glycogen and Myocardial-Ischemia

After exhaustive exercise, intravenous or oral glutamine promoted skeletal muscle glycogen storage. However, when glutamine was ingested with glucose polymer, whole-body carbohydrate storage was elevated, the most likely site being liver and not muscle, possibly due to increased glucosamine formation. The rate of tricarboxylic acid (TCA) cycle flux and hence oxidative metabolism may be limited by the availability of TCA intermediates. There is some evidence that intramuscular glutamate normally provides alpha-ketoglutarate to the mitochondrion. We hypothesized that glutamine might be a more efficient anaplerotic precursor than endogenous glutamate alone. Indeed, a greater expansion of the sum of muscle citrate, malate, fumarate and succinate concentrations was observed at the start of exercise (70% VO2(max)) after oral glutamine than when placebo or ornithine alpha-ketoglutarate was given. However, neither endurance time nor the extent of phosphocreatine depletion or lactate accumulation during the exercise was altered, suggesting either that TCA intermediates were not limiting for energy production or that the severity of exercise was insufficient for the limitation to be operational. We have also shown that in the perfused working rat heart, there is a substantial fall in intramuscular glutamine and alpha-ketoglutarate, especially after ischemia. Glutamine (but not glutamate, alpha-ketoglutarate or aspartate) was able to rescue the performance of the postischemic heart. This ability appears to be connected to the ability to sustain intracardiac ATP, phosphocreatine and glutathione.

Topics: Animals; Citric Acid Cycle; Clinical Trials as Topic; Exercise; Glucans; Glutamic Acid; Glutamine; Glutathione; Glycogen; Humans; Muscle Fatigue; Muscle, Skeletal; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Perfusion

Normal cardiac function requires a tight interaction between metabolism, contractile function and gene expression. The main perturbation challenging this equilibrium in vivo is ischemia, which alters energy flux through the control of key enzymes. The review highlights metabolic imprints and energetic aspects of programmed cell survival, programmed cell death, and of necrosis. When sustained and severe, ischemia leads to a total collapse of energy transfer, to the accumulation of metabolic endproducts, and to the development of myocardial necrosis. When moderate, ischemia results in a coordinated cellular response including enhanced anaerobic glucose metabolism, a modification of cardiac gene expression, and the development of specific mechanisms for programmed cell survival (preconditioning, stunning, hibernation). Repetitive stress results in a decrease of contractile function, a downregulation of gene expression and an impairment of energy transfer, which eventually cause the heart to fail. When the failing heart becomes energy-depleted, the programs of cell survival are no longer operational and programmed cell death ensues. To define the point of departure from programmed cell survival to cell death remains a major challenge.

Topics: Adenosine Triphosphate; Animals; Apoptosis; Cell Survival; Energy Metabolism; Gene Expression; Glucose; Glycogen; Humans; Myocardial Contraction; Myocardial Ischemia; Myocardium; Nitric Oxide

Topics: Animals; Glucose; Glycogen; Humans; Insulin; Myocardial Ischemia; Myocardium; Reperfusion Injury

Experimentally, enhanced glycolytic flux has been shown to confer many benefits to the ischemic heart, including maintenance of membrane activity, inhibition of contracture, reduced arrhythmias, and improved functional recovery. While at moderate low coronary flows, the benefits of glycolysis appear extensive, the controversy arises at very low flow rates, in the absence of flow; or when glycolytic substrate may be present in excess, such that high glucose concentrations with or without insulin overload the cell with deleterious metabolites. Under conditions of total global ischemia, glycogen is the only substrate for glycolytic flux. Glycogenolysis may only be protective until the accumulation of metabolites (lactate, H+, NADH, sugar phosphates and Pi ) outweighs the benefit of the ATP produced. The possible deleterious effects associated with increased glycolysis cannot be ignored, and may explain some of the controversial findings reported in the literature. However, an optimal balance between the rate of ATP production and rate of accumulation of metabolites (determined by the glycolytic flux rate and the rate of coronary washout), may ensure optimal recovery. In addition, the effects of glucose utilisation must be distinguished from those of glycogen, differences which may be explained by functional compartmentation within the cell.

Topics: Animals; Glucose; Glycogen; Glycolysis; Humans; Models, Biological; Myocardial Ischemia; Myocardium

The mammalian myocardium meets its high energy needs through the oxidation of a variety of substrates, chiefly fatty acids. This review examines the hypothesis that efficient energy transfer in the heart occurs through a series of moiety-conserved cycles, which makes the heart an obligatory "omnivore." Ischemia results in a transformation of efficient metabolic cycles to less-efficient linear pathways. Substrate metabolism during reperfusion requires the replenishment of depleted cycles and is a major determinant for the return of contractile function. Although there is growing recognition of the concept that regulation of substrate flux through metabolic pathways is shared by several of the pathway enzymes it is apparent that glucose oxidation and glycogen resynthesis promote the return of normal contractile function in the postischemic heart. This concept is supported by clinical observations on the beneficial effects of a solution containing glucose, insulin, and potassium (GIK) for treatment of refractory left ventricular contractile failure after hypothermic ischemic arrest during cardiac surgery.

Topics: Animals; Glucose; Glycogen; Humans; Insulin; Myocardial Ischemia; Myocardium; Oxidation-Reduction; Potassium

Topics: Animals; Disease Models, Animal; Glycogen; Ischemic Preconditioning, Myocardial; Myocardial Contraction; Myocardial Ischemia; Rabbits; Rats; Sodium-Hydrogen Exchangers; Swine

The Authors review several pharmacological interventions aimed at protecting the ischemic myocardium. Drugs which have been widely used in the treatment of ischemic heart diseases, such as beta-blockers, nitrates and calcium-antagonists, are able to delay the development of ischemic injury if administered before the beginning of ischemia, but their clinical effectiveness is limited. The new drugs which are presently investigated are designed to counteract the molecular mechanisms which mediate irreversible tissue injury, namely cytosolic calcium overload, cellular hyperosmolarity, and free radical production. In particular, interventions able to interfere with the release of calcium from its intracellular stores would be of major importance. In this regards, it is interesting to point out that derivatives of phenylalkylamine calcium-antagonists have been reported to modulate the opening probability of sarcoplasmic reticulum calcium channels.

Topics: Adenosine; Adrenergic beta-Antagonists; Calcium; Calcium Channel Agonists; Calcium Channels; Glycogen; Heart Diseases; Heat-Shock Proteins; Humans; Myocardial Ischemia; Myocardial Reperfusion; Potassium Channels; Sodium Channels

The purpose of the study was to characterize the functional and metabolic adjustments of a myocardial region subjected to low-flow ischemia. In addition, studies tested whether such myocardium retains an inotropic reserve.. Anesthetized swine were studied in which the left anterior descending coronary artery was cannulated and perfused at a constant low level causing regional contractile dysfunction (sonomicrometry for wall thickness) and the appearance of metabolic indicators of ischemia (decrease in creatine phosphate and lactate production) with only slight loss of ATP and glycogen (transmural biopsies). After 85 minutes of low-flow ischemia, dobutamine was infused into the hypoperfused artery as an inotropic challenge. Coronary hypoperfusion for 5 minutes resulted in a 54% reduction of regional systolic wall thickening, reversal of lactate consumption to lactate production, and a significant decrease in creatine phosphate. Subendocardial blood flow was reduced from 0.62 +/- 0.11 (+/- SD) to 0.16 +/- 0.07 mL.min-1.g-1. Prolonged hypoperfusion for 85 minutes resulted in no further change in regional blood flow but a partial recovery of metabolic parameters. Dobutamine infusion after 85 minutes of hypoperfusion increased regional myocardial work. However, again lactate production was significantly increase and creatine phosphate was decreased. Regional coronary hypoperfusion produces a downregulation of regional contractile function in proportion to the blood flow decrease. With prolonged hypoperfusion, after the initial adjustment phase, there is little further change in function, and metabolic markers of ischemia improve. Although the ischemic downregulated myocardium retains a significant inotropic reserve, primarily anaerobic energy production is utilized.. These data are consistent with downregulation being a protective mechanism for the ischemic myocardium to restore an energy supply-demand balance in the face of reduced blood flow. Inotropic stimulation of the downregulated myocardium enhances regional function but at the cost of worsening its metabolic status. Thus, inotropic stimulation of the hypoperfused and downregulated myocardium is probably detrimental to long-term viability.

Topics: Acute Disease; Adenosine Triphosphate; Animals; Coronary Circulation; Dobutamine; Glycogen; Heart; Myocardial Contraction; Myocardial Ischemia; Myocardium; Phosphocreatine; Swine; Time Factors

The failure to maintain the viability of ischemic myocardium is one of the mechanisms that causes ischemic heart dysfunction after revascularization. Hibernating myocardium is considered to be able to maintain long-term viability during chronic hypoperfusion. Pigment epithelium-derived factor (PEDF) decreases the contractility of hypoxic cardiomyocytes and protects cardiomyocytes against ischemic injury, which is strikingly similar to the pathophysiologic characteristics of hibernating myocardium. It was therefore postulated that PEDF may induce acute ischemic myocardium into a "hibernating-like" state to maintain its viability. Adult Sprague-Dawley rat models of acute myocardial infarction were surgically established. Lentiviral vectors carrying the

Topics: Animals; Biomarkers; Disease Models, Animal; Eye Proteins; Gene Expression; Genes, Reporter; Genetic Therapy; Genetic Vectors; Glycogen; Heart Function Tests; Hibernation; Lentivirus; Male; Myocardial Contraction; Myocardial Infarction; Myocardial Ischemia; Myocardium; Necrosis; Nerve Growth Factors; Positron-Emission Tomography; Rats; Rats, Sprague-Dawley; Serpins

Hypoxia is known to promote vasodilation of coronary vessels through several mediators including cardiac-derived adenosine and endothelium-derived prostanoids and nitric oxide. To date, the impact of endogenous glycogen depletion in vascular smooth muscle and the resultant alterations in cellular energy state (e.g., AMP-activated protein kinase, AMPK) on the contractile response to G protein-coupled receptor agonists (e.g., serotonin, 5-HT) has not yet been studied. In the present study, ex vivo exposure of endothelium-denuded human saphenous vein rings to hypoxic and glucose-deprived conditions during KCl-induced contractions for 30 min resulted in a marked depletion of endogenous glycogen by ∼80% (from ∼1.78 μmol/g under normoxia to ∼0.36 μmol/g under hypoxia). Importantly, glycogen-depleted HSV rings, which were maintained under hypoxia/reoxygenation and glucose-deprived conditions, exhibited significant increases in basal AMPK phosphorylation (∼6-fold ↑) and 5-HT-induced AMPK phosphorylation (∼19-fold ↑) with an accompanying suppression of 5-HT-induced maximal contractile response (∼68% ↓), compared with respective controls. Exposure of glycogen-depleted HSV rings to exogenous D-glucose, but not the inactive glucose analogs, prevented the exaggerated increase in 5-HT-induced AMPK phosphorylation and restored 5-HT-induced maximal contractile response. In addition, the ability of exogenous D-glucose to rescue cellular stress and impaired contractile function occurred through GLUT1-mediated but insulin/GLUT4-independent mechanisms. Together, the present findings from clinically-relevant human saphenous vein suggest that the loss of endogenous glycogen in vascular smooth muscle and the resultant accentuation of AMPK phosphorylation by GPCR agonists may constitute a yet another mechanism of metabolic vasodilation of coronary vessels in ischemic heart disease.

Topics: Aged; Allostasis; AMP-Activated Protein Kinases; Animals; Aorta, Thoracic; Biological Transport; Cell Hypoxia; Enzyme Activation; Female; Glucose; Glycogen; Glycogenolysis; Humans; In Vitro Techniques; Male; Middle Aged; Muscle, Smooth, Vascular; Myocardial Ischemia; Oxidative Stress; Phosphorylation; Protein Processing, Post-Translational; Rats, Wistar; Saphenous Vein; Vasoconstriction

Experiments on rats with acute myocardial ischemia accompanied by early postocclusive arrhythmias have shown normalizing, energy-stabilizing, and antiarrhythmic effects of uridine and uridine-5'-monophosphate. The drugs decreased lactate and restored reserves of glycogen and creatine phosphate depleted by ischemia. Uridine and uridine-5'-monophosphate significantly decreased the severity of ventricular arrhythmias. Both drugs reduced the incidence and duration of fibrillation. Uridine -5'-monophosphate demonstrated most pronounced antifibrillatory effectiveness. We hypothesize that the antiarrhythmic effect of the drugs is determined by their capacity to activate energy metabolism.

Topics: Animals; Arrhythmias, Cardiac; Coronary Vessels; Energy Metabolism; Glycogen; Lactic Acid; Ligation; Male; Myocardial Ischemia; Phosphocreatine; Rats; Rats, Wistar; Uridine; Uridine Monophosphate

Fatty acid and glucose transporters translocate between the sarcolemma and intracellular compartments to regulate substrate metabolism acutely. We hypothesised that during ischemia fatty acid translocase (FAT/CD36) would translocate away from the sarcolemma to limit fatty acid uptake when fatty acid oxidation is inhibited.. Wistar rat hearts were perfused during preischemia, low-flow ischemia, and reperfusion, using (3)H-substrates for measurement of metabolic rates, followed by metabolomic analysis and subcellular fractionation. During ischemia, there was a 32% decrease in sarcolemmal FAT/CD36 accompanied by a 95% decrease in fatty acid oxidation rates, with no change in intramyocardial lipids. Concomitantly, the sarcolemmal content of the glucose transporter, GLUT4, increased by 90% during ischemia, associated with an 86% increase in glycolytic rates, 45% decrease in glycogen content, and a 3-fold increase in phosphorylated AMP-activated protein kinase. Following reperfusion, decreased sarcolemmal FAT/CD36 persisted, but fatty acid oxidation rates returned to preischemic levels, resulting in a 35% decrease in myocardial triglyceride content. Elevated sarcolemmal GLUT4 persisted during reperfusion; in contrast, glycolytic rates decreased to 30% of preischemic rates, accompanied by a 5-fold increase in intracellular citrate levels and restoration of glycogen content.. During ischemia, FAT/CD36 moved away from the sarcolemma as GLUT4 moved toward the sarcolemma, associated with a shift from fatty acid oxidation to glycolysis, while intramyocardial lipid accumulation was prevented. This relocation was maintained during reperfusion, which was associated with replenishing glycogen stores as a priority, occurring at the expense of glycolysis and mediated by an increase in citrate levels.

Topics: Animals; CD36 Antigens; Disease Models, Animal; Energy Metabolism; Fatty Acids; Glucose Transporter Type 4; Glycogen; Glycolysis; Male; Metabolomics; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Oxidation-Reduction; Protein Transport; Rats; Rats, Wistar; Sarcolemma; Subcellular Fractions

Placental growth factor (PlGF) has a distinct biological phenotype with a predominant proangiogenic role in disease without affecting quiescent vessels in healthy organs. We tested whether systemic administration of recombinant human (rh)PlGF improves regional myocardial blood flow (MBF) and systolic function recovery in a porcine chronic myocardial ischemia model. We implanted a flow-limiting stent in the proximal left anterior descending coronary artery and measured systemic hemodynamics, regional myocardial function using MRI, and blood flow using colored microspheres 4 wk later. Animals were then randomized in a blinded way to receive an infusion of rhPlGF (15 μg·kg(-1)·day(-1), n = 9) or PBS (control; n = 10) for 2 wk. At 8 wk, myocardial perfusion and function were reassessed. Infusion of rhPlGF transiently increased PlGF serum levels >30-fold (1,153 ± 180 vs. 33 ± 18 pg/ml at baseline, P < 0.001) without affecting systemic hemodynamics. From 4 to 8 wk, rhPlGF increased regional MBF from 0.46 ± 0.11 to 0.85 ± 0.16 ml·min(-1)·g(-1), with a concomitant increase in systolic wall thickening from 11 ± 3% to 26 ± 5% in the ischemic area. In control animals, no significant changes from 4 to 8 wk were observed (MBF: 0.45 ± 0.07 to 0.49 ± 0.08 ml·min(-1)·g(-1) and systolic wall thickening: 14 ± 4% to 18 ± 1%). rhPlGF-induced functional improvement was accompanied by increased myocardial neovascularization, enhanced glycogen utilization, and reduced oxidative stress and cardiomyocyte apoptosis in the ischemic zone. In conclusion, systemic rhPlGF infusion significantly enhances regional blood flow and contractile function of the chronic ischemic myocardium without adverse effects. PlGF protein infusion may represent an attractive therapeutic strategy to increase myocardial perfusion and energetics in chronic ischemic cardiomyopathy.

Topics: Animals; Apoptosis; Coronary Circulation; Glycogen; Heart Ventricles; Hemodynamics; Magnetic Resonance Imaging; Myocardial Contraction; Myocardial Ischemia; Myocardial Revascularization; Myocytes, Cardiac; Oxidative Stress; Placenta Growth Factor; Pregnancy Proteins; Regional Blood Flow; Sus scrofa; Ventricular Dysfunction

The effects of ischemic-postconditioning (IPOC) on functional recovery and cell viability of ischemic-reperfused hearts from fed and fasted rats were studied in relation to triacylglycerol and glycogen mobilization, ATP content, glucose-6-phosphate dehydrogenase activity and reduced/oxidized glutathione (GSH/GSSG). Oxidative damage was estimated by measuring thiobarbituric acid reactive substances (TBARS). IPOC improved contractile recovery and cell viability in the fed but attenuated them in the fasted hearts. In both groups ischemia lowered glycogen. IPOC further reduced it. Triacylglycerol remained unchanged during ischemia-reperfusion in both groups, but triacylglycerol mobilization was activated by IPOC in the fasted group. ATP was increased by IPOC in the fed hearts, but lowered in the fasted ones, which appeared to be associated with the rates of ATP synthesis in isolated mitochondria. In the fed hearts IPOC raised glucose-6-phosphate dehydrogenase activity and GSH/GSSG, and lowered TBARS. These results suggest that IPOC effects are associated with changes in the ATP supply, mobilization of energy sources and glutathione antioxidant ratio.

Topics: Adenosine Triphosphate; Animals; Blood Pressure; Cell Survival; Energy Metabolism; Fasting; Female; Glucosephosphate Dehydrogenase; Glutathione; Glutathione Disulfide; Glycine; Glycogen; Heart; Heart Rate; Ischemic Postconditioning; Mitochondria, Heart; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardial Reperfusion Injury; Myocardium; Myocytes, Cardiac; Rats; Rats, Wistar; Thiobarbituric Acid Reactive Substances; Triglycerides; Ventricular Dysfunction, Left

Changes in metabolic and myofilament phenotypes coincide in developing hearts. Posttranslational modification of sarcomere proteins influences contractility, affecting the energetic cost of contraction. However, metabolic adaptations to sarcomeric phenotypes are not well understood, particularly during pathophysiological stress. This study explored metabolic adaptations to expression of the fetal, slow skeletal muscle troponin I (ssTnI). Hearts expressing ssTnI exhibited no significant ATP loss during 5 min of global ischemia, while non-transgenic littermates (NTG) showed continual ATP loss. At 7 min ischemia TG-ssTnI hearts retained 80±12% of ATP versus 49±6% in NTG (P<0.05). Hearts expressing ssTnI also had increased AMPK phosphorylation. The mechanism of ATP preservation was augmented glycolysis. Glycolytic end products (lactate and alanine) were 38% higher in TG-ssTnI than NTG at 2 min and 27% higher at 5 min. This additional glycolysis was supported exclusively by exogenous glucose, and not glycogen. Thus, expression of a fetal myofilament protein in adult mouse hearts induced elevated anaerobic ATP production during ischemia via metabolic adaptations consistent with the resistance to hypoxia of fetal hearts. The general findings hold important relevance to both our current understanding of the association between metabolic and contractile phenotypes and the potential for invoking cardioprotective mechanisms against ischemic stress. This article is part of a Special Issue entitled "Possible Editorial".

Topics: Adenosine Triphosphate; AMP-Activated Protein Kinases; Animals; Disease Models, Animal; Gene Expression Regulation; Glycogen; Glycolysis; Male; Mice; Mice, Transgenic; Muscle Fibers, Slow-Twitch; Myocardial Ischemia; Myocardium; Phosphorylation; Troponin I

Carnitine is a vital biologic substance facilitating fatty acids transport into mitochondria for ATP production. This study was to investigate the effects of pre-ischemic pharmacological preconditioning (PC) with L-carnitine (L-Car) on myocardial infarct size and cardiac functions in ischemic and reperfused isolated rat heart and meanwhile on left ventricular glycogen and lactate content. Isolated rat hearts were subjected to 30 min coronary artery occlusion followed by 120 min reperfusion. The hearts (n= 8-12) were perfused with L-Car (0.5-5 mM) only for 15 min before to 10 min after induction of ischemia. Preconditioning of the hearts with L-Car provided concentration-dependent cardioprotection as evidenced by improved postischemic ventricular functional recovery (developed pressure, left ventricular end diastolic pressure and coronary flow rate) and reduced myocardial infarct size (p<0.001). L-Car (2.5 mM) decreased both glycogen (p<0.001) and lactate (p>0.05) content in left ventricle during ischemia compared with the control. The results of this study demonstrate that L-Car pharmacologically precondition the hearts against ischemic and reperfusion injury in part by recovery of postischemic ventricular hemodynamic functions, depletion of glycogen and therefore reduction of lactate accumulation.

Topics: Adenosine Triphosphate; Animals; Carnitine; Glycogen; Hemodynamics; Ischemic Preconditioning, Myocardial; Lactic Acid; Male; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Rats; Rats, Sprague-Dawley

The goal of the present study was to assess the effects of a restricted feeding schedule (RFS) on postischemic contractile recovery in relation to triacylglycerol (TAG), glycogen, and ATP content. Glucose-6-phosphate dehydrogenase (G6PDH) activity, reduced/oxidized glutathione ratio (GSH/GSSG), and thiobarbituric acid reactive substances (TBARS) levels were also determined. Isolated rat hearts entrained to daily RFS (2 h food access starting at 1200) or fed ad libitum (FED) for 3 weeks were Langendorff-perfused (25 min ischemia, 30 min reperfusion) with Krebs-Ringer bicarbonate solution (10 mmol/L glucose). RFS improved the recovery of contractility and reduced creatine kinase (CK) release upon reperfusion. Further, at the end of reperfusion, RFS hearts exhibited increased G6PDH activity and repletion of tissue glycogen, TAG, and ATP that was not observed in the FED hearts. GSH/GSSG at the end of reperfusion fell to the same value in both nutritional states, and TBARS levels were higher in the RFS hearts. In conclusion, RFS improved postischemic functional recovery, which was accompanied by a reduction in CK release and a striking energy recovery. Although enhanced G6PDH activity was displayed, RFS was unable to reduce lipid peroxidation, supporting a clear dissociation between protection against mechanical dysfunction and CK release on the one hand and oxidative damage on the other.

Topics: Adenosine Triphosphate; Animals; Caloric Restriction; Creatine Kinase; Disease Models, Animal; Female; Glucosephosphate Dehydrogenase; Glutathione; Glycogen; Heart Function Tests; Heart Ventricles; Lipid Peroxidation; Myocardial Contraction; Myocardial Ischemia; Perfusion; Rats; Rats, Wistar; Triglycerides; Ventricular Function, Left

Ischaemic pre-conditioning (IP) is a potent protective mechanism for limiting the myocardial damage due to ischaemia. It is not fully known as to how IP protects. The metabolism of adenosine may be an important mechanistic component. We study the role of adenosine turnover together with glycolytic flow in ischaemic myocardium subjected to IP.. An acute myocardial ischaemia pig model was used, with microdialysis sampling of some metabolites (lactate, adenosine, glucose, glycerol, taurine) of ischaemic myocardium. An IP group was compared with a control group before and during a prolonged ischaemia. ¹⁴C-labelled adenosine and glucose were infused through microdialysis probes, and lactate, ¹⁴C-labelled lactate, glucose, taurine and glycerol were analysed in the effluent. The glycogen content in myocardial biopsies was determined.. The ¹⁴C-adenosine metabolism was higher as there was a higher production of ¹⁴C-lactate in IP animals compared with the controls. The glycolytic flow, measured as myocardial lactate formation, was retarded during prolonged ischaemia in IP animals. Myocardial free glucose and glycogen content decreased during the prolonged ischaemia in both groups, with higher free glucose in the IP group. We confirmed the protective effects of IP with lower myocardial concentrations of markers for cellular damage (glycerol).. This association between increased adenosine turnover and decreased glycolytic flow during prolonged ischaemia in response to IP can possibly be explained by the competitive effect for the metabolites from both glucose and adenosine metabolism for entering glycolysis. We conclude that this study provides support for an energy-metabolic explanation for the protective mechanisms of IP.

