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PubMed1976

Factors influencing rumen microbial growth rates and yields: effects of urea and amino acids over time.

Washed cell suspensions of mixed rumen bacteria were used to evaluate effects of 100% urea-nitrogen and 75% urea-nitrogen plus 25% amino acid-nitrogen in growth media upon microbial growth rate and yield, specific rate of glucose consumption, and incorporation of glucose into mixed cells, carbon dioxide, and end products. Rumen microbial dry matter, nitrogen, ribonucleic acid, deoxyribonucleic acid, glucose disappearance, and production of volatile fatty acids were considerably higher in medium containing urea plus amino acids as compared with urea only. Specific growth rates of microbes were .104 and .203 and mean doubling times were 6.7 and 3.4 h in the urea and urea plus amino acid growth media. Microbial growth in mg per 100 mg glucose used, per mole glucose and per mole adenosine triphosphate (ATP), and specific rate of glucose consumption in mmol per mg cells-h were 19.3, 34.7, 15.4, and .016 with urea, and 24.4, 44.2, 20.6, and .014 with urea plus amino acids. Percentages of catabolized glucose incorporated into microbial cells, carbon dioxide, and end products did not differ between treatments and averaged 19.5, 7.8, and 64.4%.

Adenosine Triphosphate↗
PubMed2005

Mitochondrial energy metabolism is markedly impaired by D-2-hydroxyglutaric acid in rat tissues.

Tissue accumulation of high amounts of D-2-hydroxyglutaric acid (DGA) and l-2-hydroxyglutaric acid (LGA) is the biochemical hallmark of the inherited neurometabolic disorders D-2-hydroxyglutaric aciduria (DHGA) and l-2-hydroxyglutaric aciduria (LHGA), respectively. Patients affected by DHGA predominantly present neurological and cardiomuscular symptoms, while those with LHGA have mainly severe neurological symptoms. Lactic aciduria and/or lactic acidemia may also occur in both disorders, suggesting mitochondrial dysfunction. We have previously reported that cytochrome c oxidase (COX) activity is severely inhibited by DGA in rat cerebral cortex and human skeletal muscle. In the present study, we initially evaluated the role of DGA and LGA on the mitochondrial respiratory chain complex activities, as well as CO2 on production in cardiac and skeletal muscle from 30-day-old Wistar rats. DGA significantly inhibited COX and ATP synthase (F0F1-ATP synthase) activities, in contrast to the other activities of the respiratory chain enzymes which were not affected by DGA in both muscular tissues. In addition, CO2 production was also markedly reduced by DGA in rat skeletal and cardiac muscles. On the other hand, LGA did not interfere with any of the respiratory chain complex activities studied, neither with CO2 generation. We also measured mitochondrial respiratory parameters in rat brain mitochondrial preparations in the presence of DGA and LGA. Both metabolites significantly lowered the respiratory control ratio in the presence of glutamate/malate and succinate. Since the metabolites stimulated oxygen consumption in state IV and compromised ATP formation, it can be presumed that these organic acids might act as endogenous uncouplers of mitochondria respiration. Moreover, COX activity linked to TMPD-ascorbate was significantly reduced by DGA in the brain mitochondrial enriched fractions. Finally, DGA and LGA reduced cell viability of rat cerebral cortex slices, as determined by the MTT assay. In case our in vitro data also occur in vivo, it may be presumed that impairment of energy metabolism may contribute to the understanding of the clinical features mainly of patients affected by DHGA.

Animals↗
PubMed2000

Mitochondrial defects by intracellular calcium overload versus endothelial cold ischemia/reperfusion injury.

