higher in the patients at the start and decreased towards the control level in six out of seven patients after 1 year on maintenance erythropoietin treatment. When related to the mitochondrial protein content, a significant reduction in the ATP production rate was observed. 3. The ratio of alkali-soluble protein to DNA in skeletal muscle and the concentrations of ATP, phosphocreatine and total creatine in skeletal muscle at rest were normal in the patients and did not change during the study. The maximal aerobic power improved by 25% after the correction of anaemia.(ABSTRACT TRUNCATED AT 250 WORDS)

ATP production rate in mitochondria isolated from microsamples of human muscle

Mitochondrial ATP production (MAPR) was determined using a bioluminescence method in mitochondrial preparations of human skeletal muscle. We obtained muscle samples from 21 healthy subjects using the percutaneous muscle biopsy technique. The subjects were grouped according to their degree of physical activity, i.e., from sedentary to highly active. The MAPR for each subject was related to the mitochondrial protein content and to the activity of glutamate dehydrogenase (GDH) estimated in muscle homogenates and in isolated mitochondria. With the use of GDH as a reference base, the MAPR could be expressed per muscle mass. MAPR was determined for different individual substrates and also for the combination of pyruvate, palmitoyl-carnitine, alpha-ketoglutarate, and malate (MAPRPPKM). The MAPR observed was higher for the combination of substrates than for any individual substrate tested. The relation between mitochondrial ATP production rate and GDH activity was the same for all subjects irrespective of physical activity status, but MAPRPPKM/kg muscle varied from 6.6 +/- 1.3 mmol.min-1.kg-1 in sedentary subjects to 11.0 +/- 2.2 in highly active subjects. The present method, which uses 50 mg of muscle tissue, enables mitochondrial function to be estimated in healthy subjects or patients with mitochondrial disorders.