Topics: Adenosine; Animals; Blood Glucose; Body Temperature; Energy Metabolism; Female; Glycerol; Glycogen; Glycolysis; Hemodynamics; Ischemic Preconditioning, Myocardial; Lactic Acid; Microdialysis; Myocardial Ischemia; Swine; Taurine

It is unknown what effects high levels of fatty acids have on energy metabolism and cardiac efficiency during milder forms of ischemia. To address this issue, isolated working rat hearts perfused with Krebs-Henseleit solution (5 mM glucose, 100 muU/mL insulin, and 0.4 (Normal Fat) or 1.2 mM palmitate (High Fat)) were subjected to 30 min of aerobic perfusion followed by 30 min of mild ischemia (39% reduction in coronary flow). Both groups had similar aerobic function and rates of glycolysis, however the High Fat group had elevated rates of palmitate oxidation (150%), and decreased rates of glucose oxidation (51%). Mild ischemia decreased cardiac work (56% versus 40%) and efficiency (29% versus 11%) further in High Fat hearts. Palmitate oxidation contributed a greater percent of acetyl-CoA production during mild ischemia in the High Fat group (81% versus 54%). During mild ischemia glycolysis remained at aerobic levels in the Normal Fat group, but was accelerated in the High Fat group. Triglyceride, glycogen and adenine nucleotide content did not differ at the end of mild ischemia, however glycogen turnover was double in the High Fat group (248%). Addition of the pyruvate dehydrogenase inhibitor dichloroacetate to the High Fat group resulted in a doubling of the rate of glucose oxidation and improved cardiac efficiency during mild ischemia. We demonstrate that fatty acid oxidation dominates as the main source of residual oxidative metabolism during mild ischemia, which is accompanied by suppressed cardiac function and efficiency in the presence of high fat.

Topics: Adenine Nucleotides; Animals; Dichloroacetic Acid; Fatty Acids; Glucose; Glycogen; Glycolysis; Heart; In Vitro Techniques; Male; Myocardial Ischemia; Myocardium; Oxidation-Reduction; Palmitates; Perfusion; Rats; Rats, Sprague-Dawley; Triglycerides

Activation of 5'-AMP-activated protein kinase (AMPK) may benefit the heart during ischemia-reperfusion by increasing energy production. While AMPK stimulates glycolysis, mitochondrial oxidative metabolism is the major source of ATP production during reperfusion of ischemic hearts. Stimulating AMPK increases mitochondrial fatty acid oxidation, but this is usually accompanied by a decrease in glucose oxidation, which can impair the functional recovery of ischemic hearts. To examine the relationship between AMPK and cardiac energy substrate metabolism, we subjected isolated working mouse hearts expressing a dominant negative (DN) alpha(2)-subunit of AMPK (AMPK-alpha(2) DN) to 20 min of global no-flow ischemia and 40 min of reperfusion with Krebs-Henseleit solution containing 5 mM [U-(14)C]glucose, 0.4 mM [9, 10-(3)H]palmitate, and 100 microU/ml insulin. AMPK-alpha(2) DN hearts had reduced AMPK activity at the end of reperfusion (82 +/- 9 vs. 141 +/- 7 pmol.mg(-1).min(-1)) with no changes in high-energy phosphates. Despite this, AMPK-alpha(2) DN hearts had improved recovery of function during reperfusion (14.9 +/- 0.8 vs. 9.4 +/- 1.4 beats.min(-1).mmHg.10(-3)). During reperfusion, fatty acid oxidation provided 44.0 +/- 2.8% of total acetyl-CoA in AMPK-alpha(2) DN hearts compared with 55.0 +/- 3.2% in control hearts. Since insulin can inhibit both AMPK activation and fatty acid oxidation, we also examined functional recovery in the absence of insulin. Functional recovery was similar in both groups despite a decrease in AMPK activity and a decreased reliance on fatty acid oxidation during reperfusion (66.4 +/- 9.4% vs. 85.3 +/- 4.3%). These data demonstrate that the suppression of cardiac AMPK activity does not produce an energetically compromised phenotype and does not impair, but may in fact improve, the recovery of function after ischemia.

Topics: Acetyl Coenzyme A; Adenine Nucleotides; Aerobiosis; AMP-Activated Protein Kinases; Animals; Cyclic AMP-Dependent Protein Kinases; Energy Metabolism; Enzyme Inhibitors; Fatty Acids, Nonesterified; Glycogen; Hypoglycemic Agents; In Vitro Techniques; Insulin; Mice; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion Injury; Palmitates; Recovery of Function

Nitric oxide (NO) may limit myocardial ischemia-reperfusion injury by slowing the mitochondrial metabolism. We examined whether rat heart contains catalysts potentially capable of reducing nitrite to NO during an episode of regional myocardial ischemia produced by temporary coronary artery occlusion. In intact Sprague-Dawley rats, a 15-min coronary occlusion lowered the nitrite concentration of the myocardial regions exhibiting ischemic glucose metabolism to approximately 50% that of nonischemic regions (185 +/- 223 vs. 420 +/- 203 nmol/l). Nitrite was rapidly repleted during subsequent reperfusion. The heart tissue tested in vitro acquired a substantial ability to consume nitrite when made hypoxic at neutral pH, and this ability was slightly enhanced by simultaneously lowering the pH to 5.5. More than 70% of this activity could be abolished by flushing the coronary circulation with crystalloid to remove trapped erythrocytes. Correspondingly, erythrocytes demonstrated the ability to reduce exogenous nitrite to NO under hypoxic conditions in vitro. In erythrocyte-free heart tissue, the nitrite consumption increased fivefold when the pH was lowered to 5.5. Approximately 40% of this pH-sensitive increase in nitrite consumption could be blocked by the xanthine oxidoreductase inhibitor allopurinol, whereas lowering the Po(2) sufficiently to desaturate myoglobin accelerated it further. We conclude that rat heart contains several factors capable of catalyzing ischemic nitrite reduction; the most potent is contained within erythrocytes and activated by hypoxia, whereas the remainder includes xanthine oxidoreductase and other pH-sensitive factors endogenous to heart tissue, including deoxymyoglobin.

Topics: Allopurinol; Animals; Catalysis; Disease Models, Animal; Enzyme Inhibitors; Erythrocytes; Glucose; Glycogen; Hydrogen-Ion Concentration; Hypoxia; Male; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Myoglobin; Nitric Oxide; Nitrites; Oxidation-Reduction; Rats; Rats, Sprague-Dawley; Time Factors; Xanthine Dehydrogenase

Ventricular dysfunction is reported greater in the left (LV) versus right ventricle (RV) in infants following surgically induced ischemia. Ventricle-specific differences in baseline metabolism may alter response to ischemia thus affecting postischemic functional recovery. This study identifies ventricle-specific metabolic differences in the newborn (piglet) heart at baseline (working) and during ischemia (arrested). Baseline LV citrate synthase (CS) and hydroxyacyl-CoA dehydrogenase (HAD) activities were 15% and 18% lower (p < 0.02), whereas creatine kinase (CK) and phosphofructokinase (PFK) activities were 40% and 23% higher (p < 0.04) than the RV. Baseline LV glycogen reserves were also 55% higher (p = 0.004). By 15 min of ischemia, LV ATP was 20% lower (p < 0.05), lactate was 51% higher (p = 0.001), and hydrogen ions (H) were 43% higher (p = 0.03) compared with the RV. These differences persisted for the entire ischemic period (p < 0.02). After 45 min of ischemia, the LV used 58% less (p < 0.05) glycogen than the RV. These findings demonstrate that the enhanced glycolytic capacity of the newborn LV was accompanied by greater anaerobic end-product accumulation and lower energy levels during ischemia. This profile may offer one explanation for greater LV-dysfunction relative to the RV in children following ischemia.

Topics: 3-Hydroxyacyl CoA Dehydrogenases; Adaptation, Physiological; Adenosine Triphosphate; Animals; Animals, Newborn; Citrate (si)-Synthase; Creatine Kinase; Energy Metabolism; Glycogen; Glycolysis; Heart Arrest, Induced; Heart Ventricles; Hydrogen-Ion Concentration; Lactic Acid; Myocardial Ischemia; Myocardium; Phosphofructokinases; Swine; Time Factors; Ventricular Dysfunction, Left; Ventricular Dysfunction, Right

Alterations in myocardial glucose metabolism are a key determinant of ischemia-induced depression of left ventricular mechanical function. Since myocardial glycogen is an important source of endogenous glucose, we compared the effects of ischemia on glucose uptake and utilization in isolated working rat hearts in which glycogen content was either replete (G replete, 114 micromol/g dry wt) or partially depleted (G depleted, 71 mumol/g dry wt). The effects of low-flow ischemia (LFI, 0.5 ml/min) on glucose uptake, glycogen turnover (glycogenolysis and glycogen synthesis), glycolysis, adenosine 5'-monophosphate-activated protein kinase (AMPK) activity, and GLUT4 translocation were measured. Relative to preischemic values, LFI caused a time-dependent reduction in glycogen content in both G-replete and G-depleted groups due to an acceleration of glycogenolysis (by 12-fold and 6-fold, respectively). In G-replete hearts, LFI (15 min) decreased glucose uptake (by 59%) and did not affect GLUT4 translocation. In G-depleted hearts, LFI also decreased initially glucose uptake (by 90%) and glycogen synthesis, but after 15 min, when glycogenolysis slowed due to exhaustion of glycogen content, glucose uptake increased (by 31%) in association with an increase in GLUT4 translocation. After 60 min of LFI, glucose uptake, glycogenolysis, and glycolysis recovered to near-preischemic values in both groups. LFI increased AMPK activity in a time-dependent manner in both groups (by 6-fold and 4-fold, respectively). Thus, when glycogen stores are replete before ischemia, ischemia-induced AMPK activation is not sufficient to increase glucose uptake. Under these conditions, an acceleration of glycogen degradation provides sufficient endogenous substrate for glycolysis during ischemia.

Topics: Adenosine Monophosphate; Adenosine Triphosphate; Aerobiosis; Animals; Cell Membrane; Creatine; Cyclic AMP-Dependent Protein Kinases; Energy Metabolism; Enzyme Activation; Fatty Acids; Glucose; Glucose Transporter Type 4; Glycogen; Glycolysis; Hypoglycemic Agents; In Vitro Techniques; Insulin; Male; Myocardial Ischemia; Myocardium; Phosphocreatine; Rats; Rats, Sprague-Dawley; Sarcolemma; Ventricular Function, Left

During ischemia, endogenous glycogen becomes the principal substrate for energy through glycolysis. Cardiac-specific manipulation of AMP-activated protein kinase (AMPK) by over-expression of its regulatory gamma-subunit induces glycogen storage. The aim of this study was to examine whether heart glycogen in transgenic mice overexpressing PRKAG2 may protect from ischemia and reperfusion injury. Isolated hearts were mounted on Langendorff apparatus and subjected to 30 min 'no-flow' or 'low-flow' ischemia and 60 min reperfusion. Hemodynamic measurements, tetrazolium staining, glycogen and lactate were used to monitor ischemia reperfusion damage. After low-flow ischemia, left ventricular pressure, coronary flow (CF) and the area of viable myocardium were 20-30% higher in PRKAG2 mice compared to controls. The basal levels of glycogen in PRKAG2 were 9.2 microg/g, markedly higher than in controls, but after low-flow ischemia they declined concomitantly with increased lactate washout in the coronary effluent. During no-flow ischemia there was neither protection nor consumption of glycogen in PRKAG2 hearts. Cardioprotection was also eliminated when PRKAG2 hearts were depleted of glycogen prior to low-flow ischemia. AMPK alpha Thr172 phosphorylation did not differ between PRKAG2 hearts and controls either during low-flow ischemia or reperfusion. We conclude that PRKAG2 hearts resist low-flow ischemia injury better than controls. Improved recovery was associated with increased consumption of glycogen, and was unrelated to AMPK activation. These findings demonstrate the potential of heart protection from ischemia and reperfusion injury through metabolic manipulation increasing the level and utilization of myocardial glycogen.

Topics: AMP-Activated Protein Kinases; Animals; Gene Expression Regulation, Enzymologic; Glycogen; Humans; In Vitro Techniques; Male; Mice; Mice, Transgenic; Multienzyme Complexes; Myocardial Ischemia; Myocardial Reperfusion Injury; Protein Serine-Threonine Kinases; Rabbits

Adenosine-induced acceleration of glycolysis in hearts stressed by transient ischemia is accompanied by suppression of glycogen synthesis and by increases in activity of adenosine 5'-monophosphate-activated protein kinase (AMPK). Because p38 mitogen-activated protein kinase (MAPK) may regulate glucose metabolism and may be activated downstream of AMPK, this study determined the effects of the p38 MAPK inhibitors SB202190 and SB203580 on adenosine-induced alterations in glucose utilization and AMPK activity. Studies were performed in working rat hearts perfused aerobically following stressing by transient ischemia (2 x 10-min ischemia followed by 5-min reperfusion). Phosphorylation of AMPK and p38 MAPK each were increased fourfold by adenosine, and these effects were inhibited by either SB202190 or SB203580. Neither of these inhibitors directly affected AMPK activity. Attenuation of the adenosine-induced increase in AMPK and p38 MAPK phosphorylation by SB202190 and SB203580 occurred independently of any change in tissue ATP-to-AMP ratio and did not alter glucose uptake, but it was accompanied by an increase in glycogen synthesis and glycogen content and by inhibition of glycolysis and proton production. There was a significant inverse correlation between the rate of glycogen synthesis and AMPK activity and between AMPK activity and glycogen content. These data demonstrate that AMPK is likely downstream of p38 MAPK in mediating the effects of adenosine on glucose utilization in hearts stressed by transient ischemia. The ability of p38 MAPK inhibitors to relieve the inhibition of glycogen synthesis and to inhibit glycolysis and proton production suggests that these agents may restore adenosine-induced cardioprotection in stressed hearts.

Topics: Adenosine; AMP-Activated Protein Kinases; Animals; Coronary Circulation; Enzyme Inhibitors; Glucose; Glycogen; Imidazoles; Male; Multienzyme Complexes; Myocardial Ischemia; Myocardium; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Protein Serine-Threonine Kinases; Pyridines; Rats; Rats, Sprague-Dawley

Because the question "is AMP-activated protein kinase (AMPK) alpha(2)-isoform a friend or a foe in the protection of the myocardium against ischemia-reperfusion injury?" is still in debate, we studied the functional consequence of its deletion on the contractility, the energetics, and the respiration of the isolated perfused heart and characterized the response to low-flow ischemia and reperfusion with glucose and pyruvate as substrates. alpha(2)-AMPK deletion did not affect basal contractility, respiration, and high-energy phosphate contents but induced a twofold reduction in glycogen content and a threefold reduction in glucose uptake. Low-flow ischemia increased AMPK phosphorylation and stimulated glucose uptake and phosphorylation in both alpha(2)-knockout (alpha(2)-KO) and wild-type (WT) groups. The high sensitivity of alpha(2)-KO to the development of ischemic contracture was attributed to the constitutive impairment in glucose transport and glycogen content and not to a perturbation of the energy transfer by creatine kinase (CK). The functional coupling of MM-CK to myofibrillar ATPase and the CK fluxes were indeed similar in alpha(2)-KO and WT. Low-flow ischemia impaired CK flux by 50% in both strains, showing that alpha(2)-AMPK does not control CK activity. Despite the higher sensitivity to contracture, the postischemic contractility recovered to similar levels in both alpha(2)-KO and WT in the absence of fatty acids. In their presence, alpha(2)-AMPK deletion also accelerated the contracture but delayed postischemic contractile recovery. In conclusion, alpha(2)-AMPK is required for a normal glucose uptake and glycogen content, which protects the heart from the development of the ischemic contracture, but not for contractile recovery in the absence of fatty acids.

Topics: Adenosine Triphosphate; AMP-Activated Protein Kinases; Animals; Cell Respiration; Creatine Kinase, MM Form; Energy Metabolism; Enzyme Activation; Fatty Acids; Glucose; Glycogen; In Vitro Techniques; Kinetics; Male; Mice; Mice, Knockout; Multienzyme Complexes; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Oxygen Consumption; Perfusion; Phosphocreatine; Phosphorylation; Protein Serine-Threonine Kinases; Pyruvic Acid

p38 mitogen-activated protein kinase (MAPK) and 5'-AMP-activated protein kinase (AMPK) are activated by metabolic stresses and are implicated in the regulation of glucose utilization and ischemia-reperfusion (IR) injury. This study tested the hypothesis that inhibition of p38 MAPK restores the cardioprotective effects of adenosine in stressed hearts by preventing activation of AMPK and the uncoupling of glycolysis from glucose oxidation. Working rat hearts were perfused with Krebs solution (1.2 mM palmitate, 11 mM [(3)H/(14)C]glucose, and 100 mU/l insulin). Hearts were stressed by transient antecedent IR (2 x 10 min I/5 min R) before severe IR (30 min I/30 min R). Hearts were treated with vehicle, p38 MAPK inhibitor (SB-202190, 10 microM), adenosine (500 microM), or their combination before severe IR. After severe IR, the phosphorylation (arbitrary density units) of p38 MAPK and AMPK, rates of glucose metabolism (micromol x g dry wt(-1) x min(-1)), and recovery of left ventricular (LV) work (Joules) were similar in vehicle-, SB-202190- and adenosine-treated hearts. Treatment with SB-202190 + adenosine versus adenosine alone decreased p38 MAPK (0.03 +/- 0.01, n = 3 vs. 0.48 +/- 0.10, n = 3, P < 0.05) and AMPK (0.00 +/- 0.00, n = 3 vs. 0.26 +/- 0.08, n = 3 P < 0.05) phosphorylation. This was accompanied by attenuated rates of glycolysis (1.51 +/- 0.40, n = 7 vs. 3.95 +/- 0.65, n = 7, P < 0.05) and H(+) production (2.12 +/- 0.76, n = 7 vs. 6.96 +/- 1.48, n = 7, P < 0.05), and increased glycogen synthesis (1.91 +/- 0.25, n = 6 vs. 0.27 +/- 0.28, n = 6, P < 0.05) and improved recovery of LV work (0.81 +/- 0.08, n = 7 vs. 0.30 +/- 0.15, n = 8, P < 0.05). These data indicate that inhibition of p38 MAPK abolishes subsequent phosphorylation of AMPK and improves the coupling of glucose metabolism, thereby restoring adenosine-induced cardioprotection.

Topics: Adenosine; AMP-Activated Protein Kinases; Animals; Cardiotonic Agents; Disease Models, Animal; Glucose; Glycogen; Glycolysis; Hydrogen-Ion Concentration; Imidazoles; In Vitro Techniques; Male; Multienzyme Complexes; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Protein Kinase Inhibitors; Protein Serine-Threonine Kinases; Pyridines; Rats; Rats, Sprague-Dawley; Severity of Illness Index; Ventricular Function, Left

AMP-activated protein kinase (AMPK) is a major sensor and regulator of the energetic state of the cell. Little is known about the specific role of AMPKalpha(2), the major AMPK isoform in the heart, in response to global ischemia. We used AMPKalpha(2)-knockout (AMPKalpha(2)(-/-)) mice to evaluate the consequences of AMPKalpha(2) deletion during normoxia and ischemia, with glucose as the sole substrate. Hemodynamic measurements from echocardiography of hearts from AMPKalpha(2)(-/-) mice during normoxia showed no significant modification compared with wild-type animals. In contrast, the response of hearts from AMPKalpha(2)(-/-) mice to no-flow ischemia was characterized by a more rapid onset of ischemia-induced contracture. This ischemic contracture was associated with a decrease in ATP content, lactate production, glycogen content, and AMPKbeta(2) content. Hearts from AMPKalpha(2)(-/-) mice were also characterized by a decreased phosphorylation state of acetyl-CoA carboxylase during normoxia and ischemia. Despite an apparent worse metabolic adaptation during ischemia, the absence of AMPKalpha(2) does not exacerbate impairment of the recovery of postischemic contractile function. In conclusion, AMPKalpha(2) is required for the metabolic response of the heart to no-flow ischemia. The remaining AMPKalpha(1) cannot compensate for the absence of AMPKalpha(2).

Topics: Acetyl-CoA Carboxylase; Adenine; Adenosine Triphosphate; AMP-Activated Protein Kinases; Animals; Fatty Acids; Gene Expression Regulation, Enzymologic; Glycogen; Isoenzymes; Lactates; Mice; Mice, Knockout; Multienzyme Complexes; Myocardial Contraction; Myocardial Ischemia; Myocardium; Phenotype; Protein Kinases; Protein Serine-Threonine Kinases

Hypothyroid heart displays a phenotype of cardioprotection against ischemia and this study investigated whether administration of dronedarone, an amiodarone-like compound that has been shown to preferentially antagonize thyroid hormone binding to thyroid hormone receptor alpha1 (TRalpha1), results in a similar effect. Dronedarone was given in Wistar rats (90 mg/kg, once daily (od) for 2 weeks) (DRON), while untreated animals served as controls (CONT). Hypothyroidism (HYPO) was induced by propylthiouracil administration. Isolated rat hearts were perfused in Langendorff mode and subjected to 20 minutes of zero-flow global ischemia (I) followed by 45 minutes of reperfusion (R). 3,5,3' Triiodothyronine remained unchanged while body weight and food intake were reduced. alpha-Myosin heavy chain (alpha-MHC) decreased in DRON while beta-myosin heavy chain (beta-MHC) and sarcoplasmic reticulum Ca2+ adenosine triphosphatase (ATPase) expression (SERCA) was similar to CONT. In HYPO, alpha-MHC and SERCA were decreased while beta-MHC was increased. Myocardial glycogen content was increased in both DRON and HYPO. In DRON, resting heart rate and contractility were reduced and ischemic contracture was significantly suppressed while postischemic left ventricular end-diastolic pressure and lactate dehydrogenase release (IU/L min) after I/R were significantly decreased. In conclusion, dronedarone treatment results in cardioprotection by selectively mimicking hypothyroidism. This is accompanied by a reduction in body weight because of the suppression of food intake. TRs might prove novel pharmacologic targets for the treatment of cardiovascular illnesses.

Topics: Adaptation, Physiological; Amiodarone; Animals; Calcium-Transporting ATPases; Dronedarone; Eating; Glycogen; Heart; Heart Rate; Hypothyroidism; In Vitro Techniques; Isomerism; L-Lactate Dehydrogenase; Male; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Myosins; Rats; Rats, Wistar; Sarcoplasmic Reticulum; Thyroid Hormone Receptors alpha; Thyroid Hormones; Weight Gain

ATP-sensitive potassium (KATP) channels are involved in the mechanisms underlying ischemic preconditioning. KATP channels open during ischemia, presumably secondary to intracellular metabolic alterations. The direct effects of KATP channel modulation on myocardial metabolism have not been studied. The aim of the present study was to investigate whether a KATP opener (diazoxide) and blocker (glibenclamide) modulates myocardial glycogen, lactate, and amino acid content before, during, and after ischemia. In isolated perfused rat hearts, we investigated the effect of diazoxide (30 microM) and glibenclamide (10 microM) administered 15 minutes before ischemia on myocardial glycogen, lactate, and amino acid content before, during, and after ischemia. Diazoxide increased left-ventricular developed pressure during reperfusion (P < 0.05) and decreased myocardial glycogen depletion (P < 0.05) and lactate accumulation (P < 0.05) during ischemia compared with the control group. Glibenclamide decreased myocardial glycogen content (P < 0.05) and increased myocardial lactate (P < 0.05) and alanine (P < 0.05) content before ischemia and reduced myocardial glycogen content after ischemia (P < 0.05) compared with control. KATP channel activation by diazoxide modulates myocardial metabolism. These findings suggest that activation of KATP channels protects against ischemia-reperfusion injury by a mechanism that involves decreased energy depletion.