Questions as to the critical stress factor and primary targets of cold ischemia/reperfusion (CIR) injury were addressed by comparing mitochondrial defects caused by (1) CIR injury and (2) intracellular Ca2+ overload. CIR was simulated in transformed human umbilical vein endothelial cell cultures (tEC) by 8 h cold anoxia in University of Wisconsin solution and reoxygenation at 37 degrees C. Intracellular Ca2+ concentrations were changed by permeabilization of suspended cells with digitonin in culture medium (RPMI, 0.4 mM Ca2+). Binding of free Ca2+ by ethylene glycol-bis(beta-aminoethylether)-N,N,N',N'-tetraacetic acid in RPMI or mitochondrial incubation medium served as controls. Extracellular Ca2+ protected the cell membrane against permeabilization. Mitochondrial functions were determined before and after permeabilization of the cell membrane. After CIR, mitochondrial respiratory capacity declined, but oxygen consumption remained coupled to adenosine triphosphate (ATP) production. In contrast, Ca2+ overload caused uncoupling of mitochondrial respiration. High intracellular Ca2+ overload, therefore, does not reproduce cold ischemia/reperfusion injury in endothelial cells.

Adenosine↗
PubMed1975

Energy metabolism of experimental wounds at various oxygen environments.

Energy metabolism of healing tissue was studied in experimental wounds of rats chronically breathing 11% O2, air or 55% O2. Increasing oxygen supply elevated both PO2 and PCO2 in the wound tissue. At the early phases of healing hypoxic wounds contained less DNA than normoxic or hyperoxic tissues. In hypoxia the accumulation of wound collagen was clearly retarded. Furthermore, tissue taken from wounds healing in hypoxic environments and tested ex vivo in air showed decreased capacity for glucose utilization, lactate production and oxygen consumption. Concentrations of AMP, ADP and ATP in repair tissue increased as healing progressed. The more oxygen available the higher the amounts of ADP and ATP. The AMP content was not affected by changes in local oxygen tension. These results support the earlier concept that the supply of oxygen in healing tissue may be rate-limitimg. Reduction of available oxygen either by systemic hypoxia or by increased diffusion distance impedes healing.

Animals↗
PubMed1986

Relation of actin fibrils to energy metabolism of endothelial cells.

The physiological significance of the association of glycolytic enzymes with actin fibrils was investigated in cell culture. Cytochalasin D (CD) was used to induce the known actin-based sequence of events in a culture of an endothelial-cell line (XTH-2) derived from hearts from tadpoles of Xenopus laevis. 1 min following addition of CD, ruptures in the cortical fibrillar meshwork and in stress fibres are seen. At the same time the cellular ATP level decreases by ca. 25%. This and the following reactions resulting in a kind of arborization depend on a continuous supply with metabolic energy. As shown by measurements of oxygen consumption, cells with intact energy metabolism provide the ATP needed from glycolysis; ATP produced by oxidative phosphorylation is not utilized as long as lactate dehydrogenase (LDH) reoxidizes NADH2. After inhibition of LDH, respiration in XTH-2 cells doubles. CD treatment induces a transient increase in oxygen consumption, indicating an increased energy supply by respiration. From these results we conclude: The energy needed by the actomyosin system is - under normal metabolic conditions -supplied from ATP phosphorylated in glycolysis. The processes of energy metabolism seem to be highly compartmentalized; ATP is not a parameter that is kept constant in time intervals of minutes up to one hour.

Actins↗
PubMed1995

A novel technique for preparing improved buffy coat platelet concentrates.

We evaluated in vitro platelet function of platelet concentrates stored at 22 C for 5 days prepared either by the conventional pelleting procedure or platelet concentrates prepared from buffy coats by utilizing a novel bucket designed to support a suspended bag. For platelet concentrates from buffy coat, whole blood was centrifuged at 3,000 x g for 13 min, with all but 30cc of the cell poor plasma transferred to a satellite bag, followed by a second centrifugation at 170 x g for 5 min utilizing our novel centrifugation device. For pelleted platelets, whole blood was centrifuged at 2,000 x g for 3 min, platelet rich plasma removed, centrifuged, and the pellet resuspended in plasma. Leukocyte contamination in buffy coat platelet concentrates was reduced by 95% (p < 0.001) in comparison to pelleted platelets. Further, platelets from buffy coat platelet concentrates demonstrated significantly enhanced ADP-induced aggregation, increased recovery from hypotonic shock, higher morphology scores, and reduced GMP-140 expression in comparison to pelleted preparations. No differences in O2 consumption, CO2 production, pH and total ATP were observed between the two types of preparations at day 5 of storage. Our results indicate that platelet concentrates from buffy coat, prepared by a suspended storage bag centrifugation technique, are superior with respect to in vitro platelet function when compared to pelleted platelets.