Topics: Alanine; Amino Acids; Animals; Diazoxide; Drug Interactions; Glutamic Acid; Glyburide; Glycogen; Lactic Acid; Male; Myocardial Ischemia; Myocardium; Potassium Channel Blockers; Potassium Channels; Rats; Rats, Wistar

Pre-diabetic subjects with high insulin secretory capacity have double risk of cardiovascular disease compared with subjects who do not develop insulin-resistance. It is well established that the ability of the myocardium to increase its glycolytic ATP production plays a crucial role in determining cell survival under conditions of ischemia. Up to now, whether the pre-diabetic state reduces the tolerance of the heart to ischemia by affecting its ability to increase its energy production through glycolysis remains unknown. The aim of the present study was to assess whether insulin resistance affects the ability of the myocardium to increase glycolysis under ischemic conditions. Male Wistar rats were fed for 8 weeks a fructose-enriched (33%) diet to induce a pre-diabetic state. Hearts were isolated and subjected to ex-vivo low-flow (2%) ischemia for 30 min. The fructose diet increased sarcolemmal GLUT4 localisation in myocardial cells under basal conditions compared with controls. This effect was not accompanied by increased glucose utilisation. Ischemia induced the translocation of GLUT4 to the plasma membrane in controls but did not significantly modify the distribution of these transporters in pre-diabetic hearts. Glycolytic flux under ischemic conditions was significantly lower in fructose-fed rat hearts compared with controls. The reduction of glycolytic flux during ischemia in fructose-fed rat hearts was not due to metabolic inhibition downstream hexokinase II since no cardiac accumulation of glucose-6-phosphate was detected. In conclusion, our results suggest that the pre-diabetic state reduces the tolerance of the myocardium to ischemia by decreasing glycolytic flux adaptation.

Topics: Adaptation, Physiological; Animals; Diabetes Mellitus, Experimental; Fructose; Glucose; Glucose Transporter Type 4; Glycogen; Glycolysis; Hexosephosphates; In Vitro Techniques; Insulin Resistance; Lactic Acid; Male; Monosaccharide Transport Proteins; Muscle Proteins; Myocardial Ischemia; Myocardium; Myocytes, Cardiac; Prediabetic State; Protein Transport; Rats; Rats, Wistar; Sarcolemma

A computational model of myocardial energy metabolism was used to assess the metabolic responses to normal and reduced myocardial blood flow. The goal was to examine to what extent glycolysis and lactate formation are controlled by the supply of glycolytic substrate and/or the cellular redox (NADH/NAD+) and phosphorylation (ATP/ADP) states.. Flow was reduced over a wide range and for a sufficient duration in order to investigate the sequence of events that occur during the transition to a new metabolic steady state.. Simulation results indicated multiple time-dependent controls over both glycolysis and lactate formation.. Changes in phosphorylation state and glucose uptake only significantly affect the initial phase of the glycolytic response to ischemia, while glycogen breakdown exerts control over glycolysis during the entire duration of ischemia. Similarly, changes in the redox state affect the rates of lactate formation and release primarily during the initial transient phase of the response to the reductions in blood flow, while the rate of glycolysis controls the rate of lactate formation throughout the entire period of adaptation.

Topics: Adenosine Triphosphate; Glycogen; Glycolysis; Lactic Acid; Models, Cardiovascular; Myocardial Ischemia; Myocardium; NAD; Oxidation-Reduction; Oxygen Consumption; Phosphocreatine; Phosphorylation; Pyruvic Acid

Our laboratory demonstrated that mild regional hypothermia reduced myocardial infarct size by an average of 65% in the rabbit model of regional ischemia. The exact mechanism for this benefit has not been explored. We hypothesized that a moderate reduction in regional myocardial temperature could preserve cardiac energy metabolism and thus protect the myocardium from sustained ischemic insult.. Anesthetized open-chest rabbits were randomized to normothermic sham-operated (NS, n = 6), hypothermic sham-operated (HS, n = 6), normothermic ischemic (NI, n = 10), and hypothermic ischemic (HI, n = 10) groups. Both sham-operated groups received no occlusions, and both ischemic groups were subjected to 20 minutes of coronary occlusion. To achieve regional cooling of the hearts in the hypothermic groups, a bag of ice water was placed directly on the risk area 15 minutes prior to coronary artery occlusion/no intervention and maintained for the duration of the subsequent 20 minutes of ischemia/no intervention (in the HI and HS groups respectively). Hypothermia preserved adenosine triphosphate (ATP) and glycogen stores in the ischemic area by 42.9% and 84.2%, respectively (1.20 +/- 0.11 micromoles ATP/g wet tissue vs 0.84 +/- 0.06 micromoles ATP/g wet tissue and 8.16 +/- 0.95 micromoles of glucosyl unit/g wet tissue vs 4.43 +/- 0.44 micromoles of glucosyl unit/g wet tissue in the HI and the NI groups, respectively). In addition, hypothermia resulted in a trend toward creatine phosphate preservation in the nonischemic area.. This is the first demonstration that local therapy with mild reductions in myocardial temperature preserves energy metabolism both in the ischemic and the nonischemic areas as well. The preservation in ATP is the likely mechanism by which regional hypothermia is preserving ischemic myocardium.

Topics: Adenosine Triphosphate; Animals; Energy Metabolism; Glycogen; Hypothermia, Induced; Male; Myocardial Ischemia; Myocardium; Rabbits

This study tested the robustness of our computational model of myocardial metabolism by comparing responses to two different inputs with experimental data obtained in pigs under similar conditions. Accordingly, an abrupt and a gradual reduction in coronary flow of similar magnitude were implemented and used as model input. After flow reductions reached 60% from control values, ischemia was kept constant for 60 min in both groups. Our hypotheses were that: (1) these two flow-reduction profiles would result in different transients (concentrations and flux rates) while having similar steady-state values and (2) our model-simulated responses would predict the experimental results in an anesthetized swine model of myocardial ischemia. The two different ischemia-induction patterns resulted in the same decrease in steady-state MVO2 and in similar steady-state values for metabolite concentrations and flux rates at 60 min of ischemia. While both the simulated and experimental results showed decreased glycogen concentration, accumulation of lactate, and net lactate release with ischemia, the onset of glycogen depletion and the switch to lactate efflux were more rapid in the experiments than in the simulations. This study demonstrates the utility of computer models for predicting experimental outcomes in studies of metabolic regulation under physiological and pathological conditions.

Topics: Animals; Computer Simulation; Coronary Circulation; Disease Models, Animal; Energy Metabolism; Glycogen; Lactic Acid; Myocardial Ischemia; Myocardium; Oxygen Consumption; Swine; Time Factors

H11 kinase is a serine/threonine kinase preferentially expressed in the heart, which participates in cardiac cell growth and also in cytoprotection during ischemia. A cardiac-specific transgenic mouse overexpressing H11 kinase (2- to 7-fold protein increase) has been generated, and is characterized by cardiac hypertrophy with preserved function and protection against irreversible damage during ischemia/reperfusion. In this study, we tested whether H11 kinase also participates in the metabolic adaptation to cardiac hypertrophy and ischemia.. A yeast two-hybrid screen using H11 kinase as a bait in a human heart library revealed a potential interaction with phosphoglucomutase (PGM), the enzyme converting glucose 6-phosphate into glucose 1-phosphate. Interaction between H11 kinase and PGM was confirmed by co-immunoprecipitation. To test the biochemical relevance of this interaction, PGM activity was measured in the heart from wild type and transgenic mice, showing a 20% increase of Vmax in the transgenic group, without change in KM. Glycogen content was increased proportionately to the expression of the transgene, reaching a 40% increase in high-expression transgenic mice (7-fold increase in H11 kinase protein) versus wild type (p < 0.01). Increased incorporation of glucose into glycogen was coupled to a 3-fold increase in the protein expression of the glucose transporter GLUT1 in plasma membrane of transgenic mice (p < 0.01).. H11 kinase promotes the synthesis of glycogen, an essential fuel for the stressed heart in both conditions of overload and ischemia. Therefore, H11 kinase represents an integrative sensor in the cardiac adaptation to stress by coordinating cell growth, survival and metabolism.

Topics: Animals; Blotting, Western; Cell Membrane; Cell Proliferation; Cell Survival; Dose-Response Relationship, Drug; Glucose Transporter Type 1; Glycogen; Heat-Shock Proteins; Humans; Immunoprecipitation; Kinetics; Mice; Mice, Transgenic; Molecular Chaperones; Monosaccharide Transport Proteins; Myocardial Ischemia; Myocardium; Phosphoglucomutase; Protein Serine-Threonine Kinases; Protein Structure, Tertiary; Proteins; Reperfusion Injury; Transgenes; Two-Hybrid System Techniques

Metabolic interventions improve performance during demand-induced ischemia by reducing myocardial lactate production and improving regional systolic function. We tested the hypotheses that 1) stimulation of glycolysis would increase lactate production and improve ventricular wall motion, and 2) the addition of fatty acid oxidation inhibition would reduce lactate production and further improve contractile function. Measurements were made in anesthetized open-chest swine hearts. Three groups, hyperglycemia (HG), HG + oxfenicine (HG + Oxf), and control (CTRL), were treated under aerobic conditions and during demand-induced ischemia. During demand-induced ischemia, HG resulted in greater lactate production and tissue lactate content but had no significant effect on glucose oxidation. HG + Oxf significantly lowered lactate production and increased glucose oxidation compared with both the CTRL and HG groups. Myocardial energy efficiency was greater in the HG and HG + Oxf groups under aerobic conditions but did not change during demand-induced ischemia. Thus enhanced glycolysis resulted in increased energy efficiency under aerobic conditions but significantly enhanced lactate production with no further improvement in function during demand-induced ischemia. Partial inhibition of free fatty acid oxidation in the presence of accelerated glycolysis increased energy efficiency under aerobic conditions and significantly reduced lactate production and enhanced glucose oxidation during demand-induced ischemia.

Topics: Animals; Coronary Circulation; Disease Models, Animal; Fatty Acids, Nonesterified; Glycogen; Glycolysis; Hyperglycemia; Lactic Acid; Myocardial Ischemia; Myocardium; Oxidation-Reduction; Physical Conditioning, Animal; Sus scrofa; Ventricular Function, Left

Compared with normal hearts, those with pathology (hypertrophy) are less tolerant of metabolic stresses such as ischemia. Pharmacologic intervention administered prior to such stress could provide significant protection. This study determined, firstly, whether the pentose sugar ribose, previously shown to improve postischemic recovery of energy stores and function, protects against ischemia when administered as a pretreatment. Secondly, the efficacy of this same pretreatment protocol was determined in hearts with pathology (hypertrophy). For study 1, Sprague-Dawley rats received equal volumes of either vehicle (bolus i.v. saline) or ribose (100 mg/kg) before global myocardial ischemia. In study 2, spontaneously hypertensive rats (SHR; blood pressure approximately 200/130) with myocardial hypertrophy underwent the same treatment protocol and assessments. In vivo left ventricular function was measured and myocardial metabolites and tolerance to ischemia were assessed. In normal hearts, ribose pretreatment significantly elevated the heart's energy stores (glycogen), and delayed the onset of irreversible ischemic injury by 25%. However, in vivo ventricular relaxation was reduced by 41% in the ribose group. In SHR, ribose pretreatment did not produce significant elevations in the heart's energy or improvements in tolerance to global ischemia, but significantly improved ventricular function (maximal rate of pressure rise (+dP/dt(max)), 25%; normalized contractility ((+dP/dt)/P), 13%) despite no change in hemodynamics. Thus, administration of ribose in advance of global myocardial ischemia does provide metabolic benefit in normal hearts. However, in hypertrophied hearts, ribose did not affect ischemic tolerance but improved ventricular function.

Topics: Adenosine Triphosphate; Anaerobic Threshold; Animals; Cardiotonic Agents; Disease Models, Animal; Drug Administration Schedule; Glycogen; Hypertension; Hypertrophy, Left Ventricular; Injections, Intravenous; Male; Myocardial Ischemia; Myocardium; Phosphocreatine; Rats; Rats, Sprague-Dawley; Ribose; Structure-Activity Relationship; Ventricular Function, Left; Ventricular Function, Right

AMP-activated protein kinase (AMPK) is an energy-sensing enzyme that plays a pivotal role in regulating cellular metabolism for sustaining energy homeostasis under stress conditions. Activation of AMPK has been observed in the heart during acute and chronic stresses, but its functional role has not been completely understood because of the lack of effective activators and inhibitors of this kinase in the heart. We generated transgenic mice (TG) with cardiac-specific overexpression of a dominant negative mutant of the AMPK alpha2 catalytic subunit to clarify the functional role of this kinase in myocardial ischemia. In isolated perfused hearts subjected to a 10-min ischemia, AMPK alpha2 activity in wild type (WT) increased substantially (by 4.5-fold), whereas AMPK alpha2 activity in TG was similar to the level of WT at base line. Basal AMPK alpha1 activity was unchanged in TG and increased normally during ischemia. Ischemia stimulated a 2.5-fold increase in 2-deoxyglucose uptake over base line in WT, whereas the inactivation of AMPK alpha2 in TG significantly blunted this response. Using 31P NMR spectroscopy, we found that ATP depletion was accelerated in TG hearts during no-flow ischemia, and these hearts developed left ventricular dysfunction manifested by an early and more rapid increase in left ventricular end-diastolic pressure. The exacerbated ATP depletion could not be attributed to impaired glycolytic ATP synthesis because TG hearts consumed slightly more glycogen during this period of no-flow ischemia. Thus, AMPK alpha2 is necessary for maintaining myocardial energy homeostasis during ischemia. It is likely that the functional role of AMPK in myocardial energy metabolism resides both in energy supply and utilization.

Topics: Adenosine Triphosphate; AMP-Activated Protein Kinases; Animals; Chromatography, High Pressure Liquid; Deoxyglucose; Energy Metabolism; Glucose; Glycogen; Glycolysis; Homeostasis; Magnetic Resonance Spectroscopy; Mice; Mice, Transgenic; Multienzyme Complexes; Myocardial Ischemia; Myocardium; Phosphates; Protein Serine-Threonine Kinases

To study whether ACE inhibition and AT-II receptor blockade modulates myocardial glucose uptake during ischemia and reperfusion.. We developed a method for in vivo sampling of large trans-myocardial tissue samples from beating pig hearts and in vitro measurement of sarcolemmal glucose transport, in a series of experiments in which hearts were exposed to stimuli (glucose-insulin and pacing) known to promote cellular glucose transport. In the subsequent study we compared three experimental groups: (i) ACE inhibition (ACE-I, n = 6): increasing oral doses of benazepril up to 40 mg daily for 3 weeks, (ii) angiotensin II receptor antagonist (AT II-A, n = 7): increasing oral doses of valsartan up to 320 mg for 3 weeks, (iii) control (n = 7). Samples were harvested at baseline, following 20 min of regional ischemia, and following 5 and 15 min of reperfusion. The samples were incubated with 3-O-methylglucose (MeGlu), and cellular MeGlu uptake was measured.. Insulin-glucose, pacing, and ischemia increased cellular MeGlu transport two- to fourfold (p < 0.001). Cellular MeGlu transport was increased in ACE-I and AT II-A animals during reperfusion (p < 0.001), but not at baseline or during ischemia, compared with controls.. Enhanced capacity for glucose transport during reperfusion may be a mechanism underlying the beneficial effects of ACE inhibition and AT II-antagonism in ischemic heart disease.

Topics: Adenosine Triphosphate; Angiotensin II; Angiotensin-Converting Enzyme Inhibitors; Animals; Blood Pressure; Cardiac Pacing, Artificial; Cell Membrane; Disease Models, Animal; Female; Glucose; Glucose Transporter Type 4; Glycogen; Heart Rate; Infusions, Intravenous; Insulin; Male; Models, Cardiovascular; Monosaccharide Transport Proteins; Muscle Proteins; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Renin; Swine; Systole; Time Factors

Myocardial ischaemic preconditioning is characterized by a reduction in the rate of glycolysis. Brief myocardial ischaemia also reduces the glycogen content of the heart. The first objective was to determine whether augmenting glucose oxidation by activation of the pyruvate dehydrogenase complex would prevent the infarct limitation of ischaemic preconditioning. The second part of the study evaluates whether glycogen depletion before ischaemia mimics the infarct-limiting effect of ischaemic preconditioning.. Dichloroacetate (300 + 150 mg kg(-1)), an activator of the pyruvate dehydrogenase complex, was administered intravenously in the anaesthetized open-chest rabbit. All animals underwent 45 min of regional ischaemia and 3 h of reperfusion. Ischaemic preconditiong was elicited by 5 min of coronary occlusion. Control rabbits, those with ischaemic preconditioning with no dichloroacetate, received a saline vehicle. An isolated perfused rabbit heart model was employed to test the second hypothesis. Hearts were depleted of glycogen by perfusing them with a substrate-free buffer. Infarction was assessed by triphenyl tetrazolium chloride and area at risk determined with fluorescent particles.. (a) Pyruvate dehydrogenase complex activation experiments. Treatment with dichloroacetate alone did not alter infarct size (58 +/- 7% control vs. 60 +/- 5% dichloroacetate). Addition of dichloroacetate did not attenuate the infarct-limiting effect of ischaemic preconditioning as evidenced by a similar reduction in infarct size in the ischaemic preconditioning group (22 +/- 5%) and in the ischaemic preconditioning + dichloroacetate group (27 +/- 7%). (b) Glycogen depletion experiments. Compared with control hearts with a normal glycogen content (4.84 +/- 0.15 mg g(-1) wet weight), glycogen depleted and ischaemic preconditioning hearts had reduced glycogen content before ischaemia (2.15 +/- 0.26, 1.62 +/- 0.17 mg g(-1) wet weight, respectively; P < 0.01). Glycogen depletion did not reduce infarct size: 25.0 +/- 4.5% cf. 27.9 +/- 3.4% in the control group. However, ischaemic preconditioning resulted in a significant reduction of infarct size (11.5 +/- 2.3% vs. 27.9 +/- 3.4% control; P < 0.01).. Augmentation of oxidative glycolysis by dichloroacetate in in situ rabbit hearts does not alter the effect of ischaemic preconditioning, and glycogen depletion in the isolated rabbit heart does not influence infarct size after subsequent coronary occlusion.

Topics: Analysis of Variance; Animals; Blood Pressure; Coronary Circulation; Dichloroacetic Acid; Glycogen; Heart Rate; Ischemic Preconditioning, Myocardial; Male; Myocardial Infarction; Myocardial Ischemia; Rabbits; Time Factors

Based on our observations of energy sparing in heat-acclimated (AC) rat hearts, we investigated whether changes in preischemic glycogen level, glycolytic rate, and plasma thyroxine level mediate cardioprotection induced in these hearts during ischemia-reperfusion insults. Control (C) (24 degrees C), AC (34 degrees C, 30 days), acclimated-euthyroid (34 degrees C + 3 ng/ml l-thyroxine), and control hypothyroid (24 degrees C + 0.02% 6-n-propyl-2-thiouracil) groups were studied. Preischemic glycogen was higher in AC than in C hearts [39.0 +/- 8.5 vs. 19.2 +/- 4.2 (SE) micromol glucose/g wet wt; P < 0.0006], and the lactate produced vs. glycogen level during total ischemia ((13)C-NMR spectroscopy) was markedly slower (AC: -0.82x, r = 0.98 vs. C: -4.7x, r = 0.9). Time to onset of ischemic contracture was lengthened, and the fraction of hearts experiencing ischemic contracture was lowered. Pulse pressure recovery was improved in AC compared with C animals before, but not after, absolute sodium iodoacetate-induced glycolysis inhibition. Acclimated-euthyroid hearts exhibited decreased ischemic tolerance, whereas induced hypothyroidism in C improved cardiotolerance. Thus higher preischemic glycogen and slowed glycolysis are associated with hypothyroidism and are likely important mediators of the improved ischemic tolerance exhibited by AC hearts.

Topics: Acclimatization; Animals; Carbon Isotopes; Glucose; Glycogen; Glycolysis; Hot Temperature; Hypothyroidism; Lactic Acid; Magnetic Resonance Spectroscopy; Male; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Physical Endurance; Rats; Thyroxine

We tested the hypothesis that an acute critical limitation in coronary flow reserve could rapidly recapitulate the physiological, molecular, and morphological phenotype of hibernating myocardium. Chronically instrumented swine were subjected to a partial occlusion to produce acute stunning, followed by reperfusion through a critical stenosis. Stenosis severity was adjusted serially so that hyperemic flow was severely reduced yet always higher than the preocclusion resting level. After 24 hours, resting left anterior descending coronary artery (LAD) wall thickening had decreased from 36.3+/-4.0% to 25.5+/-3.7% (P<0.05), whereas resting flow had remained normal (67+/-6 versus 67+/-8 mL/min, respectively). Although peak hyperemic flow exceeded the prestenotic value, resting flow (45+/-10 mL/min) and LAD wall thickening (17.0+/-5.0%) progressively decreased after 2 weeks, when physiological features of hibernating myocardium had developed. Regional reductions in sarcoplasmic reticulum proteins were present in hibernating myocardium but absent in stunned myocardium evaluated after 24 hours. Histological analysis showed an increase in connective tissue along with myolysis (myofibrillar loss per myocyte >10%) and increased glycogen typical of hibernating myocardium in the LAD region (33+/-3% of myocytes from animals with hibernating myocardium versus 15+/-4% of myocytes from sham-instrumented animals, P<0.05). Surprisingly, the frequency of myolysis was similar in normally perfused remote regions from animals with hibernating myocardium (32+/-7%). We conclude that the regional physiological and molecular characteristics of hibernating myocardium develop rapidly after a critical limitation in flow reserve. In contrast, the global nature of myolysis and increased glycogen content dissociate them from the intrinsic adaptations to ischemia. These may be related to chronic elevations in preload but appear unlikely to contribute to chronic contractile dysfunction.

Topics: Adaptation, Physiological; Animals; Calcium-Binding Proteins; Calcium-Transporting ATPases; Calsequestrin; Chronic Disease; Coronary Circulation; Coronary Stenosis; Disease Models, Animal; Disease Progression; Glycogen; Hemodynamics; Ischemic Preconditioning, Myocardial; Microspheres; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardial Stunning; Myocardium; Myofibrils; RNA, Messenger; Sarcoplasmic Reticulum; Sarcoplasmic Reticulum Calcium-Transporting ATPases; Swine

Repetitive myocardial ischemia increases glucose uptake, but the effect on glycogen is unclear. Thirteen swine instrumented with a hydraulic occluder on the circumflex (Cx) artery underwent 10-min occlusions twice per day for 4 days. After 24 h postfinal ischemia and in the fasted state, echocardiogram and positron emission tomography imaging for blood flow ([(13)N]-ammonia) and 2-[(18)F]fluoro-2-deoxy-D-glucose (FDG) uptake were obtained. Tissue was then collected for ATP, creatine phosphate (CP), glycogen, and glucose transporter-4 content, and hexokinase activity. After reperfusion, regional function and CP-to-ATP ratios in the Cx and remote regions were similar. Despite the absence of stunning, the Cx region demonstrated higher glycogen levels (33 +/- 11 vs. 24 +/- 11 micromol/g; P < 0.05), and this increase correlated well with the increase in FDG uptake (r(2) = 0.78; P < 0.01). Hexokinase activity was also increased relative to remote regions (0.62 +/- 0.29 vs. 0.37 +/- 0.19 IU/g; P < 0.05), with no difference in GLUT-4 content. In summary, 24 h after repetitive ischemia, glucose uptake and glycogen levels are increased at a time that functional and bioenergetic markers of stunning have recovered. The significant correlation between glycogen content and FDG accumulation in the postischemic region suggests that increased rates of glucose transport and/or phosphorylation are linked to increased glycogen levels in hearts subjected to repetitive bouts of ischemia.

Topics: Adenosine Triphosphate; Ammonia; Animals; Biological Transport; Coronary Circulation; Echocardiography; Fluorodeoxyglucose F18; Glucose; Glucose Transporter Type 4; Glycogen; Heart; Hemodynamics; Hexokinase; Monosaccharide Transport Proteins; Muscle Proteins; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Nitrogen Isotopes; Phosphocreatine; Swine; Tomography, Emission-Computed

PKC epsilon is a PKC isoform that translocates during preconditioning and may mediate cardioprotection. To investigate whether PKC epsilon activation is cardioprotective, Langendorff-perfused hearts from wild-type (WT) mice and from mice expressing constitutively active mutant PKC epsilon were subjected to 20 min ischemia and 40 min reperfusion while(31)P NMR spectra were acquired. Pre-ischemic glycogen levels were similar in WT and PKC epsilon hearts. During ischemia, ATP fell less in PKC epsilon than in WT hearts. Ischemic intracellular pH, however, was similar in WT and PKC epsilon hearts. During reperfusion, recovery of contractile function and ATP were greater in PKC epsilon than WT hearts. In conclusion, expression of activated PKC epsilon protected hearts from post-ischemic energetic and contractile dysfunction, consistent with the proposed cardioprotective role of PKC epsilon. Protection occurred in the PKC epsilon hearts without attenuation of ischemic H(+) production, implying that, at least in this ischemic model, reduced acidification during ischemia is not necessary for cardioprotection.

Topics: Adenosine Triphosphate; Animals; Enzyme Activation; Glycogen; Heart Rate; Hydrogen-Ion Concentration; Isoenzymes; Mice; Mice, Transgenic; Myocardial Contraction; Myocardial Ischemia; Nuclear Magnetic Resonance, Biomolecular; Phosphates; Protein Kinase C; Protein Kinase C-epsilon; Protons; Time Factors

During stress, patients with coronary artery disease frequently fail to increase coronary flow and myocardial oxygen consumption (MVO(2)) in response to a greater demand for oxygen, resulting in "demand-induced" ischemia. We tested the hypothesis that dobutamine infusion with flow restriction stimulates nonoxidative glycolysis without a change in MVO(2) or fatty acid uptake. Measurements were made in the anterior wall of anesthetized open-chest swine hearts (n = 7). The left anterior descending (LAD) coronary artery flow was controlled via an extracorporeal perfusion circuit, and substrate uptake and oxidation were measured with radiotracers. Demand-induced ischemia was produced with intravenous dobutamine (15 microg x kg(-1) x min(-1)) and 20% reduction in LAD flow for 20 min. Despite no change in MVO(2), there was a switch from lactate uptake (5.9 +/- 3.1) to production (74.5 +/- 16.3 micromol/min), glycogen depletion (66%), and increased glucose uptake (105%), but no change in anterior wall power or the index of anterior wall energy efficiency. There was no change in the rate of tracer-measured fatty acid uptake; however, exogenous fatty acid oxidation decreased by 71%. Thus demand-induced ischemia stimulated nonoxidative glycolysis and lactate production, but did not effect fatty acid uptake despite a fall in exogenous fatty acid oxidation.

Topics: Adenosine Diphosphate; Adenosine Triphosphate; Animals; Blood Flow Velocity; Coronary Vessels; Dobutamine; Energy Metabolism; Fatty Acids; Fatty Acids, Nonesterified; Glycogen; Glycolysis; Hemodynamics; Kinetics; Lactic Acid; Myocardial Ischemia; Myocardium; Oxidation-Reduction; Oxygen Consumption; Swine; Triglycerides; Ventricular Function, Left

A moderate reduction in coronary blood flow results in decreased myocardial oxygen consumption, accelerated glycolysis, decreased pyruvate oxidation, and lactate accumulation. To quantitatively understand cardiac metabolism during ischemia, we have developed a mechanistic, mathematical model based on biochemical mass balances and reaction kinetics in cardiac cells. By numerical solution of model equations, computer simulations showed the dynamic responses in glucose, fatty acid, glucose-6-phosphate, glycogen, triglyceride, pyruvate, lactate, acetyl-CoA, and free-CoA as well as CO2, O2, phosphocreatine/creatine, nicotinamide adenine dinucleotide (reduced form)/nicotinamide adenine dinucleotide (oxidized form) (NADH/NAD+), and adenosine diphosphate/adenosine triphosphate (ADP/ATP). When myocardial ischemia was simulated by a 60% reduction in coronary blood flow, the model generated myocardial concentrations, uptakes, and fluxes that were consistent with experimental data from in vivo pig studies. After 60 min of ischemia the concentrations of glycogen, phosphocreatine, and ATP were decreased by 60%, 75%, and 50%, respectively. With the onset of ischemia, myocardial lactate concentration increased and the myocardium switched from net consumer to net producer of lactate. Our model predicted a rapid 13-fold increase in NADH/NAD+, but only a twofold increase in the ratio of acetyl-CoA to free-CoA. These findings are consistent with the concept that pyruvate oxidation is inhibited during ischemia partially by the rise in NADH/NAD+.

Topics: Adenosine Diphosphate; Adenosine Triphosphate; Animals; Blood Glucose; Computer Simulation; Coronary Circulation; Energy Metabolism; Fatty Acids; Glycogen; Heart; Humans; Lactic Acid; Models, Cardiovascular; Myocardial Ischemia; Myocardium; NAD; Oxygen; Sensitivity and Specificity; Swine

Rhythm disorders are common complications in diabetic patients, due to their enhanced sensitivity to ischaemia. However, experimental studies are inconsistent, and both higher and lower vulnerability to injury has been reported. Our objectives were to compare susceptibility to ventricular arrhythmias in rats with prolonged duration of diabetes induced by streptozotocin (45 mg/kg, i.v.), utilising two different models. Following 8 weeks, either anaesthetised open-chest rats in vivo or isolated Langendorff-perfused hearts were subjected to 30 min regional zero-flow ischaemia induced by occlusion of LAD coronary artery. In addition, cardiac glycogenolysis and lactate production were measured. In open-chest rats, 90 % of the controls exhibited ventricular tachycardia (VT) which represented 55.4 % of total arrhythmias, whereby only 19.9 % of arrhythmias occurred as VT in 44 % of the diabetic rats (P < 0.05 vs controls). Duration of VT and ventricular fibrillation (VF) was reduced from 35.5 +/- 11.1 and 224.8 +/- 153.9 s in the controls to 4.8 +/- 2.5 and 2.2 +/- 0.2 s in the diabetics, respectively (P < 0.05). Accordingly, severity of arrhythmias (arrhythmia score, AS) was also lower in the diabetics (2.0 +/- 0.38 vs 3.3 +/- 0.3 in the controls; P < 0.05). In the isolated hearts, high incidence of VF was decreased in the diabetic hearts, and although VT occurred in almost all of the diabetic hearts, the duration of VT and VF was substantially shorter (61.5 +/- 14.5 and 5.5 +/- 0.5 s vs 221.5 +/- 37 and 398.5 +/- 55 s in the controls, respectively; P < 0.05). AS was reduced to 2.9 +/- 0.12 from 4.1 +/- 0.3 in the controls (P < 0.05). Postischaemic accumulation of lactate was lower in the diabetic than in the non-diabetic myocardium (20.4 +/- 1.9 vs 29.5 +/- 2.9 micromol/l/g w.wt.; P < 0.05). These results suggest that rat hearts with chronic diabetes, despite some differences in the arrhythmia profiles between the in vivo model and isolated heart preparation, are less sensitive to ischaemic injury and exhibit lower susceptibility to ventricular arrhythmias and reduced accumulation ofglycolytic metabolites.

Topics: Animals; Blood Glucose; Coronary Disease; Diabetes Mellitus, Experimental; Glycogen; In Vitro Techniques; Lactic Acid; Male; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Rats; Rats, Wistar; Tachycardia, Ventricular

Acute ischemia comes with two phases of life-threatening arrhythmias, early (within 10 minutes, 1A) and late (after about 15 minutes, 1B). The mechanism of the latter is unknown and in this paper, we test the hypothesis that a phase of intermediate coupling between surviving epicardium and inexcitable midmyocardium underlies 1B arrhythmias.. Pig hearts (n=26) were retrogradely perfused with a blood Tyrode's mixture. The left anterior descending artery was occluded. We investigated (1) inducibility of ventricular fibrillation (VF) with programmed stimulation, (2) tissue impedance (Rt) heterogeneity within the ischemic zone, (3) multiple subepicardial and midmyocardial electrograms, (4) subepicardial lactate dehydrogenase (LDH) and glycogen content.. In nine of ten hearts, one--three premature stimuli caused VF between 14 and 53 min of ischemia. This typically happened when the Rt of the ischemic zone had increased up to 40% of its final value. More uncoupling terminated the period of VF inducibility. The excitability of the surviving subepicardial layer was depressed during the same period with partial uncoupling, but recovered when the uncoupling from the midmyocardium had progressed further.. We show that 1B-VF can be induced within a distinct time window and coincides with a distinct range of Rt rise. Subepicardium is electrically depressed, presumably through coupling with midmyocardium, complete uncoupling causes subepicardial recovery and terminates the substrate for 1B-VF. Hence, we suggest that the substrate for 1B-VF consists of intermediate coupling of subepicardium and midmyocardium.

Topics: Animals; Cell Communication; Death, Sudden, Cardiac; Electrocardiography; Female; Glycogen; Heart Conduction System; L-Lactate Dehydrogenase; Male; Myocardial Ischemia; Myocardium; Organ Culture Techniques; Swine; Tachycardia, Ventricular

The ischemic heart is dependent on glycolysis for ATP generation, and therapies that increase glucose utilization during ischemia improve survival. Myocardial ischemia results in the translocation of the glucose transporter proteins GLUT1 and GLUT4 to the sarcolemma. The increased glucose entry via these transporters contributes to enhanced glycolysis during ischemia.. To determine the role of GLUT4 in mediating increased glycolytic flux during ischemia, hearts from mice with cardiac-selective GLUT4 deficiency (G4H-/-) were subjected to global low-flow ischemia. During normal perfusion, hearts from fed G4H-/- mice showed increased GLUT1-mediated glucose uptake, higher concentrations of glycogen and phosphocreatine, but delayed recovery after ischemia. When these compensatory changes were eliminated by a 20-hour fast, G4H-/- hearts exhibited depressed glucose utilization during ischemia and developed profound and irreversible systolic and diastolic dysfunction associated with accelerated ATP depletion during ischemia and diminished regeneration of high-energy phosphate compounds on reperfusion.. GLUT4 is an important mediator of enhanced glycolysis during ischemia and represents an important protective mechanism against ischemic injury.

Topics: Adenosine Triphosphate; Animals; Blood Pressure; Creatine; Fasting; Glucose; Glucose Transporter Type 1; Glucose Transporter Type 4; Glycogen; Glycolysis; Heart; Heart Rate; In Vitro Techniques; Insulin; Lactic Acid; Magnetic Resonance Spectroscopy; Mice; Monosaccharide Transport Proteins; Muscle Proteins; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Phosphates; Phosphocreatine

Topics: Analysis of Variance; Animals; Body Weight; Diabetes Mellitus, Experimental; Disease Models, Animal; Glycogen; Heart; Hemodynamics; Humans; Isoproterenol; L-Lactate Dehydrogenase; Lactates; Male; Myocardial Infarction; Myocardial Ischemia; Myocardium; Necrosis; Organ Size; Rats; Rats, Wistar

We have previously demonstrated that perfusion of isolated hearts with high concentrations of glucose results in increased glycolysis during ischemia, diminished ischemic injury, and improved functional recovery with reperfusion.. To evaluate a possible mechanism by which glucose conferred this protection. We examined the hypothesis that increased exogenous glucose concentrations results in increased concentrations of fructose-2,6-bisphosphate, a potent activator of phosphofructokinase-1, and thus increases glycolysis.. Perfused rabbit hearts were subjected to 60 min of low-flow ischemia. Control hearts were perfused with buffer containing 0.4 mmol/l palmitate, 5 mmol/l glucose, and 70 mU/l insulin, and treated hearts were perfused with buffer containing 0.4 mmol/l palmitate, 15 mmol/l glucose and 210 mU/l insulin.. Ischemic contracture was attenuated by perfusion of high concentrations of glucose (high glucose) (P < 0.05 compared with control). Glucose uptake and lactate production were greater in hearts perfused with high glucose, as was the ATP concentration at the end of ischemia (P < 0.05 compared with controls). Exogenous glucose uptake and lactate production correlated well with fructose-2,6-bisphosphate content (P = 0.007).. Enhancement of glycolysis in hearts perfused with high glucose may be the result of stimulation of phosphofructokinase-1 by fructose-2,6-bisphosphate. Accordingly, this may serve as an important mechanism by which cardioprotection may be achieved.

Topics: Adenosine Triphosphate; Animals; Glucose; Glycogen; Glycolysis; Hemodynamics; Lactic Acid; Myocardial Ischemia; Phosphofructokinase-1; Phosphofructokinase-2; Phosphoric Monoester Hydrolases; Rabbits

Metabolic disorders due to changes in cytosolic glucose utilisation are suspected to be involved in the increased sensitivity of the aged myocardium to ischemia. This study presents the first direct measurement of glucose utilisation in hearts from senescent rats during low-flow ischemia under different conditions of substrate delivery and glycogen stores. Isolated hearts from young adult (4-months-old) and senescent (24-months-old) rats were subjected to 30 min coronary flow restriction (residual flow rate=2% of control flows). Experiments were performed using glucose-free or glucose-enriched (11 mmol/L) perfusion media. The effects of increased glycogen stores were assessed after 24-h fasting in both age groups. Ischemic contracture was measured via a left-ventricular balloon. Ageing increased ischemic contracture under both conditions of substrate delivery in fed rat hearts. The increase in ischemic tolerance induced by fasting in senescent rat hearts was less than that seen in young rat hearts. Moreover, fasting decreased glucose utilisation in hearts from young rats, an effect which was not found in hearts from old rats. Furthermore, myocardial glycogen utilisation was increased in all groups of aged rats compared with that of young adults, particularly under fasting conditions. It is concluded that fasting is less detrimental to the aged myocardium during low-flow ischemia than to the young myocardium because it does not further reduce exogenous glucose utilisation, and it stimulates glycogen consumption. Moreover, a reduction in exogenous glucose utilisation, which is only partly compensated for by increased glycogenolytic flux could be, at least in part, responsible for the increased ischemic contracture in hearts from old fed rats. Finally, our glucose-free experiments suggest that residual oxidative phosphorylation during low-flow ischemia might be less relevant in hearts from senescent rats than in those from young adults.

Topics: Aging; Animals; Coronary Circulation; Energy Metabolism; Fasting; Glucose; Glycogen; Glycolysis; In Vitro Techniques; Lactic Acid; Male; Myocardial Ischemia; Myocardium; Oxidative Phosphorylation; Perfusion; Rats; Rats, Wistar

This study was undertaken to define the contributions of left ventricular hypertrophy (LVH) and increased adrenergic activity to the acceleration of ischemic contracture (IC) that occurs in chronic hyperthyroid rat heart. Acute and chronic hyperthyroidism (THYR) were induced by thyroxine administration for 2 and 14 days, respectively, and normal animals (NORM) served as controls. Isolated hearts were perfused in a Langendorff mode. NORM alpha acute, n = 6; THYR alpha acute, n = 8; and THYR alpha, n = 13; and NORM alpha, n = 13 were subjected to 20-min zero-flow global ischemia (I) and 45-min reperfusion (R). Additional THYR and NORM hearts treated with propranolol (prop) were subjected to 30-min I; THYR beta prop, n = 6 and NORM beta prop, n = 8, and THYR beta, n = 6, NORM beta, n = 8 served as controls. Acceleration of IC was measured by the time to peak contracture (Tmax). Left ventricular hypertrophy (LVH) was assessed by the ratio of left ventricular weight in milligrams (LVW) to animal body weight (BW) in grams. Cardiac hypertrophy developed in chronic but not acute hyperthyroidism. Propranolol reduced the extent of LVH. Contracture occurred earlier in chronic than in acute hyperthyroid and normal hearts. Propranolol did not alter contracture. In conclusion, IC is accelerated by thyroxine administration, and this is probably not due to LVH or increased beta-adrenergic activity. Propranolol diminishes LVH in hyperthyroidism.

Topics: Adrenergic beta-Antagonists; Animals; Glycogen; Hyperthyroidism; Hypertrophy, Left Ventricular; Male; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Propranolol; Rats; Rats, Wistar

Diabetic hearts are suggested to exhibit either increased or lower sensitivity to ischemia. Detrimental effects of prolonged ischemia can be attenuated by preconditioning, however, relatively little is known about its effects in the diseased myocardium. This study was designed to test the susceptibility to ischemia-induced arrhythmias and the effect of preconditioning in the diabetic heart. Rats were made diabetic with streptozotocin (45 mg/kg, i.v.). After 1 week, isolated Langendorff-perfused hearts were subjected to 30 min occlusion of LAD coronary artery without or with preceding preconditioning induced by one cycle of 5 min ischemia and 10 min reperfusion. Glycogen and lactate contents were estimated in the preconditioned and non-preconditioned hearts before and after ischemia. Diabetic hearts were more resistant to ischemia-induced arrhythmias: incidence of ventricular tachycardia (VT) decreased to 42% and only transient ventricular fibrillation (VF) occurred in 17% of the hearts as compared to the non-diabetic controls (VT 100% and VF 70% including sustained VF 36%; p < 0.05). Preconditioning effectively suppressed the incidence and severity of arrhythmias (VT 33%, VF 0%) in the normal hearts. However, this intervention did not confer any additional protection in the diabetic hearts. Despite higher glycogen content in the diabetic myocardium and greater glycogenolysis during ischemia, production of lactate in these hearts was significantly lower than in the controls. Preconditioning caused a substantial decrease in the accumulation of lactate in the normal hearts, whereby in the diabetic hearts, this intervention did not cause any further reduction in the level of lactate. In conclusion, diabetic rat hearts exhibit lower susceptibility to ischemic injury and show no additional response to preconditioning. Reduced production of glycolytic metabolites during ischemia can account for the enhanced resistance of diabetic hearts to ischemia as well as for the lack of further protection by preconditioning.

Topics: Animals; Arrhythmias, Cardiac; Blood Glucose; Diabetes Mellitus, Experimental; Glycogen; Heart; Heart Rate; In Vitro Techniques; Ischemic Preconditioning, Myocardial; Lactic Acid; Male; Myocardial Ischemia; Myocardium; Rats; Rats, Wistar

Using (13)C NMR, we tested the hypothesis that protection by preconditioning is associated with reduced glycogenolysis during ischemia. Preconditioned rat hearts showed improved postischemic function and reduced ischemic damage relative to ischemic controls after 30 min stop-flow ischemia and 30 min reperfusion (contractility: 30+/-10 vs. 2+/-2%; creatine kinase release: 41+/-4 vs. 83+/-15 U/g; both P<0.05). Preconditioning decreased preischemic [(13)C]glycogen by 24% (a 10% decrease in total glycogen), and delayed ischemic [(13)C]glycogen consumption by 5-10 min, reducing ischemic glycogenolysis without changing acidosis relative to controls. Upon reperfusion, glycogen synthesis resumed only after preconditioning. Glutamate (13)C-isotopomer analysis showed recovery of Krebs cycle activity with higher anaplerosis than before ischemia (23+/-4 vs. 11+/-3%, P<0.05), but in controls reperfusion failed to restore flux. Compared to control, preconditioning before 20 min ischemia increased contractility (86+/-10 vs. 29+/-14%, P<0.05) and restored preischemic anaplerosis (13+/-3 vs. 39+/-9%, P<0.05). Preconditioning is associated with reduced glycogenolysis early during ischemia. However, protection does not rely on major variations in intracellular pH, as proposed earlier. Our isotopomer data suggest that preconditioning accelerates metabolic and functional recovery during reperfusion by more efficient/active replenishment of the depleted Krebs cycle.

Topics: Alanine; Animals; Citric Acid Cycle; Glutamic Acid; Glycogen; Heart; Hydrogen-Ion Concentration; In Vitro Techniques; Lactic Acid; Magnetic Resonance Spectroscopy; Male; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Rats; Rats, Sprague-Dawley; Tissue Extracts

We examined whether pharmacological inhibition of glycogenolysis by N-methyl-1-deoxynojirimycin (MOR-14), a new compound which reduces the glycogenolytic rate by inhibiting the alpha-1,6-glucosidase activity of the glycogen-debranching enzyme, can protect the heart against postischemic left ventricular dysfunction. The hearts of male Sprague-Dawley rats were excised, and perfused on a Langendorff apparatus with Krebs-Henseleit solution with a gas mixture of 95% O2 and 5% CO2. The hearts were paced at 320 beats/min except during the ischemia. Left ventricular developed pressure (LVDP, mmHg), +/-dP/dt (mmHg/s), and coronary flow (ml/min) were continuously monitored. All hearts were perfused for a total of 120 min including a 30-min preischemic period followed by a 30-min episode of global ischemia and 60 min reperfusion. with or without 0.5 or 2 mM of MOR-14 during the 30-min preischemic period or the first 30 min of reperfusion. In another series of experiments, the myocardial content of glycogen and lactate was measured during the 30-min episode of ischemia in groups treated with and without 2mM of MOR-14. Preischemic but not postischemic treatment with MOR-14 significantly improved LVDP and +/-dP/dt without altering coronary flow during reperfusion in a dose-dependent manner. MOR-14 significantly preserved the glycogen content and significantly attenuated the lactate accumulation during the 30-min episode of ischemia. Preischemic treatment with MOR-14 is protective against postischemic left ventricular dysfunction through the inhibition of glycogenolysis in the isolated rat heart.

Topics: 1-Deoxynojirimycin; Adenosine Triphosphate; Animals; Blood Flow Velocity; Disease Models, Animal; Enzyme Inhibitors; Glycogen; Glycoside Hydrolase Inhibitors; Heart; Lactic Acid; Male; Myocardial Ischemia; Myocardial Reperfusion; Rats; Rats, Sprague-Dawley; Time Factors; Ventricular Dysfunction, Left

In order to highlight severity of myocardial injury during the course of global ischemia and reperfusion, cytochemistry of glycogen and succinate dehydrogenase (SDH) as well as hematoxylin and eosin staining (H & E) and electron microscopy were observed in canine myocardium. Seven mongrel dogs were selected for reperfusion injury after global ischemia in this study. Myocardial biopsies were taken from the anterior wall of the left ventricle (a) after cardiopulmonary bypass (the first biopsy); (b) at the end of the aortic crossclamp (the second biopsy); and (c) 30 minutes after crossclamp removal (the third biopsy). All biopsies were cytochemically assessed, and the latter two, for electron microscopic studies. The averages of myocardial necrotic rate and surface to volume ratio of myocardial mitochondria were calculated under electron microscopy and in electron microscopic slices, respectively. Myofibrillae were of normal morphology in the first biopsy; in wave-shape and partly vacuolated, with large and deformed nuclei in the second one; and in wave-shape and severely vacuolated in the third one, in H & E. Glycogen granules were variously stained in moderate, weak and intensive positive reactions in the three biopsies respectively in glycogen staining. SDH was stained in intensive, weak, and moderate positive reactions in three, respectively. By electron microscopy, Z bands twisted severely, and local dissolution of cristae and matrix occurred in a minority of the mitochondria in the second biopsy; and majority of the Z bands in necrotic region had disappeared, the myofibrillae were obscure and patchily dissolved. Clustered and deformed mitochondria could be found in the third biopsy. Significant difference could be noted between the averages of the second and third biopsies (14.88 +/- 3.09% vs. 60.25 +/- 8.55%, p < 0.001). The surface to volume ratio of the ischemic mitochondria was much bigger than that of the reperfused (3.95 +/- 1.09 micron-1 vs. 2.77 +/- 0.93 micron-1, p = 0.041). Myocardial injury was more severe in reperfusion than in ischemia myocardium. There were correlations between histobiochemical and ultrastructural alterations in damaged canine myocardium.

Topics: Animals; Dogs; Glycogen; Microscopy, Electron; Mitochondria, Heart; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Succinate Dehydrogenase

The authors tested the hypothesis that ischemia stimulates glucose uptake in rat heart independent of the insulin signaling pathway and independent of endogenous catecholamines. Isolated working rat hearts were perfused with Krebs-Henseleit buffer containing [2-3H]glucose (5 mmol/l, 0.05 muCi/ml) and Na-oleate (0.4 mmol/l) with or without the phosphatidylinositol 3-kinase inhibitor wortmannin (3 mumol/l). Insulin (1 mU/ml) was added to the perfusate in the middle of the experiments or the hearts were subjected to 30 min of low-flow ischemia (75% reduction in coronary flow) followed by 15 min of reperfusion. In a separate group, hearts were subjected to ischemia and reperfusion in the presence of propranolol (10 mumol/l) plus phentolamine (10 mumol/l). Cardiac power was stable but decreased (-75%) during ischemia. Both insulin and ischemia increased glucose uptake (P < 0.01). Glucose uptake returned to pre-ischemic values during reperfusion. Wortmannin completely inhibited insulin-stimulated glucose uptake and glycogen synthesis, but did not affect the ischemia-stimulated glucose uptake or glycogen resynthesis during reperfusion. Full adrenergic blockade did not abolish the ischemia-stimulated glucose uptake. The authors conclude that: (1) insulin and ischemia increase glucose uptake through different mechanisms; (2) ischemia-stimulated glucose uptake is not catecholamine mediated: and (3) glycogen resynthesis during reperfusion is independent of PI3-K.