Adenosine Diphosphate↗
PubMed1983

Insulin release : the fuel concept.

Insulin release evoked by nutrient secretagogues invariably coincides with an increase in the catabolism of exogenous and/or endogenous nutrients in pancreatic islet cells, resulting in an increased generation rate of reducing equivalents and ATP, and an increase in O2 consumption. This situation was documented in response to a number of carbohydrates, including the anomers of D-glucose and D-mannose, various 2-keto acids, including pyruvate and its poorly oxidized phenylated analog 3-phenylpyruvate, and selected amino acids, including the nonmetabolized analog of L-leucine 2-aminobicyclo[2, 2, 1]heptane-2-carboxylic acid. These convergent observations indicate that the B-cell should be considered as a fuel-sensor organ. Further progress in this field requires both the identification of those circulating nutrients used by the B-cell whether in the resting or stimulated state, and a better understanding of the coupling between the generation of second messengers (H+, NAD(P)H, ATP) and more distal events in the secretory sequence, such as the remodelling of ionic fluxes across the plasma membrane and within the pancreatic B-cell.

Amino Acids↗
PubMedDate not supplied

[Quantitative model of human erythrocyte glycolysis. Relationship between erythrocyte energy metabolism and Na+, K+-ATPase activity].

The influence of the changes of the transport Na+, K+-ATPase activity on the human red blood cell (RBC) glycolysis was studied. Two different types of energetic regulation were found out under the strophanthin inhibition. A decrease of ATP comsumption results in a drop of glucose consumption and lactate production in the first type regulation of RBC. In the second type regulation of RBC the glucose consumption does not change after ATPase inhibition and the excess metabolic flux is directed through the hexosemonophosphate shunt. In both cases the resulting decrease of ATP production leads to the ATP level stabilization. An increase of potassium ions concentration in the external medium does not influence the RBC glycolysis. The valinomycin added increases the glucose consumption and the lactate production. The cell volume decreases under the effect of valinomycin.

Erythrocytes↗
PubMed1986

Effects of various inhibitors of oxidative phosphorylation on energy metabolism, macromolecular synthesis and cyclic AMP production in isolated rat thymocytes. A regulating role for the cellular energy state in macromolecular synthesis and cyclic AMP production.

Inhibitors of oxidative phosphorylation such as several triorganotin compounds, oligomycin, 2,4-dinitrophenol and carbonylcyanide p-trifluoromethoxyphenylhydrazone suppress energy metabolism of isolated rat thymocytes as indicated by a reduction of ATP levels, an increase in glucose consumption and by a marked accumulation of lactate. Also these compounds effectively inhibit the incorporation of DNA, RNA and protein precursors into acid-precipitable material of thymocytes. Moreover, the prostaglandin E1-induced elevation of cAMP is markedly reduced by these inhibitors. A correlation is observed between the effects on energy metabolism, macromolecular synthesis and cAMP production, since from a series of trialkyltin chlorides, tri-n-propyltin, tri-n-butyltin and tri-n-hexyltin are very effective inhibitors of these functions, while trimethyltin and tri-n-octyltin affect neither of them; other inhibitors of oxidative phosphorylation, each of them with quite different mechanisms of action, also inhibit macromolecular synthesis and cAMP production. The finding that a rise in intracellular ATP concentrations leads to a reversion of the tri-n-butyltin-induced inhibition of cAMP production and uridine incorporation, indicates a regulating role for the cellular energy state in these aspects of cellular function.

2,4-Dinitrophenol↗
PubMed2000

Hibernation in noncontracting mammalian cardiomyocytes.