Topics: Animals; Catecholamines; Deoxyglucose; Glucose; Glycogen; Heart; Male; Myocardial Ischemia; Myocardium; Rats; Rats, Sprague-Dawley

We undertook this study to determine if the metabolism of exogenous glucose and glycogen in hypertrophied hearts differed from that in normal hearts during severe ischemia. Thus, rates of glycolysis (3H2O production) and oxidation (14CO2 production) from exogenous glucose and glycogen were measured in isolated working control (n = 13) and hypertrophied (n = 12) hearts from sham-operated and aortic-banded rats during 40 min of severe low-flow ischemia. Hearts, in which glycogen was prelabelled with [5-3H]- or [14C]-glucose, were paced and perfused with Krebs-Henseleit solution containing 1.2 mM palmitate, 5.5 mM [5-3H]- or [14C]-glucose (different from the isotope used to label glycogen), 0.5 mM lactate and 100 microU/ml insulin during ischemia. Rates of glycolysis from exogenous glucose (3301 +/- 122 v 2467 +/- 167 nmol/min/g dry wt, mean +/- S.E.M., P < 0.05) and glucose from glycogen (808 +/- 27 v 725 +/- 21 nmol/min/g dry wt, P < 0.05) were accelerated in hypertrophied hearts compared to control hearts. However, rates of oxidation of exogenous glucose and glucose from glycogen were not significantly different between the two groups. As observed in normoxic non-ischemic hearts, glucose from glycogen was preferentially oxidized compared to exogenous glucose. Additionally, rates of glycogen synthesis (167 +/- 7 v 140 +/- 9 nmol/min/g dry wt, P < 0.05) were increased in hypertrophied hearts compared to control hearts during severe low-flow ischemia indicating that glycogen turnover (i.e. simultaneous synthesis and degradation) was accelerated in the hypertrophied heart. Thus, we demonstrate that glucose utilization and glycogen turnover are accelerated in the hypertrophied heart during severe low-flow ischemia as compared to the normal heart.

Topics: Animals; Body Weight; Cardiomegaly; Glucose; Glycogen; Male; Myocardial Ischemia; Rats; Rats, Sprague-Dawley; Time Factors

The objective of this study was to determine whether hypoxic preconditioning (HP) could lessen the myocardial increase in [Na+]i, thus protecting the aging myocardium against ischemia.. A decrease in ischemic tolerance with aging is associated with an accelerated increase in [Na+]i during ischemia. Ischemic preconditioning fails to protect the middle-aged and senescent myocardium against ischemia.. Isolated hearts of young adult (12-week-old), middle-aged (50-week-old) and senescent (100-week-old) Fischer 344 rats were subjected to 25 min of ischemia with or without HP followed by 30 min of reperfusion. Left ventricular (LV) function, myocardial energy metabolites and [Na+]i were measured.. In the older groups, the recovery of LV function and high-energy phosphates (HEPs) was lower with an increased release of creatine kinase (CK) during reperfusion than in the young group. The increased [Na+]i at the end of ischemia was greater in the former groups than in the young group. HP decreased myocardial glycogen and lessened the increased [Na+]i in the young group, resulting in an improved recovery of LV function and HEPs, as well as decreased CK release. However, the levels of glycogen before HP in the older groups were higher than in the young group and its levels after HP were similar to that before HP in the young group. HP did not affect the [Na+]i, exacerbated CK release and inhibited the recovery of LV function and HEPs in the older groups.. HP failed to lessen the increased [Na+]i or to protect the aging hearts, probably due to the preexistence of increased glycogen level.

Topics: Aging; Analysis of Variance; Animals; Calcium; Chi-Square Distribution; Creatine Kinase; Glycogen; Homeostasis; Ischemic Preconditioning, Myocardial; Male; Myocardial Ischemia; Myocardium; Perfusion; Phosphates; Rats; Rats, Inbred F344; Sodium; Ventricular Dysfunction, Left

1. Cardioprotection by adenosine A1 receptor activation limits infarct size and improves post-ischaemic mechanical function. The mechanisms responsible are unclear but may involve alterations in myocardial glucose metabolism. 2. Since glycogen is an important source of glucose during ischaemia, we examined the effects of N6-cyclohexyladenosine (CHA), an A1 receptor agonist, on glycogen and glucose metabolism during ischaemia as well as reperfusion. 3. Isolated working rat hearts were perfused with Krebs-Henseleit solution containing dual-labelled 5-3H and 14C glucose and palmitate as energy substrates. Rates of glycolysis and glucose oxidation were measured directly from the production of 3H2O and 14CO2. Glycogen turnover was measured from the rate of change of [5-3H and 14C]glucosyl units in total myocardial glycogen. 4. Following low-flow (0.5 ml min-1) ischaemia (60 min) and reperfusion (30 min), left ventricular minute work (LV work) recovered to 22% of pre-ischaemic values. CHA (0.5 microM) improved the recovery of LV work 2 fold. 5. CHA altered glycogen turnover in post-ischaemic hearts by stimulating glycogen synthesis while having no effects on glycogen degradation. CHA also partially inhibited glycolysis. These changes accelerated the recovery of glycogen in CHA-treated hearts and reduced proton production. 6. During ischaemia, CHA had no measurable effect on glycogen turnover or glucose metabolism. Glycogen phosphorylase activity, which was elevated after ischaemia, was inhibited by CHA, possibly in response to CHA-induced inhibition of AMP-activated protein kinase activity. 7. These results indicate that CHA-induced cardioprotection is associated with alterations of glycogen turnover during reperfusion as well as improved metabolic coupling of glycolysis to glucose oxidation.

Topics: Adenosine; Adenylate Kinase; Aerobiosis; Animals; Glucose; Glycogen; Glycogen Synthase; Glycolysis; Heart Ventricles; In Vitro Techniques; Kinetics; Male; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Palmitic Acid; Phosphates; Phosphorylases; Protons; Purinergic P1 Receptor Agonists; Rats; Rats, Sprague-Dawley; Receptors, Purinergic P1; Ventricular Function

Fasting protects the ischemic heart from injury and infarction. Previous studies have shown that hearts from fasted animals have greater glycogen utilization and a lower cytosolic redox state (NADH/NAD+) during global ischemia. While the mechanisms of increased glycogen utilization in fasted animals have not been elucidated, animals that hibernate or are tolerant of anoxia are known to increase the tissue content of the active form of glycogen phosphorylase, phosphorylase a. Therefore, this study was designed to (a) determine whether hearts from fasted animals have increased activity of glycogen phosphorylase during ischemia and (b) define those mechanisms responsible for this increase.. Hearts isolated from either fed or fasted (24 h) rats were perfused and freeze-clamped at baseline, and after 1 and 10 min of ischemia, for measurement of phosphorylase activity, phosphorylase kinase activity, and glucose-6-phosphate concentrations.. Fasting increased the phosphorylase a/b ratio under both baseline and ischemic conditions. This increase was not accompanied by an increase in the activity of phosphorylase kinase, either with maximal [Ca2+] or under physiologic [Ca2+]. Glucose 6-phosphate concentrations were lower in hearts from fasted animals under baseline, but not ischemic, conditions.. Fasting enhances glycogen utilization during ischemia by increasing the active form of glycogen phosphorylase. This increase is not due to a change in phosphorylation by phosphorylase kinase nor end-product inhibition by G-6P. While the precise mechanism of increased glycogen phosphorylase activity in fasted animals is not clear, one likely explanation may be the lower cytosolic redox state demonstrated in the myocardium of fasted animals.

Topics: Analysis of Variance; Animals; Fasting; Glucose-6-Phosphate; Glycogen; Male; Myocardial Ischemia; Myocardium; Perfusion; Phosphorylase Kinase; Phosphorylases; Rats; Rats, Sprague-Dawley

The present study was carried out to investigate the effect of orotic acid on ischaemic/reperfused myocardial function and cardiac glycogen content in isolated working rat hearts. In a preliminary series of studies, hearts isolated from male Wistar rats (300-350 g) were perfused with oxygenated Krebs-Henseleit buffer containing cumulative concentrations of orotic acid from 0.01 to 10.00 mg l-1. In the concentration range of 0.01-0.10 mg l-1, orotic acid significantly improved left ventricular function. Therefore, in the second series of studies, rats were treated intravenously with 0.01 mg kg-1 orotic acid for 4 days. Hearts were then isolated and subjected to 30 min of global no-flow ischaemia followed by 10 min of reperfusion. Orotic acid treatment significantly improved post-ischaemic myocardial function and increased pre-ischaemic and post-ischaemic glycogen content of the heart. We conclude that orotic acid improves ischaemic/reperfused cardiac performance and this effect may be based on the elevation of myocardial glycogen content.

Topics: Animals; Glycogen; Heart; Heart Rate; Male; Myocardial Ischemia; Myocardial Reperfusion; Myocardial Reperfusion Injury; Myocardium; Orotic Acid; Rats; Rats, Wistar

Both hypothermia and ischemic preconditioning are known to provide tolerance to myocardial ischemia and reperfusion. The aim of this study was to determine whether hypothermia during the ischemic preconditioning period attenuates the protective effect of ischemic preconditioning.. Experiments were performed in buffer-perfused isolated rabbit hearts. All hearts underwent 45 minutes of regional ischemia, followed by 2 hours of reperfusion. Ischemic preconditioning was elicited by either one or four periods of 5 minutes of regional ischemia. Hypothermia (25 degrees C) was induced beginning either 20 or 50 minutes before the 45-minute period of regional ischemia; normothermia (38 degrees C) was restored 10 minutes before the 45-minute period of regional ischemia. Except for the hypothermic periods noted, hearts were maintained at 38 degrees C.. Normothermic ischemic preconditioning with either one or four cycles of 5 minutes of coronary occlusion resulted in a profound reduction of infarct size (58% reduction with one cycle, p < 0.05; 95% reduction with four cycles, p < 0.01). Hypothermic ischemic preconditioning with one cycle of 5-minute coronary occlusion resulted in no reduction of infarct size but hypothermic ischemic preconditioning with four cycles of 5-minute coronary occlusions resulted in a 94% reduction of infarct size (p < 0.01). Myocardial glycogen and lactate levels were maintained near control levels during hypothermic ischemia.. From these data we conclude that hypothermia during the preconditioning period increases the threshold for eliciting the infarct limitation of ischemic preconditioning.

Topics: Animals; Body Temperature; Coronary Circulation; Disease Models, Animal; Glycogen; Hypothermia, Induced; Ischemic Preconditioning, Myocardial; Lactic Acid; Male; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Rabbits; Ventricular Function, Left

Substrate depletion and increased intracellular acidity are believed to underlie clinically important manifestations of myocardial ischaemia. Recent advances in measuring ion concentrations and metabolite changes have provided a wealth of detail on the processes involved. Coupled with the rapid increase in computing power, this has allowed the development of a mathematical model of cardiac metabolism in normal and ischaemic conditions. Pre-existing models of cardiac cells such as Oxsoft HEART contain highly developed dynamic descriptions of cardiac electrical activity. While biophysically detailed, these models do not yet incorporate biochemical changes. Modelling of bioenergetic changes was based and verified against whole heart NMR spectroscopy. In the model, ATP hydrolysis and generation are calculated simultaneously as a function of [Pi]i. Simulation of pH regulation was based on the pHi dependency of acid efflux, examined in time-course studies of pHi recovery (measured in myocytes with the fluorophore carboxy-SNARF-1) from imposed acid and alkali loads. The force-[Ca2+]i relationship of myofibrils was used as the basis of modelling H+ competition with Ca2+, and thus of pH effects on contraction. This complex description of biochemically important changes in myocardial ischaemia was integrated into the OXSOFT models. The model is sufficiently complete to simulate calcium-overload arrhythmias during ischaemia and reperfusion-induced arrhythmias. The timecourse of both metabolite and pH changes correlates well with clinical and experimental studies. The model possesses predictive power, as it aided the identification of electrophysiological effects of therapeutic interventions such as Na(+)-H+ block. It also suggests a strategy for the control of cardiac arrhythmias during calcium overload by regulating sodium-calcium exchange. In summary, we have developed a biochemically and biophysically detailed model that provides a novel approach to studying myocardial ischaemia and reperfusion.

Topics: Adenine Nucleotides; Algorithms; Animals; Computational Biology; Energy Metabolism; Glycogen; Heart; Humans; Hydrogen-Ion Concentration; Models, Cardiovascular; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Sarcolemma

Experimental data from isolated rat hearts suggest that glycolysis in severe myocardial ischemia is inhibited by accumulation of glycolytic metabolites. In contrast, positron emission tomography (PET) in patients with myocardial ischemia records a 'mismatch' between the decreased coronary flow in viable ischemic tissue and an increased fluorodeoxyglucose (18FDG) signal. To resolve this contradiction, we investigated glucose uptake at very low coronary flows in the isolated rat heart model.. Rates of glucose uptake were measured in the isolated Langendorff-perfused Wistar rat heart, at control (12-15 ml/g wet wt/min) and low coronary flows (0.1, 0.2 and 0.5 ml/g wet wt/min) and at a range of glucose concentrations (2.75, 5.5, 11 and 22 mM).. The steady-state rate of glucose uptake versus glucose concentration could be described by a double rectangular hyperbola at each coronary flow. Glucose uptake fell to levels significantly below control at low coronary flows. However, the extraction of glucose (glucose uptake as % of glucose delivered) rose sharply, from 1% at control coronary flows, to 25-30% at low coronary flows. Crossover analysis of glycolytic intermediates in freeze-clamped tissue indicated little inhibition at any specific site, although phosphofructokinase activity was restricted when glycolytic substrate availability was high. Insulin and preconditioning both increased glucose uptake with 11 mM glucose, possibly by increasing membrane transporter density and thus increasing glucose delivery to the cytosol.. Despite the reduction in absolute glucose uptake at low coronary flow-rates, the extraction of glucose was greatly increased, possibly following GLUT4 translocation. Delivery of glucose to the glycolytic pathway appears to be a major controlling site of glycolysis in low-flow ischemia. Downstream regulation is then distributed along the pathway with no one site exerting greater inhibition than reduced glucose delivery.

Topics: Animals; Blood Flow Velocity; Coronary Circulation; Dose-Response Relationship, Drug; Fructosephosphates; Glucose; Glucose-6-Phosphate; Glycogen; Glycolysis; Heart; In Vitro Techniques; Insulin; Ischemic Preconditioning, Myocardial; Lactic Acid; Male; Myocardial Ischemia; Myocardium; Perfusion; Phosphates; Rats; Rats, Wistar

Topics: Adenosine Triphosphate; Animals; Glucose; Glycogen; Glycolysis; Heart; In Vitro Techniques; Male; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Nuclear Magnetic Resonance, Biomolecular; Pyruvic Acid; Rats; Rats, Wistar; Sodium; Sodium-Potassium-Exchanging ATPase

Glycogen and its turnover are important components of myocardial glucose metabolism that significantly impact on postischemic recovery. We developed a method to measure glycogen turnover (rates of glycogen synthesis and degradation) in isolated working rat hearts using [3H]- and [14C]glucose. In aerobic hearts perfused with 11 mM glucose, 1.2 mM palmitate, and 100 microU/ml insulin, rates of glycogen synthesis and degradation were 1.24 +/- 0.3 and 0.53 +/- 0. 25 micromol. min-1. g dry wt-1, respectively. Low-flow ischemia (0.5 ml/min, 60 min) elicited a marked glycogenolysis; rates of glycogen synthesis and degradation were 0.54 +/- 0.16 and 2.12 +/- 0.14 micromol. min-1. g dry wt-1, respectively. During reperfusion (30 min), mechanical function recovered to 20% of preischemic values. Rates of synthesis and degradation were 1.66 +/- 0.16 and 1.55 +/- 0. 21 micromol. min-1. g dry wt-1, respectively, and glycogen content remained unchanged (25 +/- 3 micromol/g dry wt). The assessment of glycogen metabolism needs to take into account the simultaneous synthesis and degradation of glycogen. With this approach, a substantial turnover of glycogen was detectable not only during aerobic conditions but also during ischemia as well as reperfusion.

Topics: Animals; Glycogen; Glycolysis; Male; Myocardial Ischemia; Myocardial Reperfusion Injury; Rats; Rats, Sprague-Dawley

This study tested the hypothesis that endogenous adenosine depresses anaerobic glycolysis in preischaemic and moderately ischaemic myocardium.. Isolated, working guinea-pig hearts, perfused with glucose-fortified Krebs-Henseleit buffer, were subjected to 15 min mild hypoperfusion (coronary flow 60% of baseline) followed by 10 min ischaemia (coronary flow 20% of baseline). Adenosine A1 receptors were blocked with 8-p-sulfophenyl theophylline (8-SPT; 20 microM). Glucose oxidation and lactate production from exogenous glucose were assessed from 14CO2 and [14C]lactate formation, respectively, from [U-14C]glucose. Energy metabolites, glycolytic intermediates and glycogen were measured in extracts of stop-frozen preischaemic, mildly hypoperfused and ischaemic myocardium.. Adenosine receptor blockade did not affect left ventricular function assessed from heart rate x pressure product and pressure x volume work although coronary flow was slightly reduced. Adenosine receptor blockade increased glucose uptake (P < 0.05) by 100% during preischaemia and by 74% during mild hypoperfusion, and increased lactate production from exogenous glucose (P < 0.05) by 89% during preischaemia and fourfold during mild hypoperfusion, but did not stimulate glucose oxidation under any condition. Glycogen degradation was not increased by adenosine receptor blockade during ischaemia. Crossover plots of glycolytic intermediates revealed that phosphofructokinase was activated by adenosine receptor blockade at all three levels of perfusion.. Endogenous adenosine attenuates anaerobic glycolysis in normally perfused, hypoperfused and ischaemic myocardium by blunting phosphofructokinase activity; this effect is mediated by adenosine A1 receptors.

Topics: Adenosine; Animals; Enzyme Activation; Glucose; Glycogen; Glycolysis; Guinea Pigs; In Vitro Techniques; Lactic Acid; Myocardial Ischemia; Myocardium; Oxidation-Reduction; Perfusion; Phosphofructokinase-1; Purinergic P1 Receptor Antagonists; Theophylline; Ventricular Function, Left

Substantial release of bradykinin has been demonstrated to occur during short periods of myocardial ischaemia in various species. The aim of the present study was to investigate the protective effect of bradykinin in ischaemia and whether bradykinin could be involved in ischaemic preconditioning in the rat heart.. Isolated, buffer-perfused hearts were subjected to 30 min of global ischaemia, followed by 30 min of reperfusion. Postischaemic functional recovery was recorded in the following groups: (1) control; (2) treatment with 0.1 microM bradykinin for 10 min before ischaemia (BK); (3) bradykinin treatment combined with pretreatment with the specific bradykinin B2-receptor antagonist, HOE 140; (4) ischaemic preconditioning by 5 min ischaemia +5 min reperfusion prior to sustained ischaemia (i.p.); and (5) ischaemic preconditioning combined with HOE 140 administration.. Postischaemic myocardial function was significantly improved in both BK and i.p. groups (developed pressure 66.9 +/- 6.8 and 67.6 +/- 7.1 mmHg, respectively, vs. 43.1 +/- 5.9 mmHg in controls, P < 0.05). Pretreatment with 1 microM HOE 140 completely abolished the effect of bradykinin, while protection achieved by i.p. was unaltered by this drug. None of the protective interventions was associated with any significant improvement in myocardial adenosine triphosphate, creatine phosphate, glycogen, lactate or glucose tissue levels, detected either at the end of ischaemia or after 30 min of reperfusion.. Bradykinin, acting via B2-receptors, can protect against postischaemic contractile dysfunction to a similar extent as i.p.. An involvement of B2-receptors in the ischaemic preconditioning phenomenon could, however, not be demonstrated.

Topics: Adenosine Triphosphate; Animals; Bradykinin; Bradykinin Receptor Antagonists; Glucose; Glycogen; Ischemic Preconditioning, Myocardial; Lactic Acid; Male; Myocardial Contraction; Myocardial Ischemia; Myocardium; Phosphocreatine; Rats; Rats, Wistar

We investigated, at first in low-flow global ischemia and then with ischemic preconditioning, the effects of a compound, (4-isopropyl-3-methylsulphonylbenzoyl)guanidine hydrochloride (HOE 642), known to inhibit the Na+/H+ exchange in rat cardiomyocytes. In rat isolated hearts, perfused on a Langendorff apparatus with Krebs-Henseleit carbonate buffer, the action potentials and the contractile function were measured during a 25-min period of global low-flow ischemia (coronary flow, 0.3 mL.min-1) followed by a 30-min reperfusion. In hearts previously preconditioned, two intermittent periods of total ischemia for 5 min each, separated by 5 min reflow, were performed before low-flow ischemia. Treated hearts received HOE 642 (3.0 x 10(-8) mol.min-1) exclusively during low-flow ischemia. Treatment with HOE 642 during low-flow ischemia improves cardiac performance and lowers the rise in diastolic tension during reperfusion. Concomitantly HOE 642 shortens the action potential, and has striking effects on ventricular arrhythmias during reperfusion as well. These results support the concept that Na+/H+ exchange activation is a contributing factor to low-flow ischemia-reperfusion injuries. HOE 642 exhibited minor effects when combined with the preconditioning protocol, but a lengthening in action potential was observed and ventricular arrhythmias were mostly affected. Preconditioned hearts demonstrated marked glycogen depletion compared with controls. These results support the hypothesis that preconditioning could decrease glycogenolysis and therefore subsequently limit acidification during low-flow ischemia.

Topics: Action Potentials; Animals; Electrophysiology; Female; Glycogen; Guanidines; Heart; Hemodynamics; Myocardial Contraction; Myocardial Ischemia; Myocardium; Rats; Rats, Wistar; Reperfusion Injury; Sodium-Hydrogen Exchangers; Sulfones

Vanadium compounds have been shown to cause a variety of biological and metabolic effects including inhibition of certain enzymes, alteration of contractile function, and as an insulin like regulator of glucose metabolism. However, the influence of vanadium on metabolic and ionic changes in hearts remains to be understood. In this study we have examined the influence of vanadate on glucose metabolism and sodium transport in isolated perfused rat hearts. Hearts were perfused with 10 mM glucose and varying vanadate concentrations (0.7-100 microM) while changes in high energy phosphates (ATP and phosphocreatine (PCr)), intracellular pH, and intracellular sodium were monitored using 31P and 23Na NMR spectroscopy. Tissue lactate, glycogen, and (Na+, K+)-ATPase activity were also measured using biochemical assays. Under baseline conditions, vanadate increased tissue glycogen levels two fold and reduced (Na+, K+)-ATPase activity. Significant decreases in ATP and PCr were observed in the presence of vanadate, with little change in intracellular pH. These changes under baseline conditions were less severe when the hearts were perfused with glucose, palmitate and beta-hydroxybutyrate. During ischemia vanadate did not limit the rise in intracellular sodium, but slowed sodium recovery on reperfusion. The presence of vanadate during ischemia resulted in attenuation of acidosis, and reduced lactate accumulation. Reperfusion in the presence of vanadate resulted in a slower ATP recovery, while intracellular pH and PCr recovery was not affected. These results indicate that vanadate alters glucose utilization and (Na+, K+)-ATPase activity and thereby influences the response of the myocardium to an ischemic insult.