BACKGROUND: -The aim of the present study was to establish whether isolated neonatal mammalian cardiomyocytes were capable of downregulating energy-using processes other than contraction while maintaining metabolic stability when oxygen availability was reduced. METHODS AND RESULTS: Metabolic response of cardiomyocytes was investigated under moderate (5 to 6 micromol/L) and severe (2 to 3 micromol/L) forms of hypoxia. Cells exposed to oxygen concentrations of 5 to 6 micromol/L exhibited rates of oxygen consumption, which were decreased to 64% of normoxic rates. Rates of cellular energy usage were decreased because this reduced rate of oxygen consumption was not associated with either decreased intracellular ATP and phosphocreatine concentrations or a compensatory switch to glycolysis. When cells were exposed to oxygen concentrations of 2 to 3 micromol/L, rates of oxygen consumption decreased to 9% of normoxic rates. This decreased rate of oxygen consumption was associated with energetic stress, because a significant switch to glycolysis occurred and intracellular phosphocreatine concentrations were decreased by 40%. Rates of cellular energy usage were further decreased as indicated by stable intracellular ATP concentrations. CONCLUSIONS: -Our results suggest that isolated cardiomyocytes are capable of downregulating energy-consuming processes other than contraction when oxygen supply is decreased. Regions of myocardial tissue are also capable of downregulating metabolic activity during ischemia by shutting down contractile activity (myocardial hibernation). We suggest that metabolic downregulation associated with myocardial hibernation may not be exclusively due to reduced rates of contractile activity. Other energy-using processes (eg, protein synthesis, mRNA synthesis, ion channel activity, and proton leak) may also be shut down.

Adenosine Triphosphate↗
PubMed1975

Respiratory control in isolated perfused rat heart. Role of the equilibrium relations between the mitochondrial electron carriers and the adenylate system.

The effects of KCl-induced cardiac arrest on the redox state of the fluorescent flavoproteins and nicotinamide nucleotides and on that of cytochromes c and a were studied by surface fluorometric and reflectance spectrophotometric methods. These changes were compared with measurements of the concentrations of the adenylate system, creatine phosphate, some intermediates of the tricarboxylic acid cycle and reactants of the glutamate dehydrogenase system. KCl-induced cardiac arrest caused reduction of the fluorescent flavoproteins and nicotinamide nucleotides, oxidation of cytochromes c and a, inhibition of oxygen consumption and an increase in the ATP/(ADP X Pi) ratio. The increase in the latter was due mainly to a decrease in the concentration of Pi and an equivalent increase in creatine phosphate. The cytochromes c and a were maintained at equal redox potential and changed in parallel. When the redox state of the mitochondrial NAD couple was calculated from the glutamate dehydrogenase equilibrium, the free energy change (deltaG) corresponding to the potential difference between the NAD couple and cytochrome c was 115.8 kj/mol in the beating heart and 122.2 kj/mol in the arrested heart. The deltaG values of ATP hydrolysis calculated from the concentrations of ATP, Pi and ADP, corrected for bound ADP, were 111.1 kj/2 mol and 115.4 kj/2 mol in the beating and arrested heart respectively. The accumulation of citrate and the direction of the redox changes in the respiratory carriers indicate that the tricarboxylic acid cycle flux is controlled by the respiratory chain. The data also show a near equilibrium between the electron carriers and the adenylate system and suggest that the equilibrium hypothesis of mitochondrial respiratory control is applicable to intact myocardial tissue.

Adenosine Diphosphate↗
PubMed1990

[Respiration and motility of epididymal sperm from slaughtered bulls].

The simple method of retrograde flushing of spermatozoa from the epididymal cauda of slaughter bulls yielded an average of 2 x 10(9) spermatozoa from one cauda. The plasma membrane was intact in something between 80 and 85 percent of all epididymal spermatozoa. Respiratory rates and motility were clearly below values typical of ejaculated spermatozoa, though sizeable differences were found to exist for both parameters between individual preparations. The endogenous substrates available proved to be sufficient for such low metabolic activity, whereas higher energy turnover required the presence of exogenous substrates, such as lactate, pyruvate or fructose. Respiratory capacity, determined by uncoupling of oxidative phosphorylation in the presence of lactate, was comparable to that of ejaculated bull spermatozoa. The above findings are likely to suggest that the lower respiratory rates of epididymal spermatozoa were attributable to the fact that their consumption for motility of adenosine triphosphoric acid (ATP) was lower than that of ejaculated spermatozoa.

Abattoirs↗
PubMed1989

What causes silent myocardial ischemia?