Topics: 3-Hydroxybutyric Acid; Adenosine Triphosphate; Animals; Energy Metabolism; Glucose; Glycogen; Glycolysis; Heart; Hydrogen-Ion Concentration; Hydroxybutyrates; In Vitro Techniques; Kinetics; Lactates; Magnetic Resonance Spectroscopy; Male; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Palmitic Acid; Perfusion; Phosphocreatine; Rats; Rats, Sprague-Dawley; Sodium; Sodium-Potassium-Exchanging ATPase; Vanadates

Interruption of ischemia by brief reperfusions (I/R) is better tolerated by the heart than continuous ischemia. The present study aims to determine the metabolic profiles of isolated rat hearts during intermittent ischemia, the possible cardioprotective role of adenosine and the influence of I/R on intracellular volumes, using multinuclear NMR spectroscopy. After five I/R (5/5 min) episodes, hearts paced at 5 Hz developed pressures comparable to those of hearts continuously perfused for 50 min at 37 degrees C (CP). Following the first 5 min episode of no-flow ischemia, [ADP] dropped from 72 +/- 9 to 43 +/- 5 microM (P < 0.001) and remained stable at the end of the following reperfusions, despite a 2.5-4-fold increase during each episode of 5 min ischemia. Intracellular volumes were stable during CP at a value of 2.50 +/- 0.06 ml/g dry weight, and decreased by 4, 8, and 12% after 1, 3, and 5 I/R episodes. The phosphorylation potentials decreased from 54 +/- 8 to 4 mM-1 during each period of 5 min ischemia and were 40 +/- 6 and 28 +/- 6 mM-1 after CP and I/R5, respectively. Cardiac glycogen had decreased during 50 min of CP from 103 +/- 13 to 81 +/- 9 mumol/g dry weight and lactate production was 116 +/- 15 mumol/heart. Five I/R episodes decreased glycogen to 46 +/- 7 mumol/g dry weight (P < 0.005 v CP) and increased lactate efflux to 262 +/- 31 mumol/ heart (P < 0.005 v CP). These findings suggest that a brief ischemia/reperfusion episode increases anaerobic metabolism of exogenous glucose, reduces [ADP] and induces cellular shrinkage. Administration of the adenosine receptor blocker 8-phenyl theophylline (8PT) during intermittent perfusion depressed the developed pressure to 78 +/- 7%, accentuated the decrease in phosphorylation potential (14 +/- 4 mM-1), abolished cellular shrinkage, reduced lactate efflux and blunted the decrease in ADP following the first I/R episode. In variance, no detectable changes were observed during intermittent ischemia when the ATP-sensitive potassium channel blocker glibenclamide was administered. These data demonstrate: (a) a brief episode of ischemia/reperfusion stimulates anaerobic metabolism of exogenous glucose and lowers intracellular ADP concentration: (b) adenosine receptors are partially responsible for the glycolytic stimulation during intermittent ischemia; (c) cellular shrinkage is related to the rate of glycolysis during intermittent ischemia/reperfusion.

Topics: Adenosine; Animals; Cell Size; Creatine; Energy Metabolism; Glyburide; Glycogen; Glycolysis; In Vitro Techniques; Ischemic Preconditioning, Myocardial; Lactic Acid; Magnetic Resonance Spectroscopy; Male; Myocardial Contraction; Myocardial Ischemia; Myocardium; Perfusion; Phosphorus; Phosphorylation; Potassium Channels; Rats; Rats, Sprague-Dawley; Receptors, Purinergic P1; Theophylline

Ischemic preconditioning (IPC) attenuates acidosis during prolonged ischemia and improves contractile and metabolic parameters during subsequent reperfusion. Glycogen depletion induced by IPC is proposed as a potential mechanism.. We studied the influence of manipulations of preischemic glycogen levels (Pre-G, micromol glucose/g wet wt) on contractile and metabolic (via 31P-nuclear magnetic resonance) parameters during 30 minutes of ischemia and recovery in four groups of isovolumic rat hearts: First, control (Con, n=18, mean Pre-G, 21.5+/-0.8); second, after two 5-minute IPC periods (IPC, n=12, Pre-G, 11.3+/-0.7); third, a control group in which Pre-G was depleted by glucose-free, acetate perfusion (Con-LowG, n=9, Pre-G, 7.9+/-1.2); and fourth, an IPC group in which Pre-G was raised by glucose and lactate perfusion such that Pre-G was similar to Con (IPC-HiG, n=11, Pre-G, 20+/-1.4). Manipulation of Pre-G significantly altered the pH fall during 30 minutes of ischemia (Con, 5.76+/-.03, Con-LowG, 6.26+/-.07; IPC-HiG, 5.91+/-.02, IPC, 6.05+/-.09). IPC-HiG hearts had significantly worse metabolic recovery (PCr, 70+/-7 versus 91+/-3% initial; IPC-HiG versus IPC, P<.05) and contractile recovery (end-diastolic pressure, 52+/-5 versus 29+/-5 mm Hg, P<.05) than IPC hearts but better recovery than Con (%PCr, 56+/-6% and end-diastolic pressure, 72+/-6 mm Hg). An ischemic rise in intracellular magnesium occurred and was atttenuated in preconditioned hearts.. Pre-G levels before ischemia influence but are not the sole determinants of the extent of acidosis during prolonged ischemia and of metabolic and contractile recovery during reperfusion in control and preconditioned hearts.

Topics: Animals; Glycogen; Hydrogen-Ion Concentration; In Vitro Techniques; Ischemic Preconditioning; Magnesium; Magnetic Resonance Spectroscopy; Male; Myocardial Contraction; Myocardial Ischemia; Myocardium; Rats; Rats, Wistar; Time Factors

Topics: Adaptation, Physiological; Adenosine Triphosphate; Animals; Energy Metabolism; Glycogen; Hypoxia; In Vitro Techniques; Kinetics; Lactic Acid; Male; Monitoring, Physiologic; Myocardial Ischemia; Myocardium; Oxygen; Perfusion; Rats; Rats, Sprague-Dawley

The effects of BAY o 1248, an inhibitor of alpha-amylo-1, 6-glucosidase, on glycogenolysis and post-ischemic functional recovery were investigated in isolated perfused rat hearts. Working rat hearts were perfused during 30 min with 11 mm glucose (controls) and, in some hearts, with 1 microM insulin or 5 mM lactate to increase their glycogen concentration. The hearts were then submitted to 10 min of no-flow ischemia and reperfused during 15 min with 11 mM glucose alone. Glycogen content was increased by 50% in hearts perfused with insulin or lactate. During ischemia, glycogen breakdown was similar in the control and lactate groups, but was abolished in the insulin-group. At reperfusion, functional recovery was improved in glycogen-loaded hearts compared to controls. When hearts were perfused with 1 mM BAY o 1248, added before ischemia, glycogenolysis was inhibited in the three groups and functional recovery was hampered in both the control and lactate groups. In the insulin group, however, the functional recovery was barely affected by BAY o 1248. We conclude that: (i) BAY o 1248 is an inhibitor of heart glycogen breakdown; (ii) the consequences of inhibition of ischemic glycogenolysis on post-ischemic functional recovery depend on the conditions; and (iii) the protective effect of insulin does not result from ischemic glycogenolysis.

Topics: 1-Deoxynojirimycin; Animals; Coronary Circulation; Depression, Chemical; Enzyme Inhibitors; Glucosamine; Glucose; Glycogen; Glycogen Debranching Enzyme System; Heart; Insulin; Lactates; Male; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Perfusion; Rats; Rats, Wistar

Fasting increases myocardial glycogen content and has been shown to limit injury and improve recovery following no-flow ischaemia in the isolated heart. However, the protective role of glycogen loading per se in fed animals has been questioned by data in preconditioned animals showing that reduced glycogenolysis may be protective prior to no-flow ischemia. Therefore, we hypothesized that fasting protects the globally ischemic heart by mechanisms separate from glycogen loading.. Isolated hearts from rats fasted for 24 h were retrogradely perfused using glucose substrate and subjected to 20 min of global no-flow ischemia. Fed rats were identically perfused either under control conditions (glucose substrate) or with an intervention chosen to increase tissue glycogen (glucose plus insulin, [insulin]) prior to ischemia. Functional recovery and creatine kinase (CK) release were measured during reperfusion. Nuclear magnetic resonance spectroscopy was used to measure intracellular pH, phosphorylated glycolytic intermediates and high-energy phosphates, while the lactate and pyruvate contents of the hearts were measured prior to and at the end of ischemia.. Heart from fasted animals had significantly increased glycogen content prior to ischemia (76.6 +/- 2 vs. 40.9 +/- 3 mumol glu/gdw in control hearts, P < 0.05) as did hearts exposed to insulin (88.6 +/- 10 mumol glu/gdw), but only hearts from fasted animals had greater glycogen utilization during ischemia. Hearts from fasted animals also had lower levels of lactate relative to pyruvate (L/P) under baseline conditions and, on reperfusion, reduced CK release (fasted: 183 +/- 48 versus control: 756 +/- 56 IU/gdw, P < 0.05). Conversely, insulin hearts had increased CK release (1831 +/- 190 IU/gdw, P < 0.001 vs control) and worse functional and metabolic recovery on reperfusion. Compared to the insulin hearts, hearts from fasted animals had both less acidosis and less rapid depletion of ATP during ischemia, as well as lower accumulation of glycolytic intermediates.. Fasting protects the heart from ischemic injury and is associated with a lower L/P ratio and increased glycogen utilization during ischemia. In contrast, increasing glycogen content in hearts from fed animals using insulin limits glycogen utilization, increases ischemic injury, and impairs both functional and metabolic recovery under conditions of 20 min of global no-flow ischemia.

Topics: Animals; Creatine Kinase; Fasting; Glycogen; Glycolysis; Hydrogen-Ion Concentration; Lactic Acid; Magnetic Resonance Spectroscopy; Male; Myocardial Ischemia; Myocardium; Perfusion; Pyruvic Acid; Rats; Rats, Sprague-Dawley

Although hypertrophied hearts have increased rates of glycolysis under aerobic conditions, it is controversial as to whether glucose metabolism during ischemia is altered in the hypertrophied heart. Because endogenous glycogen stores are a key source of glucose during ischemia, we developed a protocol to label the glycogen pool in hearts with either [3H]glucose or [14C]glucose, allowing for direct measurement of both glycogen and exogenous glucose metabolism during ischemia. Cardiac hypertrophy was produced in rats by banding the abdominal aorta for an 8-week period. Isolated hearts from aortic-banded and sham-operated rats were initially perfused under substrate-free conditions to decrease glycogen content to 40% of the initial pool size. Resynthesis and radiolabeling of the glycogen pool with [3H]glucose or [14C]glucose were accomplished in working hearts by perfusion for a 60-minute period with 11 mmol/L [3H]glucose or [14C]glucose, 0.5 mmol/L lactate, 1.2 mmol/L palmitate, and 100 mumol/mL insulin. Although glycolytic rates during the aerobic perfusion were significantly greater in hypertrophied hearts compared with control hearts, glycolytic rates from exogenous glucose were not different during low-flow ischemia. The contribution of glucose from glycogen was also not different in hypertrophied hearts compared with control hearts during ischemia (1314 +/- 665 versus 776 +/- 310 nmol.min-1.g dry wt-1, respectively). Glucose oxidation rates decreased during ischemia but were not different between the two groups. However, in both hypertrophied and control hearts, the ratio of glucose oxidation to glycolysis was greater for glucose originating from glycogen than from exogenous glucose. Our data demonstrate that glycogen is a significant source of glucose during low-flow ischemia, but the data do not differ between hypertrophied and control hearts.

Topics: Animals; Cardiomegaly; Coronary Circulation; Glucose; Glycogen; Glycolysis; Heart; Male; Myocardial Ischemia; Myocardium; Oxidation-Reduction; Rats; Rats, Sprague-Dawley

Hearts from hyperinsulinemic, insulin-resistant JCR:LA-cp rats do not properly regulate intracellular Ca2+ concentration. We hypothesized, therefore, that these hearts may be unusually sensitive to ischemic insults in which Ca2+ overload would be expected. We investigated the response to global ischemia of hearts from JCR: LA-cp animals at three different ages. At 3 mo of age, isolated hearts from insulin-resistant cp rats were mildly resistant to both mild and severe ischemic insults in comparison to the lean control rat hearts. However, at 6 and 9 mo of age, the cp rats demonstrated a poorer recovery of developed tension after ischemia and/or a higher level of resting tension during reperfusion than the lean controls. Postischemic glycogen and ATP contents were significantly lower and lactate content was higher in hearts from 6-mo-old cp rats compared with controls. The results demonstrate that an insulin-resistant animal model exhibits an increased sensitivity to ischemic myocardial injury that develops with advancing age. The mechanism responsible for the enhanced sensitivity may involve augmented glycolytic metabolism. The data also emphasize the importance of the type of diabetes when cardiac dysfunction is examined.

Topics: Adenosine Triphosphate; Aging; Animals; Glycogen; Heart; In Vitro Techniques; Insulin Resistance; Lactates; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Rats; Rats, Mutant Strains; Reference Values; Triglycerides

We tested the hypothesis that low-flow ischemia increases glucose uptake and reduces insulin responsiveness. Working hearts from fasted rats were perfused with buffer containing glucose alone or glucose plus a second substrate (lactate, octanoate, or beta-hydroxybutyrate). Rates of glucose uptake were measured by 3H2O production from [2-3H]glucose. After 15 min of perfusion at a physiological workload, hearts were subjected to low-flow ischemia for 45 min, after which they were returned to control conditions for another 30 min. Insulin (1 mU/ml) was added before, during, or after the ischemic period. Cardiac power decreased by 70% with ischemia and returned to preischemic values on reperfusion in all groups. Low-flow ischemia increased lactate production, but the rate of glucose uptake during ischemia increased only when a second substrate was present. Hearts remained insulin responsive under all conditions. Insulin doubled glucose uptake when added under control conditions, during low-flow ischemia, and at the onset of the postischemic period. Insulin also increased net glycogen synthesis in postischemic hearts perfused with glucose and a second substrate. Thus insulin stimulates glucose uptake in normal and ischemic hearts of fasted rats, whereas ischemia stimulates glucose uptake only in the presence of a cosubstrate. The results are consistent with two separate intracellular signaling pathways for hexose transport, one that is sensitive to the metabolic requirements of the heart and another that is sensitive to insulin.

Topics: 3-Hydroxybutyric Acid; Animals; Biological Transport; Caprylates; Citrates; Coronary Circulation; Glucose; Glycogen; Heart; Hydroxybutyrates; In Vitro Techniques; Insulin; Lactates; Male; Myocardial Ischemia; Myocardium; Perfusion; Rats; Rats, Sprague-Dawley

The cardioprotective effect of preconditioning is associated with glycogen depletion and attenuation of intracellular acidosis during subsequent prolonged ischemia. This study determined the effects of increasing preconditioning ischemia time on myocardial glycogen depletion and on infarct size reduction. In addition, this study determined whether infarct size reduction by preconditioning correlates with glycogen depletion before prolonged ischemia. Anesthetized rats underwent a single episode of preconditioning lasting 1.25, 2.5, 5, or 10 minutes or multiple episodes cumulating in 10 (2 x 5 min) or 20 minutes (4 x 5 or 2 x 10 min) of preconditioning ischemia time, each followed by 5 minutes of reperfusion. Then both preconditioned and control rats underwent 45 minutes of ischemia induced by left coronary artery (LCA) occlusion and 120 minutes of reperfusion. After prolonged ischemia, infarct size was determined by dual staining with triphenyltetrazolium chloride and phthalocyanine blue dye. Glycogen levels were determined by an enzymatic assay in selected rats from each group before prolonged ischemia. We found that increasing preconditioning ischemia time resulted in glycogen depletion and infarct size reduction that could both be described by exponential functions. Furthermore, infarct size reduction correlated with glycogen depletion before prolonged ischemia (r = 0.98; p < 0.01). These findings suggest a role for glycogen depletion in reducing ischemic injury in the preconditioned heart.

Topics: Acidosis; Animals; Female; Glycogen; Myocardial Infarction; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Rats; Rats, Sprague-Dawley; Time Factors

A high glycogen level may be beneficial to the ischemic heart by providing glycolytic ATP or detrimental by increasing intracellular lactate and protons. To determine the effect of high glycogen on the ischemic myocardium, the glycogen content of Langendorff-perfused rat hearts was either depleted or elevated before 32 minutes of low-flow (0.5 mL/min) ischemia with Krebs-Henseleit buffer with or without 11 mmol/L glucose, followed by 32 minutes of reperfusion with buffer containing 11 mmol/L glucose. 31P nuclear magnetic resonance spectra were acquired sequentially throughout. Further experiments involved early reperfusion or the addition of HOE 694, a Na+-H+ exchange inhibitor, during reperfusion. When glucose was supplied throughout ischemia, no ischemic contracture occurred, and postischemic recovery of contractile function was highest, at 88% of preischemic function. In the absence of glucose, normal-glycogen hearts underwent ischemic contracture at 5 minutes, had an end-ischemic pH of 6.87, and recovered to 54%, whereas in high-glycogen hearts, contracture was delayed to 13 minutes, the end-ischemic pH was 6.61, and functional recovery decreased to 13%. Contracture onset coincided with the decrease in glycolysis, which occurred as glycogen became fully depleted. Functional recovery in the high-glycogen hearts increased to 89% when reperfused before contracture and to 56% when reperfused in the presence of HOE 694. Thus, during brief ischemia in the high-glycogen hearts, ischemic glycogen depletion and contracture were avoided, and the hearts were protected from injury. In contrast, during prolonged ischemia in the high-glycogen hearts, glycogen became fully depleted, and myocardial injury occurred; the injury was exacerbated by the lower ischemia pH in these hearts, leading to increased Na+-H+ exchange during reperfusion. The contradictory findings of past studies concerning the effect of high glycogen on the ischemic myocardium may thus be due to differences in the extent of glycogen depletion during ischemia.

Topics: Adenosine Triphosphate; Animals; Coronary Circulation; Glycogen; Glycolysis; Hydrolysis; In Vitro Techniques; Lactates; Lactic Acid; Male; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Rats; Rats, Wistar

Short-term myocardial hibernation of 3 hours resulting from a moderate resting coronary flow reduction has been reproduced in pigs. This study was designed to determine whether any structural changes accompany short-term hibernation caused by a moderate flow reduction maintained for 24 hours and whether any such structural alterations are reversible after reperfusion.. A severe left anterior descending coronary artery (LAD) stenosis was created with a reduction of resting flow to approximately 60% of baseline and maintained for 24 hours. Regional coronary flow was measured by a flowmeter; wall thickening was determined by echocardiography, and local metabolic changes were measured. Of 17 pigs, 11 completed the study protocol of 24 hours. The LAD flow was reduced from 0.91 +/- 0.11 to 0.52 +/- 0.13 mL.min-1.g-1, a 43% mean decrease, at 15 minutes after the LAD stenosis and was maintained at 0.56 +/- 0.11 mL.min-1.g-1 at 24 hours. The reduction of regional coronary flow initially produced acute myocardial ischemia, as evidenced by reduced regional wall thickening (from 37.2 +/- 6.9% at baseline to 11.5 +/- 6.8%), regional lactate production (-0.34 +/- 0.28 mumol.g-1.min-1), and a decrease in regional coronary venous pH (from 7.41 +/- 0.035 at baseline to 7.30 +/- 0.030). At 24 hours, the reductions in coronary flow and wall thickening were maintained relatively constant and the rate-pressure product was relatively unchanged, but lactate production ceased and regional H+ concentration normalized, with a tendency toward a further reduction in regional oxygen consumption, from 3.10 +/- 0.90 mL.min-1.100 g-1 at 15 minutes after stenosis to 2.52 +/- 0.95 mL.min-1.100 g-1 at 24 hours (P = .06), indicating metabolic adaptation of the hypoperfused regions. Of 11 pigs, 6 were free of myocardial infarction; 3 had patchy necrosis involving 4%, 5%, and 6% of the area at risk; and 2 other pigs had a few scattered myocytes with necrosis, detected only by light and electron microscopy. Ultrastructural changes consisted of a partial loss of myofibrils and an increase in mitochondria and glycogen deposition. Regional wall thickening recovered 1 week after reperfusion in most pigs, and the ultrastructural changes reverted to normal.. In this pig model, moderately ischemic myocardium undergoes metabolic and structural adaptations but preserves the capacity to recover both functionally and ultrastructurally after reperfusion.

Topics: Actin Cytoskeleton; Animals; Coronary Circulation; Coronary Disease; Glycogen; Mitochondria, Heart; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Necrosis; Oxygen Consumption; Swine

We previously reported that adenosine A1 receptor activation protects against the cardiodepressant effects of hydrogen peroxide in isolated rat hearts. The present study examined whether a transient ischemic period of 5 min duration, which preconditions the heart against ischemic and reperfusion-induced dysfunction, can bestow protection against 30-min exposure to hydrogen peroxide in isolated rat hearts. Transient ischemia on its own failed to alter the cardiac response to hydrogen peroxide. However, when transient ischemia was carried out in the presence of the nucleoside transport inhibitor S-(4-Nitrobenzyl)-6-thioguanosine and the adenosine deaminase inhibitor erythro-9-(2-Hydroxy-3-nonyl)adenine, a significant attenuation of the hydrogen peroxide-induced loss in contractility was evident and this was associated with significant preservation of tissue glycogen content. The protective effect of the transient ischemia/drug combination on both functional changes and glycogen levels was abolished by the adenosine A1 receptor antagonist 8-cyclopentyl-1, 3-dipropylxanthine as well as by glibenclamide, a blocker of the ATP-sensitive potassium channel (KATP). To further assess the role of glycogen in the protection against hydrogen peroxide, we compared the effects of the adenosine A1 agonist N6-cyclopentyl adenosine (CPA) and insulin. While both treatments protected against hydrogen peroxide the effect of insulin was superior to any other treatment. Moreover, while all protective modalities preserved glycogen stores after hydrogen peroxide treatment, the protection afforded by insulin was also associated with significantly elevated glycogen levels prior to hydrogen peroxide administration. No protection by either CPA or insulin was evident in the absence of exogenous glucose. Taken together, our results demonstrate that a brief period of ischemia with concomitant administration of agents which increase interstitial adenosine levels protects against hydrogen peroxide toxicity. The effect is mediated by activation of adenosine A1 receptors and is linked to KATP stimulation. Moreover, our results are strongly suggestive of an important role of glycogen preservation in bestowing protective effects against hydrogen peroxide cardiotoxicity.

Topics: Adenosine Deaminase; Adenosine Deaminase Inhibitors; Animals; Depression, Chemical; Enzyme Inhibitors; Glycogen; Guanosine; Hydrogen Peroxide; Male; Myocardial Contraction; Myocardial Ischemia; Oxidants; Rats; Rats, Sprague-Dawley; Thionucleosides

The role of lactate accumulation in lethal ischemic myocardial cell injury was assessed by partially depleting hearts of glycogen before ischemia by using glucagon. Isolated adult rat hearts were perfused with glucose-free Krebs-Henseleit buffer containing acetate as substrate. After stabilization, treated hearts were perfused briefly (3 min) with buffer containing 2 micrograms/ml glucagon to reduce tissue glycogen stores, followed by 10 min of perfusion with control buffer, and 60 or 90 min of global ischemia. Before the onset of ischemia, glucagon-treated hearts contained 40% less glycogen than untreated hearts, but myocardial function and tissue levels of high-energy phosphates, lactate, and glucose 6-phosphate were similar. Lactate production during ischemia in the glucagon-treated hearts was 50% less than in untreated hearts. However, there was no decrease in the amount of creatine kinase release during reperfusion after either 60 or 90 min of ischemia. Thus although partial glycogen depletion reduced lactate accumulation during ischemia, this did not decrease the amount of lethal myocardial cell injury.

Topics: Animals; Creatine Kinase; Energy Metabolism; Glucagon; Glycogen; In Vitro Techniques; Lactic Acid; Male; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Perfusion; Phosphates; Rats; Rats, Sprague-Dawley; Time Factors

Prior transient episodes of ischemia ("ischemic preconditioning") reduce lactate accumulation and attenuate acidosis during a subsequent prolonged ischemic insult. The mechanisms responsible for attenuated glycolytic catabolite accumulation have not been established but may include earlier exhaustion of glycogen stores, slowed glycogenolysis before complete glycogen depletion, and/or inhibition of glycolysis. Simultaneous repeated measures of myocardial glycogen and the rates of glycolysis, glycogenolysis, glucose utilization, and glycolytic ATP production were obtained during total ischemia by 13C nuclear magnetic resonance spectroscopy in control and ischemia-preconditioned isolated rat hearts. Both [13C]glycolytic and [13C]glycogenolytic rates were significantly lower during total ischemia in preconditioned compared with control hearts (0.77 +/- 0.04 versus 1.06 +/- 0.06 mumol/min per gram wet weight [P < .01] for glycolysis and 0.15 +/- 0.07 versus 0.78 +/- 0.12 mumol/ min per gram wet weight [P < .001] for glycogenolysis, respectively, at 2.5 minutes of ischemia). Slowed glycolysis was present even during the early minutes of ischemia, when significant amounts of available [13C]glycogen were still present. Importantly, the reduction in the rate of glycogenolysis was larger and out of proportion to the reduction in glycolysis and occurred despite an increase in glucose utilization in preconditioned hearts (2.23 +/- 0.15 versus 1.5 +/- 0.10 mumol/min per gram wet weight at 1.25 minutes, P < .01). During early ischemia, conversion of glycogen phosphorylase to the a or "active" form was less in preconditioned than in control hearts (29.1 +/- 2.6% versus 41.2 +/- 9.8%, respectively; P < .05). Taken together, these findings demonstrate that ischemic preconditioning significantly depresses glycolytic catabolite accumulation during sustained ischemia not by more severe glycolytic inhibition or exhaustion of glycogen stores but by depressed glycogenolysis from the onset of ischemia.

Topics: Animals; Glycogen; Glycolysis; Magnetic Resonance Spectroscopy; Male; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Rats; Rats, Wistar; Time Factors

In the adult heart an increase in extracellular [Ca2+] can contribute to the severity of ischemic injury. While experimental studies have suggested that the immature heart is more resistant to ischemia than the mature heart, the reasons for this are unclear. In this study, we determined the effects of increasing perfusate [Ca2+] from 1.25 to 2.5 mM on reperfusion recovery of mechanical function and energy substrate metabolism following ischemia. Isolated bi-ventricular working hearts from 2-week-old rabbits were subjected to a 55-min period of global ischemia followed by 40 min of aerobic reperfusion. Perfusate contained 11 mM glucose, 0.5 mM lactate, and 1.2 mM palmitate, containing either: (i) 1.25 mM Ca2+ throughout the perfusion period (n = 22), (ii) 1.25 mM Ca2+ prior to and during ischemia and 2.5 mM Ca2+ following ischemia (n = 19), or (iii) 2.5 mM Ca2+ throughout the perfusion period (n = 18). In hearts perfused with 1.25 mM Ca2+ throughout, a 57% recovery of preischemic function was seen following ischemia. If [Ca2+] was increased to 2.5 mM during reperfusion a significant improvement of function was seen (hearts recovered 127% of preischemic function). A concentration of 2.5 mM Ca2+ throughout the perfusion resulted in an increase in both pre- and post-ischemic function compared to hearts perfused with 1.25 mM Ca2+ throughout. In both experimental groups reperfused with 2.5 mM Ca2+ a greater than 200% increase in both glucose and lactate oxidation was seen during reperfusion. Fatty acid oxidation rates also returned to pre-ischemic levels in both groups reperfused with 2.5 mM Ca2+, while rates returned to only 53% in hearts reperfused with 1.25 mM Ca2+. As a result, increasing [Ca2+] from 1.25 to 2.5 mM resulted in a 100% increase in ATP production rates during reperfusion. In conclusion, this study demonstrates that increasing [Ca2+] significantly improves post-ischemic recovery of function in isolated bi-ventricular working immature rabbit hearts subjected to a 55-min period of ischemia. The beneficial effects of Ca2+ in these immature hearts may be due to both a direct inotropic effect and a marked increase in carbohydrate oxidation and ATP production during reperfusion.

Topics: Adenosine Triphosphate; Animals; Animals, Newborn; Calcium; Dichloroacetic Acid; Dose-Response Relationship, Drug; Fatty Acids; Glucose; Glycogen; Heart; Heart Rate; Heart Ventricles; Lactic Acid; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Palmitates; Phosphates; Rabbits

Myocardial injury following ischemia and reperfusion is increased in the aging heart. The mechanisms underlying the increased susceptibility of the aging heart to ischemic injury remain unknown. We investigated whether decreased glycogen utilization with a more rapid depletion of ATP occurred during ischemia in the aging heart. Isolated buffer-perfused hearts from adult (6 months old) and aging (24 months old) Fischer 344 rats were subjected to 0, 2, 5, 10, 15 or 25 min of global stop-flow ischemia following a 15 min equilibration period (n = 5-6 for each ischemic time at each age). ATP level were decreased at preischemic baseline in aging hearts. ATP levels remained lower in the aging heart throughout ischemia (P < 0.001) with a similar pattern of decrease in both age groups. The decrease in tissue glycogen and increase in lactate contents was similar during ischemia in both age groups, suggesting that comparable glycogen utilization occurred during ischemia in adult and aging hearts. ATP catabolism leads to ADP, AMP and then adenosine. Tissue levels of adenosine, an important cardioprotective metabolite, were measured during ischemia. Tissue adenosine levels were decreased by 50% in the aging heart at 5 and 10 min, and remained depressed at 15 min and 25 min of ischemia compared to adult controls. Thus, increased ischemic injury in the aging heart is not related to differences in glycogen consumption. Lower tissue ATP levels and decreased adenosine levels were observed during ischemia. The differences in ATP content between adult and aging hearts occurred only during early ischemia and are unlikely to provide a mechanism for the increased damage observed following more prolonged periods of ischemia in the aging heart. The potential contribution of these decreases in tissue adenosine content to the increased injury observed in the aging heart will require further study.

Topics: Adenosine; Adenosine Triphosphate; Aging; Animals; Cardiovascular Agents; Glycogen; Male; Myocardial Ischemia; Rats; Rats, Inbred F344

Topics: Animals; Glycogen; Iron; Male; Myocardial Ischemia; Rats; Rats, Wistar

We tested the hypothesis that improved ischemia tolerance in an isolated working rat heart preparation can be achieved by interventions other than ischemic preconditioning. Hearts were perfused at near-physiological workload with bicarbonate buffer containing glucose (10 mM). A preischemic period of 25 min was followed by 15 min of global ischemia and 30 min of reperfusion under preischemic conditions. Hearts came from either fed or fasted animals (groups 1 and 2). In group 3 lactate (10 mM) and insulin (10 mU/ml) were added to the perfusate of fasted animals. In group 4 hearts from fed animals were perfused with glucose (10 mM) and were ischemically preconditioned by one cycle of ischemia between 10 and 15 min of the preischemic perfusion. Cardiac power and glucose uptake were measured continuously to assess functional and metabolic recovery. In addition, we measured the time to return of aortic flow. Glucose metabolites and the ratio of latent of free citrate synthase activity (citrate synthase ratio, a marker for the structural integrity of mitochondria) were determined at selected time points. Groups 2, 3, and 4 recovered significantly faster than group 1, whereas recovery of power showed an improvement in groups 3 and 4 only. In addition, there was an early increase in glucose uptake during reperfusion in these two groups, suggesting an early need for glucose substrate. Glycogen levels decreased with ischemia in all groups and returned to preischemic levels in groups 2, 3, and 4. The citrate synthase ratio was low in the control group and preserved in the groups showing improved functional recovery. We conclude that metabolic interventions may be as effective as ischemic preconditioning in protecting the heart from ischemic injury.

Topics: Animals; Fasting; Glucose; Glycogen; Heart; Insulin; Ischemic Preconditioning, Myocardial; Lactic Acid; Male; Mitochondria, Heart; Myocardial Ischemia; Myocardium; Rats; Rats, Sprague-Dawley

This study compared the effects of adenosine (Ado) on the coupling of glycolysis and glucose oxidation and on mechanical function in normal hearts and in hearts subjected to transient ischemia. Isolated working rat hearts were perfused with Krebs containing 1.2 mM palmitate and 100 microU/ml insulin. After 15 min of aerobic perfusion, hearts underwent either two cycles of 10 min of ischemia and 5 min of reperfusion (stressed) or 30 min of aerobic perfusion (control). After 45 min, hearts underwent either aerobic perfusion for 35 min (series A) or 30 min of ischemia and 30 min of reperfusion (series B). In series A, left ventricular minute work (LV work) was similar in control and stressed hearts and was not affected by Ado (500 microM) or N6-cyclohexyladenosine (CHA 0.5 microM). Ado reduced glycolysis by 49% in control hearts but increased glycolysis by 74% in stressed hearts. CHA inhibited glycolysis in both groups by 50 and 62%, respectively. In series B, LV work during reperfusion recovered to a similar extent in untreated control and stressed hearts. In control hearts, Ado reduced glycolysis by 50% while enhancing LV work to 81% of preischemic values. In stressed hearts, Ado increased glycolysis by 34% and depressed LV work to 9%, whereas CHA inhibited glycolysis by 53% and LV work to 91%. These data indicate that coupling of glycolysis to glucose oxidation is a key determinant of mechanical function of the postischemic myocardium. They also show that the metabolic and protective effects of Ado depend on the status of the heart before sustained ischemia.

Topics: Adenosine; Animals; Cardiovascular Agents; Glycogen; Glycolysis; Myocardial Ischemia; Myocardial Reperfusion; Rats; Rats, Sprague-Dawley; Stress, Physiological; Ventricular Function, Left

The mechanism of ischemic preconditioning remains unknown. The role of glycogen depletion prior to prolonged ischemia was examined as a potential mechanism of ischemic preconditioning. The glycogen content of the rat heart varies in a 24-h rhythm. In a retrospective study, the relationships between the time of day the animals were sacrificed, pre-ischemic myocardial glycogen content, and post-ischemic functional recovery were assessed in non-conditioned and ischemically preconditioned hearts. The analyses were performed on previously published data (Asimakis et al.. 1992, 1993). After an equilibration perfusion, isolated rat hearts were given 40 min of global ischemia followed by 30 min of reperfusion. Preconditioned hearts received 5 min of ischemia followed by a 5-min recovery period prior to the 40-min ischemic period. Some of the hearts were freeze-clamped immediately prior to the 40-min ischemic period to determine pre-ischemic glycogen content. Pre-ischemic glycogen was higher in the morning than afternoon. The time of day correlated significantly with the pre-ischemic glycogen content of non-conditioned (r = 0.67; P < 0.005) and preconditioned (r = 0.79; P < 0.001) hearts. However, time of day did not correlate significantly with post-ischemic recovery of heart rate x developed pressure (HR x DP) on end-diastolic pressure (EDP) in either the non-conditioned or preconditioned hearts. The relationships were also assessed by subdividing the groups into either morning (a.m.) or afternoon (p.m.) hearts. The pre-ischemic glycogen content was lower in the non-conditioned-p.m. (n = 5) hearts compared to the non-conditioned-a.m. (n = 10) hearts (67.6 +/- 9.0 nu 128.1 +/- 13.3 nmol glucose/mg protein P < 0.005). However, there were no significant differences between p.m. (n = 13) and a.m. (n = 9) non-conditioned hearts with respect to post-ischemic recovery of HR x DP (20.6 +/- 4 nu 12.0 +/- 4% of baseline, respectively, P = N.S.). In contrast, preconditioned-p.m. (n = 6) and -a.m. (n = 7) had pre-ischemic glycogen contents of 49.6 +/- 6 and 76.6 +/- 5.0 nmol glucose/mg protein, respectively. These glycogen values were not significantly different from the non-conditioned-p.m. hearts (67.6 nmol/mg protein). However, post-ischemic recovery of HR x DP in the preconditioned-p.m. (n = 5) and -a.m. (n = 6) hearts were 54.6 +/- 5 and 51.4 +/- 8% of baseline, respectively (these values were significantly higher (P < 0.05) than the recovery for the non-conditioned-p.

Topics: Adenosine Triphosphate; Animals; Glucose-6-Phosphate; Glycogen; In Vitro Techniques; Ischemic Preconditioning, Myocardial; Lactic Acid; Male; Myocardial Ischemia; Myocardium; Phosphocreatine; Rats; Rats, Sprague-Dawley; Retrospective Studies

The effect of ischemic preconditioning (IPC) on glycolysis, glucose oxidation, adenine nucleotide and nucleoside levels, and mechanical function was studied in isolated paced working rat hearts under aerobic conditions or when reperfused following sustained ischemia. IPC inhibited glycolysis in aerobic hearts (4.48 +/- 0.66 vs. 3.18 +/- 0.39 mumol.min-1.g dry wt-1) and calculated proton production attributable to exogenous glucose (7.79 +/- 1.31 vs. 4.73 +/- 0.81 mumol.min-1.g dry wt-1). In hearts subjected to ischemia and reperfusion, IPC decreased, relative to controls, glycogen content before the onset of ischemia (116.6 +/- 4.3 vs. 158.0 +/- 8.4 mumol/dry wt) and decreased consumption of glycogen during ischemia (54 +/- 6 vs. 76 +/- 7 mumol/dry wt). During reperfusion, glycolysis was lower in IPC hearts (2.45 +/- 0.16 vs. 4.4 +/- 0.46 mumol.min-1.g dry wt-1), as was calculated proton production (3.57 +/- 0.30 vs. 8.38 +/- 0.93 mumol.min-1.g dry wt-1). Glucose oxidation was similar in control and IPC hearts. Adenosine and ATP content of IPC hearts, relative to controls, were higher at the end of ischemia, being 0.86 +/- 0.08 vs. 0.34 +/- 0.15 mumol/g dry wt and 11.3 +/- 0.8 vs. 5.0 +/- 1.6 mumol/g dry wt, respectively. IPC enhanced recovery of ventricular work during reperfusion of ischemic hearts from 37 to 68%. These results indicate that IPC is associated with a reduction in glycogen content, inhibition of glycolysis during ischemia and reperfusion, and a decrease in proton production from glucose. These changes may, in part, explain the enhanced recovery of mechanical function observed in IPC hearts.

Topics: Adenosine Triphosphate; Aerobiosis; Animals; Cardiac Pacing, Artificial; Glucose; Glycogen; Glycolysis; Heart; In Vitro Techniques; Lactates; Lactic Acid; Male; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Oxidation-Reduction; Perfusion; Protons; Rats; Rats, Sprague-Dawley

Polymorphonuclear neutrophils (PMN) play an important role in myocardial ischemia/reperfusion (MI/R) injury; however, the role of neutrophilic proteases is less understood. The effects of a novel serine protease inhibitor (serpin), LEX032, were investigated in a murine model of MI (20 min) and R (24 hr) injury in vivo. LEX032 is a recombinant human alpha 1-antichymotrypsin in which six amino acid residues were replaced around the active center with those of alpha-1 protease inhibitor. LEX032 has the ability to inhibit both neutrophil elastase and cathepsin G, two major neutral serine proteases in neutrophils, as well as superoxide generation. LEX032 (25 or 50 mg/kg) administered i.v. 1 min before reperfusion significantly attenuated myocardial necrotic injury evaluated by cardiac creatine kinase loss compared to MI/R rats receiving only vehicle (P < .001). Moreover, cardiac myeloperoxidase activity, an index of PMN accumulation, in the ischemic myocardium was significantly attenuated by LEX032 as compared with rats receiving vehicle (P < .001). LEX032 also moderately attenuated leukotriene B4-stimulated PMN adherence to rat superior mesenteric artery endothelium and markedly diminished superoxide radical release from LTB4-stimulated PMN in vitro. In a glycogen-induced rat peritonitis model, LEX032 (50 mg/kg) significantly attenuated PMN transmigration into the peritoneal cavity in vivo. In conclusion, the recombinant serine protease inhibitor, LEX032, appears to be an effective agent for attenuating MI/R injury by inhibiting neutrophil-accumulation into the ischemic-reperfused myocardium and by inactivating cytotoxic metabolites (proteases and superoxide radical) released from neutrophils.

Topics: Animals; Cell Adhesion; Chemotaxis, Leukocyte; Creatine Kinase; Endothelium, Vascular; Glycogen; Interleukin-1; Leukotriene B4; Male; Muscle, Smooth, Vascular; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Neutrophils; Peritoneal Cavity; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Serine Proteinase Inhibitors; Serpins; Superoxides

Nitric oxide (NO) exerts both protective and detrimental actions in a variety of biological systems. During acute reperfusion following myocardial ischaemia, a rapid overproduction of free radicals, including NO, may occur. We investigated the effects of the NO synthase inhibitors NG-nitro-L-arginine methyl ester (L-NAME) and NG-monomethyl-L-arginine (L-NMMA), and the substrate for NO synthesis, L-arginine, on heart function during ischaemia and reperfusion injury.. Spontaneously beating, isolated working rabbit hearts, perfused with modified Krebs-Henseleit buffer containing 1.2 mM palmitate bound to 3% bovine serum albumin, were subjected to 15 min of aerobic perfusion followed by 35 min of global, no-flow ischaemia and 30 min of aerobic reperfusion.. Throughout the reperfusion period there was a marked impairment in the recovery of mechanical function, measured as the product of heart rate x peak systolic pressure (rate-pressure product). Addition of L-NAME (3 microM) prior to the onset of ischaemia, but not at reperfusion, caused an immediate and significant increase in the recovery of mechanical function throughout the reperfusion period. The protective action of L-NAME was abolished by L- (but not D-) arginine (100 microM). L-NAME did not cause ischaemia as it did not alter glycogen or lactate content of aerobically perfused hearts. Furthermore, it did not prevent glycogen loss or lactate accumulation during 35 min of ischaemia, suggesting that the effects of L-NAME were not due to metabolic alterations during ischaemia itself. L-NMMA (30 microM) added prior to ischaemia, but not at reperfusion, also had a protective effect which was seen later in the reperfusion period. Addition of L- (but not D-) arginine (100 microM) prior to the onset of ischaemia resulted in an improved recovery of mechanical function only at 15 min of reperfusion.. These results suggest that: (1) the recovery of mechanical function of hearts subjected to ischaemia-reperfusion injury can be improved by modulation of myocardial NO synthesis, (2) inhibition of NO synthesis (with L-NAME or L-NMMA) may offer prolonged protection whereas its stimulation (with L-arginine) provides only brief protection, and (3) the reasons for the pharmacological effectiveness of these divergent strategies may be due to the formation of peroxynitrite from NO and superoxide anion during reperfusion.

Topics: Animals; Arginine; Glycogen; Heart; Lactates; Lactic Acid; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; NG-Nitroarginine Methyl Ester; Nitric Oxide; omega-N-Methylarginine; Perfusion; Rabbits

The purpose of this report was to directly measure the influence of antecedent ischemia or repetitive ischemia on subsequent rates of intermediary metabolism, specifically exogenous glucose utilization and fatty acid oxidation, with the use of myocardial equilibrium labeling with [U-14C]palmitate and [5-3H]glucose. Twenty-one intact, working, extracorporeally perfused pig hearts were prepared and divided into three groups. These groups included 7 control hearts and 14 comparison hearts, which were exposed to either one cycle (cycle 1, n = 7) or four cycles (cycle 4, n = 7) of brief (5-10 min), moderate (70% decrease in flow below aerobic values) precursory ischemia to the left anterior descending (LAD) circulation followed by aerobic reperfusion. All groups then underwent a 40 min sustained LAD ischemia (60% decrease in flow below aerobic levels) and 40 min aerobic reperfusion. Treatment with one cycle of transient ischemia did not significantly modify the pattern of glycolytic flux from control values during sustained ischemia (over a ninefold increase in average control and cycle 1 values above aerobic levels). However, repetitive ischemia in cycle 4 hearts demonstrably attenuated glycolytic flux during the same interval (-45% from control hearts, P < 0.046). Glucose utilization rapidly returned to near-aerobic values in all three groups during reperfusion but was again appreciably lower (P < 0.004 from control values) in cycle 4 hearts. Fatty acid oxidation averaged 12.3 +/- 1.2 mumol.h-1.g dry wt-1 in all three groups during sustained ischemia and 21.3 +/- 2.0 mumol.h-1.g dry wt-1 during reperfusion (not significant among groups for either perfusion interval).(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenine Nucleotides; Animals; Fatty Acids; Glucose; Glycogen; Glycolysis; Lactates; Lactic Acid; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Oxidation-Reduction; Recurrence; Swine

This study was designed to examined the effects of inhalation anaesthetics on function and metabolism in isolated ischaemic rat hearts. Four volatile anaesthetics in two different concentrations (1.0 to 1.5 MAC) were used before whole heart ischaemia was induced for 15 min followed by reperfusion for 30 min. The data were compared with a control group in which inhalation anaesthetics were not used. Before ischaemia, volatile anaesthetics depressed ventricular function. During reperfusion, ventricular function and coronary flow in both halothane groups were significantly lower than those in the control group. Myocardial ATP concentrations in the 1.0 MAC of enflurane and isoflurane groups were significantly higher than those in the control group. We conclude that halothane had more depressant effects than the other anaesthetics and that enflurane and isoflurane may enhance metabolic recovery in the ischaemic working rat heart.

Topics: Adenosine Triphosphate; Anesthetics, Inhalation; Animals; Cardiac Output; Coronary Circulation; Enflurane; Ethers; Glycogen; Halothane; Heart; Heart Rate; Isoflurane; Male; Methyl Ethers; Myocardial Ischemia; Random Allocation; Rats; Rats, Wistar; Sevoflurane; Ventricular Function

Preconditioning in the setting of global ischemia, using functional recovery during reperfusion as the endpoint, has recently been demonstrated in the isolated perfused rat heart. It has been suggested that its beneficial actions have a metabolic basis. The isolated rat heart has not been fully characterized with respect to the metabolic, functional, and structural changes associated with this phenomenon in the setting of global ischemia. The purpose of this study was to determine (1) the time course of protection conferred by a single episode (5 minutes) of preconditioning; (2) changes in tissue high energy phosphates, lactate, and glycogen levels at different time intervals; and (3) morphological appearance of the heart at the end of ischemia as well as after reperfusion.. Isolated perfused working rat hearts were used. Preconditioning consisted of a single episode of 5 minutes of global ischemia and 15 minutes of reperfusion. Preconditioned and non-preconditioned hearts were subjected to global ischemia of 20-35 minutes duration. Functional recovery, energy metabolism (high energy phosphates, lactate, and glycogen), and structural appearance were studied at different stages.. The functional recovery of the preconditioned hearts was significantly higher than in the corresponding nonpreconditioned group during reperfusion for all durations of ischemia longer than 25 minutes. The degree of protection observed was less than reported previously. A minor degree of energy sparing was reflected by differences in the rate of depletion of glycogen and accumulation of tissue lactate during the sustained episode of ischemia. Semiquantitative light microscopy evaluation revealed that ischemia-induced structural damage was less in the preconditioned hearts, both at the end of the sustained ischemic episode as well as after reperfusion.. A single episode of global ischemia successfully preconditions the isolated working rat heart. The protection elicited was demonstrated on a functional and structural level, and was accompanied by a small energy-sparing effect.

Topics: Animals; Chromatography, High Pressure Liquid; Energy Metabolism; Glycogen; Heart; Heart Arrest; In Vitro Techniques; Lactates; Lactic Acid; Male; Microscopy, Electron; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Phosphates; Rats; Rats, Wistar

Glycogen is consumed during ischemic preconditioning and synthesized during the subsequent period of ischemic tolerance. To better understand this sequence, we examined the effect of brief coronary artery occlusions on regional myocardial glycogen metabolism in intact, anesthetized rats. Sequential 2-min periods of left coronary artery occlusion reduced the glycogen concentration of the anterior left ventricle approximately 30% relative to the posterior region. During subsequent reperfusion, the activity of the physiologically active glycogen synthase I form of glycogen synthase increased threefold in the anterior region (0.58 +/- 0.11 vs. 0.18 +/- 0.08 mumol.g-1.min-1, P < 0.01), stimulating a similar regional increase in glycogen synthesis rate (0.24 +/- 0.04 vs. 0.08 +/- 0.03 mumol.g-1.min-1, P < 0.01). These events were preceded by a rise in regional glucose 6-phosphate concentration, which increased the activity of a myocardial glycogen synthase phosphatase. In diabetic rats glycogen synthase phosphatase activity was significantly lower, and postischemic glycogen synthase activation was significantly impaired. These data suggest the operation of a feedback loop in which transient ischemia leads to a glucose 6-phosphate-mediated increase in the activity of a phosphoprotein phosphatase active toward glycogen synthase. This suggests phospho-protein phosphatase activation may be a feature of the preconditioned myocardium.

Topics: Animals; Coronary Vessels; Diabetes Mellitus, Experimental; Enzyme Induction; Glucose; Glucose-6-Phosphate; Glucosephosphates; Glycogen; Glycogen Synthase; Glycogen-Synthase-D Phosphatase; Kinetics; Male; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Rats; Rats, Sprague-Dawley; Reference Values; Regression Analysis; Time Factors

A spin-echo method is presented for obtaining high resolution, 13C coupled, proton spectra of lactate and alanine in intact, beating rat hearts. All hearts were depleted of glycogen prior to prolonged perfusion with either 10 mM unenriched glucose or [1-13C]glucose to restore glycogen. These two groups of hearts were then examined by 1H NMR during prolonged global (zero flow) or low pressure (low flow) ischemia. During global ischemia, lactate was derived from both glucose and glycogen, with endogenous glycogen contributing twice as much lactate as exogenous glucose. During low perfusion pressure ischemia, however, lactate was derived exclusively from exogenous glucose. The entire pool of lactate (both 12C and 13C) was visible by NMR in intact, glucose perfused hearts while alanine was not detected. However, upon adding 10 mM pyruvate to the perfusate, the entire alanine pool became NMR visible while some of the lactate became NMR invisible. These observations indicate that the NMR visibility of small, usually highly mobile metabolites such as alanine and lactate is not always 100% in intact hearts and that the NMR visibility of these molecules may depend upon which exogenous substrate is presented to the heart.