Myocardial ischemia is an imbalance between consumption and production of adenosine triphosphate (ATP) that leads to ATP depletion and a cascade of biochemical events. Why some patients have pain during these events while others do not is unclear, but some studies indicate that a combination of pain threshold and magnitude of ischemia may be at work. Ischemia can occur during vigorous or daily activities or at rest and can be influenced by mental stress. It is most common in the morning hours, possibly because heart rate, blood pressure, and contractility rise rapidly in the morning and factors that increase coronary vessel tone and reduce blood flow also increase during these hours. The specific pathophysiology of an ischemic episode depends on whether the patient also has chronic stable coronary disease, variant angina, or unstable angina, not on whether the episode is silent or symptomatic.

Angina Pectoris↗
PubMed2005

3-Hydroxyglutaric acid moderately impairs energy metabolism in brain of young rats.

3-Hydroxyglutaric acid (3HGA) accumulates in the inherited neurometabolic disorder known as glutaryl-CoA dehydrogenase deficiency. The disease is clinically characterized by severe neurological symptoms, frontotemporal atrophy and striatum degeneration. Because of the pathophysiology of the brain damage in glutaryl-CoA dehydrogenase deficiency is not completed clear, we investigated the in vitro effect of 3HGA (0.01-5.0mM) on critical enzyme activities of energy metabolism, including the respiratory chain complexes I-V, creatine kinase isoforms and Na(+),K(+)-ATPase in cerebral cortex and striatum from 30-day-old rats. Complex II activity was also studied in rat C6-glioma cells exposed to 3HGA. The effect of 3HGA was further investigated on the rate of oxygen consumption in mitochondria from rat cerebrum. We observed that 1.0mM 3HGA significantly inhibited complex II in cerebral cortex and C6 cells but not the other activities of the respiratory chain complexes. Creatine kinase isoforms and Na(+),K(+)-ATPase were also not affected by the acid. Furthermore, no inhibition of complex II activity occurred when mitochondrial preparations from cerebral cortex or striatum homogenates were used. In addition, 3HGA significantly lowered the respiratory control ratio in the presence of glutamate/malate and succinate under stressful conditions or when mitochondria were permeabilized with digitonin. Since 3HGA stimulated oxygen consumption in state IV and compromised ATP formation, it can be presumed that this organic acid might act as an endogenous uncoupler of mitochondria respiration. Finally, we observed that 3HGA changed C6 cell morphology from a round flat to a spindle-differentiated shape, but did not alter cell viability neither induced apoptosis. The data provide evidence that 3HGA provokes a moderate impairment of brain energy metabolism and do not support the view that 3HGA-induced energy failure would solely explain the characteristic brain degeneration observed in glutaryl-CoA dehydrogenase deficiency patients.

Animals↗
PubMed1966

The effects of adenine nucleotides on carbohydrate metabolism in pigeon-liver homogenates.

1. The effects of added AMP on carbohydrate metabolism were investigated in pigeon-liver homogenates, which can degrade glucose and synthesize it from lactate. Suitable experimental conditions were established for studying such effects, including the addition of P(i) (20mm) to stabilize adenine nucleotides and supplementation with NAD(+) (0.5mm). 2. Lactate increased the rate of oxygen consumption and kept the concentration of ATP high and that of AMP relatively low. 3. Added AMP (1.25-5mm) raised the net rate of carbohydrate removal and inhibited the net formation of glucose from lactate, as well as the incorporation of lactate into glucose. These effects were accompanied by a fall in the concentrations of hexose 6-phosphates and a rise in those of fructose diphosphate and triose phosphates. When the activity of glyceraldehyde 3-phosphate dehydrogenase was limited experimentally by a low concentration of NAD(+) or when it was blocked by iodoacetate, the accumulations of fructose diphosphate and triose phosphates were large and accounted for most of the carbohydrate degraded in the presence of AMP. 4. AMP also inhibited the conversion of pyruvate into phosphoenolpyruvate. Data on the concentrations of pyruvate, phosphoenolpyruvate and intermediates of the tricarboxylic acid cycle, as well as on isotope distribution, suggest that the effect was due to inhibition of phosphoenolpyruvate carboxykinase. 5. The results indicate that in the homogenates phosphofructokinase and fructose diphosphatase, controlled in their activity by adenine nucleotides and other cell constituents, are enzymes which regulate the direction of carbohydrate metabolism (degradation or synthesis) in the liver. 6. It is suggested that active transport of adenine nucleotides, citrate, Mg(2+), Ca(2+), P(i) and other cell constituents may play a role in regulating the activity of enzymes which are affected by these substances. 7. A procedure is described for generating alkali in a closed manometer vessel, by mixing mercuric oxide and a solution of sodium iodide, for use in a method for measuring the oxygen consumption at physiological bicarbonate concentrations.

Adenine Nucleotides↗
PubMed1979

Respiration in Relation to Adenosine Triphosphate Content during Desiccation and Rehydration of a Desiccation-tolerant and a Desiccation-intolerant Moss.

O(2) consumption by the desiccation-tolerant moss Tortula ruralis and the desiccation-intolerant Cratoneuron filicinum increased markedly during the latter stages of desiccation. ATP content of the mosses during desiccation was not correlated with O(2) consumption, but was influenced by the rate at which the mosses lost water. The more rapid the water loss, the more ATP that was present in the dry mosses. The pattern of O(2) consumption on rehydration also was influenced by the previous rate of desiccation. After rapid desiccation of T. ruralis O(2) consumption upon rehydration was considerably elevated, and for up to 24 hours. After very slow desiccation the elevation was small and brief. Normal O(2) consumption did not occur in C. filicinum after rapid desiccation, but did so within a few hours of rehydration after slower speeds of drying. ATP levels in T. ruralis returned to normal within 5 to 10 minutes of rehydration. In C. filicinum, increases in ATP were closely correlated with O(2) consumption. These observations are considered to be related to differential damage caused to mitochondria and to cellular integrity by different speeds of water loss. The desiccation-tolerant moss appears to be able to repair the severe damage imposed by rapid desiccation whereas the desiccation-intolerant moss cannot.

Journal Article↗
PubMed1998

Impairment of the mitochondrial electron chain transport prevents NF-kappa B activation by hydrogen peroxide.

A large body of work has been devoted to mechanisms leading to the activation of the transcription factor NF-kappa B in various cell types. Several studies have indicated that NF-kappa B activation by numerous stimuli depends on the intracellular generation of reactive oxygen species (ROS). In this report, we first demonstrated that inhibition of the electron transport chain by either rotenone or antimycine A gave rise to dose-dependent inhibition of NF-kappa B translocation induced by 150 microM of hydrogen peroxide (H2O2). Conversely, the impairment of the mitochondrial respiratory chain did not affect T lymphocyte treatment by TNF-alpha (tumor necrosis factor alpha) or pre-B lymphocyte treatment with LPS (lipopolysaccharide). We also showed that oligomycine which inhibits ATP synthase and FCCP, which uncouples respiration also led to dose-dependent inhibition of NF-kappa B activation by H2O2. All these inhibitors were also shown to inhibit mitochondrial respiration in lymphocytes assessed by oxygen consumption. Although only a transient drop in ATP concentration was observed when lymphocytes were treated by H2O2, this effect was remarkably reinforced in the presence of oligomycine demonstrating the crucial role of ATP in the signal transduction pathway induced by H2O2.

Adenosine Triphosphate↗
PubMed1997

Hepatic oxygen supply, energy charge, and histological findings in dogs with portal vein arterialization.

Hepatic oxygen supply, energy charge (EC), and histology were examined comparatively in dogs with portal vein anastomosis (PVA group), and PA in addition to PVA (PA group). The PVA group showed a lower level of hepatic oxygen supply than those of the PA group throughout the experimental period, and also showed decreases of adenosine triphosphate (ATP) and EC level after blood perfusion. In contrast, the oxygen supply and consumption were stable in the PA group. A temporary fall of ATP level was followed by recovery to the preperfusion level in the PA group. Histological examination indicated the collapse of hepatic cords with granular and vacuolar degeneration in only the PVA group. These findings suggested that PA, when supplemented to PVA, is an available technique for preventing hepatic failure caused by ischemic conditions.

Adenosine Triphosphatases↗