Topics: Alanine; Animals; Glycogen; Heart; Isoproterenol; Lactates; Lactic Acid; Magnetic Resonance Spectroscopy; Male; Myocardial Ischemia; Myocardium; Perfusion; Rats; Rats, Sprague-Dawley

Limitation of myocardial injury and infarction has been demonstrated by interventions such as ischemic preconditioning or the use of pyruvate as a substrate, which reduces glycogen content before, and acidosis during, ischemia. An isolated perfused rat heart model of global ischemia was employed to test the hypothesis that glycogen depletion reduces ischemic injury as measured by creatine kinase release. 31P-nuclear magnetic resonance spectroscopy was used to measure high-energy phosphates (ATP and phosphocreatine), phosphomonoesters (PME), and intracellular pH. Compared with control glucose-perfused hearts with normal glycogen content (1.49 +/- 0.13 mg Glc/g wet wt), glycogen-depleted pyruvate, ischemic preconditioned, and glycogen-depleted glucose hearts all had reduced glycogen content before ischemia (0.62 +/- 0.16, 0.81 +/- 0.10, and 0.67 +/- 0.12 mg Glc/g wet wt, respectively; P = 0.003) and significantly higher pH at the end of ischemia (5.85 +/- 0.02, 6.33 +/- 0.06, 6.24 +/- 0.04, and 6.12 +/- 0.02 in control, glycogen-depleted pyruvate, preconditioned, and glycogen-depleted glucose-perfused hearts, respectively; P < 0.01), although acidification during the initial phase of ischemia was differentially affected by the three interventions. Glycogen-depleted pyruvate and preconditioned hearts had reduced PME accumulation, greater recovery of function and phosphocreatine, and lower creatine kinase release on reperfusion, whereas glycogen-depleted glucose-perfused hearts were similar to control hearts. In summary, glycogen depletion by these three methods limits the fall in pH during global ischemia, although glycogen depletion in the absence of preconditioning does not limit ischemic injury.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenosine Triphosphate; Animals; Creatine Kinase; Disease Models, Animal; Glucose; Glycogen; Glycolysis; Hydrogen-Ion Concentration; In Vitro Techniques; Intracellular Fluid; Magnetic Resonance Spectroscopy; Male; Myocardial Ischemia; Perfusion; Phosphates; Phosphocreatine; Rats; Rats, Sprague-Dawley; Ventricular Function, Left

Various methods for extraction and isolation of myocardial glycogen show different yields and identify different glycogen subsets. The aim of the present study was to identify a glycogen fraction exposed to changes during myocardial ischaemia. Endomyocardial biopsies from 10 pigs were sampled before cardioplegia, after cardioplegic arrest, and after reperfusion. Glycogen yields were compared following five extraction procedures: (1) hot alkaline tissue digestion, (2) homogenization in perchloric acid and subsequent determination in homogenate, (3) homogenization in perchloric acid and subsequent determination in supernatant, (4) homogenization in perchloric acid and subsequent determination in the precipitate redissolved in hot alkaline and (5) homogenization in homogenisation buffer with lysating capacity. Glycogen was isolated on filter-paper and determined enzymatically. Hot alkaline tissue digestion yielded the highest glycogen amounts (63.5 +/- 18.3 nmol/mg wet weight). Glycogen yields in perchloric homogenate and supernatant were 51%, perchloric precipitate 47%, and buffer 30% of these obtained with hot alkaline. Glycogen yields in hot alkaline were comparable to the sum of those obtained in perchloric supernatant ("acid extractable glycogen") and redissolved precipitate ("heavily extracted glycogen") confirming that glycogen yields obtained with hot alkaline digestion represent "total glycogen". Acid extractable glycogen showed superior analytical characteristics compared with the other methods. Acid extractable glycogen demonstrated a consistent decrease during ischaemia whereas total glycogen and glycogen extracted in homogenization buffer tended to decrease. Glycogen in perchloric precipitate remained unchanged during ischaemia. These findings support a revival of the concept that tissue contains two forms of glycogen. Decreases in myocardial glycogen content during myocardial ischaemia are best observed with acid extractable glycogen.

Topics: Animals; Evaluation Studies as Topic; Female; Glycogen; Heart Arrest, Induced; Male; Methods; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Perchlorates; Swine

An increase in intracellular Na+ during ischaemia has been associated with myocardial injury. In this study, we determined whether inhibition of Na+/K+ ATPase activity contributes to this increase and whether Na+/K+ ATPase activity can be maintained by provision of glucose to perfused rat hearts during low flow, 0.5 ml/min, ischemia. We used 31P NMR spectroscopy to determine changes in myocardial energetics and intracellular and extracellular volumes. 23Na NMR spectroscopy, with DyTTHA3- present as a shift reagent, was used to measure changes in intracellular Na+ and 87Rb NMR spectroscopy was used to estimate Na+/K+ ATPase activity from Rb+ influx rates, Rb+ being an NMR-sensitive congener of K+. In hearts provided with 11 mM glucose throughout ischemia, glycolysis continued and ATP was twofold higher than in hearts without glucose. In the glucose-hearts, Rb+ influx rate was threefold higher, intracellular Na+ was fivefold lower at the end of ischemia and functional recovery during reperfusion was twofold higher. We propose that continuation of glycolysis throughout low flow ischemia allowed maintenance of sufficient Na+/K+ ATPase activity to prevent the increase in intracellular Na+ that would otherwise have led to myocardial injury.

Topics: Animals; Energy Metabolism; Glucose; Glycogen; Glycolysis; Hydrogen-Ion Concentration; In Vitro Techniques; Lactates; Lactic Acid; Magnetic Resonance Spectroscopy; Male; Myocardial Contraction; Myocardial Ischemia; Myocardium; Phosphorus; Rats; Rats, Wistar; Rubidium; Rubidium Radioisotopes; Sodium; Sodium-Potassium-Exchanging ATPase

Although Ca2+ is an important regulator of energy metabolism, the effects of increasing extracellular [Ca2+] on energy substrate preference are not clear. We determined the relationship between [Ca2+], fatty acids, and ischemia on rates of glycolysis, glucose oxidation, and palmitate oxidation in isolated working rat hearts. Hearts were perfused with Krebs-Henseleit buffer containing 11 mM glucose, 100 microU/mL insulin, and either 1.25 or 2.5 mM Ca2+, in the presence or absence of 1.2 mM palmitate. Rates of glycolysis and glucose oxidation or palmitate oxidation were measured in the hearts using [5-3H,14C(U)]glucose or [1-14C]palmitate, respectively. In the absence of fatty acids, glycolysis and glucose oxidation rates were similar, regardless of whether [Ca2+ was 1.25 or 2.5 mM. Addition of 1.2 mM palmitate to the perfusate of hearts perfused with 1.25 mM Ca2+ significantly decreased rates of both glycolysis (from 4623 +/- 438 to 1378 +/- 238 nmol.min-1.g-1 dry weight) and glucose oxidation (from 1392 +/- 219 to 114 +/- 22 nmol.min-1.g-1 dry weight). When [Ca2+] was increased from 1.25 to 2.5 mM in hearts perfused with 1.2 mM palmitate, glycolysis and glucose oxidation increased by 164 and 271%, respectively, with no change in palmitate oxidation rates. Increasing [Ca2+] from 1.25 to 2.5 mM increased the contribution of glucose to ATP production from 9.3 to 18.7%. When hearts were subjected to low-flow ischemia (by reducing coronary flow to 0.5 mL.min-1) oxidative metabolism was essentially abolished. Under these conditions, glycolytic rates were not dependent on either [Ca2+] or the presence or absence of fatty acids. These results demonstrate that perfusate [Ca2+] is an important determinant of myocardial glucose metabolism in aerobic hearts, and that glycolysis and glucose oxidation are more responsive to changes in [Ca2+] than is fatty acid oxidation.

Topics: Adenosine Triphosphate; Aerobiosis; Animals; Calcium; Fatty Acids; Glucose; Glycogen; Glycolysis; In Vitro Techniques; Male; Myocardial Contraction; Myocardial Ischemia; Myocardium; Oxidation-Reduction; Palmitic Acids; Perfusion; Rats; Rats, Sprague-Dawley

Working rat hearts perfused with 5.5 mM glucose were submitted to a 10-min period of no-flow ischaemia or anoxia. Both conditions stimulated glycogenolysis, activated phosphorylase and increased cyclic GMP content, although the time course of these changes differed in anoxia and ischaemia. Changes in cyclic GMP content were not correlated with glycogenolysis or phosphorylase activation. Perfusion with 1 microM L-nitroarginine methylester, an inhibitor of nitric oxide synthase, decreased cGMP concentration under normoxic conditions and abolished the ischaemia-induced increase in cGMP. The inhibitor decreased the coronary flow without affecting the overall working performance of the hearts under normoxic conditions.

Topics: Adenosine Triphosphate; Amino Acid Oxidoreductases; Animals; Arginine; Coronary Circulation; Cyclic GMP; Fructosediphosphates; Glucose; Glycogen; Heart; Hexosephosphates; Hypoxia; In Vitro Techniques; Kinetics; Male; Myocardial Ischemia; Myocardium; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Perfusion; Rats; Rats, Wistar; Time Factors

The aim was to assess the abilities of exogenous noradrenaline, isoprenaline, and phenylephrine to precondition the isolated rat heart against ischaemic and reperfusion injury.. The isovolumetric Langendorff rat heart model was used to determine postischaemic recovery of left ventricular function. The hearts were subjected to 30 min of normothermic global ischaemia followed by 30 min reperfusion. Treated hearts were perfused with noradrenaline (10(-7) M), isoprenaline (10(-8) M), or phenylephrine (10(-6) M, 10(-5) M, and 10(-4) M) for 5 min followed by 5 min washout before the 30 min ischaemic period.. Control hearts recovered 47.6(SEM 4.3)% of baseline heart rate x developed pressure after 30 min reperfusion, whereas noradrenaline and isoprenaline treated hearts recovered 75.1(4.6) and 76.4(4.6)%, respectively (p < 0.001 v control). Left ventricular end diastolic pressures at the end of reperfusion were 48.8(4.0), 20.0(2.4), and 21.6(2.7)mm Hg for control, noradrenaline treated (p < 0.001 v control), and isoprenaline treated (p < 0.001 v control) hearts respectively. beta Blockade with propranolol during noradrenaline treatment blocked the protective effects. No concentration of phenylephrine used was able to enhance postischaemic heart rate x developed pressure significantly, or result in improved (lower) postischaemic left ventricular end diastolic pressure. During treatment with noradrenaline and phenylephrine (10(-5) M), lactate release was 13.0(1.0) and 11.0(0.9) mumol.5 min-1, respectively (p = NS); these values were significantly (p < 0.001) greater than baseline value of 3.7(0.5) mumol.5 min-1. Immediately before the 30 min ischaemic period, control and phenylephrine treated groups had glycogen levels of 132(14) and 128(5) nmol.mg-1 protein, respectively (p = NS), whereas the glycogen content of the noradrenaline treated group was only 96(5) nmol.mg-1 protein (p < 0.05 v control and phenylephrine treated).. Transient beta adrenergic but not alpha 1 adrenergic stimulation can precondition the isolated perfused rat heart. The mechanism of protection may, at least in part, be due to transient demand ischaemia. Partial depletion of glycogen following treatment may play a role in the observed protective effects.

Topics: Adrenergic beta-Agonists; Animals; Arrhythmias, Cardiac; Glycogen; Heart; Isoproterenol; Male; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Norepinephrine; Perfusion; Phenylephrine; Rats; Rats, Sprague-Dawley; Stimulation, Chemical

It has recently been shown that hypoxia and ischemia are equally effective to precondition the myocardium of the rat. A comparison of the metabolic changes caused by transient ischemia and hypoxia has not yet been made and may help to elucidate the metabolic factors involved in eliciting preconditioning. The aim of this study was to compare the changes in tissue high energy phosphates, glycogen and lactate during and after hypoxic and ischemic preconditioning in isolated perfused rat hearts. Isolated rat hearts were subjected to global ischemia of 30 minutes duration, with and without preconditioning consisting of a single episode of 5 minutes global ischemia or hypoxia (PO2 = 12kPa). The post-ischemic recovery of aortic flow of the nonpreconditioned group was significantly less than that of the two preconditioned groups: 0.5 +/- 0.5 ml/min vs. 23.3 +/- 3.4 and 20.7 +/- 3.6 ml/min for ischemic and hypoxic preconditioning respectively. The only common metabolic factor between the two preconditioned groups was the similar extent of glycogenolysis after transient ischemia or hypoxia: glycogen decreased from 22 +/- 0.8 in non-preconditioned hearts to 16 +/- 0.5 and 16 +/- 1.5 mumoles glucose per g wet tissue in ischemic and hypoxic preconditioned hearts respectively. There was also no difference in lactate production between the two groups during the sustained episode of ischemia. We conclude that oxygen deprivation, rather than other metabolic factors, is the important factor in eliciting preconditioning.

Topics: Animals; Coronary Circulation; Glycogen; Hypoxia; Lactates; Male; Models, Biological; Myocardial Ischemia; Perfusion; Phosphates; Rats; Rats, Wistar; Reperfusion

The role of low-dose aspirin (3 mg/kg, i.v.) in attenuating ischemic reperfusion injury was studied in a canine model. Regional ischemia for 40 min was produced by temporary occlusion of the left anterior descending coronary artery and thereafter reperfusion instituted for 3 h. Mean arterial pressure (MAP), heart rate (HR), left ventricular end diastolic pressure (LVEDP), positive (+) LV dP/dtmax and negative (-) LV dP/dtmax were monitored along with myocardial adenosine triphosphate (ATP), creatine phosphate (CP), glycogen and lactate. Following reperfusion, there was a significant fall in (i) MAP, (ii) (+) LV dP/dtmax and (iii) (-) LV dP/dtmax. LVEDP was corrected after about 2 h of reperfusion. Replenishment of only myocardial CP occurred, without any change in ATP and glycogen, although lactate accumulation was corrected. Aspirin administered 15 min before reperfusion (post-treatment) caused normalisation of LVEDP within 15 min and prevented any deterioration in (-) LV dP/dtmax, although it had no effect on MAP and (+) LV dP/dtmax. After 3 h of reperfusion (post-treatment), myocardial ATP, CP, glycogen and lactate contents became normal. The number of premature ventricular complexes was significantly reduced after aspirin treatment. The present study indicates that low-dose aspirin post-treatment can ameliorate at least some of the deleterious consequences of reperfusion injury of the myocardium.

Topics: Adenosine Triphosphate; Analysis of Variance; Animals; Aspirin; Blood Pressure; Creatinine; Disease Models, Animal; Dogs; Glycogen; Heart Rate; Injections, Intravenous; Lactates; Lactic Acid; Myocardial Ischemia; Myocardial Reperfusion Injury

This study evaluated the importance of ATP-dependent potassium channels (KATP) for ischemic preconditioning (IP) in swine. Swine were studied because due to the sparsity of their innate collateral circulation infarct size (IS) development closely resembles that observed in humans. Ninety minutes of ischemia at a blood flow reduction sufficient to reduce regional myocardial work by 90% caused 13.2 +/- 8.9% (SD) IS of the area at risk. A single cycle of 10-min preconditioning ischemia followed by 15-min reperfusion reduced IS after 90 min of ischemia to 2.8 +/- 2.7%. The epicardial monophasic action potential duration at 50% repolarization (MAP50) was decreased more markedly during the initial 10 min of the prolonged ischemia than during the first 10 min of the preconditioning ischemic period (84 +/- 4 vs. 89 +/- 2%). Transmural myocardial adenosine (ADO) uptake was reversed to net release during both ischemic periods and during the initial phase of reperfusion. Glibenclamide (0.5 mg/kg, followed by 50 micrograms/min i.v.) abolished the reduction in MAP50 without altering ADO release. Glibenclamide did not alter IS per se (13.0 +/- 7.6%) but abolished the beneficial effect of IP (IS: 13.6 +/- 6.2%). Thus blockade of KATP with glibenclamide abolishes the IS-reducing effect of IP in swine but does not reduce ADO release.

Topics: Action Potentials; Adenosine Triphosphate; Animals; Blood Glucose; Coronary Circulation; Coronary Vessels; Female; Glyburide; Glycogen; Heart; Heart Rate; Humans; Male; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Phosphocreatine; Potassium Channels; Regression Analysis; Swine; Swine, Miniature; Time Factors; Ventricular Function, Left

The efficacy of increasing glycolysis during ischemia for enhancing the salutary effects of reperfusion was evaluated in isolated perfused rabbit hearts subjected to low-flow ischemia followed by reperfusion. Control hearts were perfused with buffer containing 0.4 mM palmitate, 5 mM glucose, and 70 mU/l insulin. Additional groups of hearts were perfused with double glucose/insulin and 1 mM dichloroacetate or were subjected to substrate priming to increase preischemic glycogen content. Ischemic contracture was completely prevented in hearts perfused with high glucose/insulin and was delayed markedly by either dichloroacetate or enhanced preischemic glycogen [45 +/- 14 and 31 +/- 20 min, respectively; P < 0.01 each vs. control (11 +/- 10 min)] and inversely related to the rate of lactate production. With reperfusion, recovery of developed pressure was 56 +/- 23% of baseline in control hearts, 90 +/- 8% in hearts receiving high glucose/insulin, 92 +/- 5% in hearts receiving dichloroacetate, and 79 +/- 19% in hearts with increased glycogen (P < 0.05 each vs. control hearts). Creatine kinase release was reduced by > 55% in treated hearts. Thus enhancement of glycolysis by diverse mechanisms during ischemia decreased ischemic damage and improved the recovery of contractile function with reperfusion.

Topics: Animals; Biomarkers; Blood Pressure; Body Water; Dichloroacetic Acid; Energy Metabolism; Fatty Acids, Nonesterified; Glucose; Glycogen; Glycolysis; Heart; Heart Rate; In Vitro Techniques; Insulin; Lactates; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Oxygen Consumption; Rabbits; Time Factors; Triglycerides; Ventricular Function, Left

The purpose of this study was to determine whether metabolites produced by glycolysis during ischemia significantly contribute to myocardial injury of hypertrophied hearts. The accumulation of glycolytic metabolites during ischemia was reduced by means of glycogen reduction or by treatment with the glycolytic inhibitor, 2-deoxy-D-glucose (2-DG) before ischemia. Hearts from aortic-banded (Band) and sham-operated (Sham) rats (8 wk postop) were isolated, perfused with Krebs buffer, and had a left ventricular (LV) balloon to measure developed pressure. A 15-min perfusion with hypoxic buffer (glycogen reduction, GR) or a 10-min perfusion with 10 mM 2-DG (glycolytic inhibition) was followed by 25 min global, normothermic, no-flow ischemia and 30 min normoxic reperfusion. Heart weights were greater in Band than Sham [2.76 +/- 0.06 vs. 1.5 +/- 0.04 (mean +/- SE) g; P < 0.001]. GR and 2-DG each resulted in reduced ATP levels measured at the beginning of ischemia in both Band and Sham groups compared with untreated groups, but there were no differences among groups after 25 min of ischemia. Myocardial lactate levels at the end of ischemia were significantly reduced in both Band and Sham hearts with GR or 2-DG compared with untreated controls. Recovery of LV function after ischemia and reperfusion was significantly improved in Band after GR (206% increase) and after 2-DG treatment (126% increase) compared with their respective untreated controls. Diastolic dysfunction during reperfusion was ameliorated in Band by preischemic GR but not by 2-DG treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Cardiomegaly; Deoxyglucose; Diastole; Glycogen; Glycolysis; Male; Myocardial Ischemia; Myocardial Reperfusion; Rats; Rats, Sprague-Dawley; Ventricular Function, Left

Functional recovery following ischemia and reperfusion in the isolated working rat heart perfused with glucose (11 mM) was examined in relation to pre- and postischemic levels of ATP, glycogen, glucose 6-phosphate, and the lactate-to-pyruvate ratio. The following variables were studied: feeding and fasting in vivo, addition of L-lactate (10 mM), dl-beta-hydroxybutyrate (10 mM), glucagon (0.01 and 1 micrograms/ml), and a 15-min anoxic perfusion before ischemia in vitro. Recovery was assessed as the percentage of preischemic power. Good correlation was found between functional recovery and the postischemic content of glycogen. Glycogen depletion by anoxia or glucagon before ischemia impaired recovery. There was no relationship among lactate produced, or the lactate-to-pyruvate ratio, and recovery. The addition of lactate or beta-hydroxybutyrate to hearts from fed rats increased the content of glycogen and glucose 6-phosphate, whereas addition of lactate, but not beta-hydroxybutyrate, improved recovery. There was a linear relationship between glycogen content and glucose 6-phosphate levels. In conclusion, the degree of return of oxidative metabolism and of net glycogen resynthesis reflects postischemic recovery of function. The results also suggest a role for anaplerosis of the citric acid cycle as an additional determinant of postischemic recovery.

Topics: 3-Hydroxybutyric Acid; Adenosine Triphosphate; Animals; Fasting; Glucose-6-Phosphate; Glucosephosphates; Glycogen; Heart; Hydroxybutyrates; Hypoxia; In Vitro Techniques; Lactates; Lactic Acid; Male; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Pyruvates; Pyruvic Acid; Rats; Time Factors

The level of systemic hypoxia required to alter neonatal myocardial metabolism and its resultant effect on tolerance to global ischemia is unknown. This study examines myocardial purine nucleotides, glycogen (MG), lactate, creatine phosphate (CP) and the subsequent tolerance to ischemia in hearts exposed to varying levels of hypoxia (2 h). Three-day-old swine were randomly allocated into five study groups. Animals were anaesthetized and ventilated (2 h) with varying mixtures of medical air and nitrogen to achieve their target PaO2 (mmHg): normoxia (PaO2 = 80, n = 18), mild (PaO2 = 60, n = 10), moderate (PaO2 = 40, n = 12), moderately-severe (PaO2 = 30, n = 7) and severe (PaO2 = 20, n = 9). Arterial blood gases verified PaO2 and normal PaCO2 (39.5 +/- 0.5 mmHg). Subsequently, the heart was exposed and the metabolic profile determined from a freeze-clamp LV biopsy. The heart was excised and tolerance to ischemia determined by time (min) to ischemic contracture onset (TICo) and peak (TICp). The results demonstrated a tendency to decreased MG with progressive hypoxia which reached significance in severe hypoxia (6.6 +/- 2.7 mumol/g, P < 0.05). Despite a doubling of myocardial lactate with moderately-severe hypoxia, increases only reached significance with severe hypoxia (27.8 +/- 6.3 mumol/g, P < 0.0001). Despite the reduction in LV adenosine triphosphate (ATP) with severe hypoxia (2.16 +/- 0.68 mumol/g, P < 0.05), CP was unaltered.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenosine Diphosphate; Adenosine Monophosphate; Adenosine Triphosphate; Animals; Animals, Newborn; Blood Gas Analysis; Glycogen; Hypoxia; Inosine Monophosphate; Lactates; Male; Myocardial Ischemia; Myocardium; Phosphocreatine; Severity of Illness Index; Swine; Time Factors

The mechanical effects of ischemic contracture may be important in the development of irreversible cellular damage as it increases mechanical stress on sarcolemmal membranes and restricts endocardial perfusion. To assess the relative importance of these mechanical effects compared with decreased energy supply in the development of irreversible injury, the effects of inhibiting ischemic contracture with 2,3-butanedione monoxime (BDM), an agent that disrupts excitation-contraction coupling, were delineated in isovolumically contracting isolated rabbit hearts. Administration of 20 mmol/L BDM in 12 hearts subjected to 60 minutes of low-flow ischemia prevented ischemic contracture (left ventricular end-diastolic pressure [LVEDP], 12 +/- 3 compared with 48 +/- 14 mm Hg in 20 control hearts; P < .001), reduced membrane damage (creatine kinase [CK] release, -54% compared with control hearts; P < .05), and enhanced functional recovery during reperfusion (left ventricular developed pressure [LVDP], 86 +/- 10% of baseline compared with 56 +/- 23% in control hearts; P < .01). These observations were not related to increased intracavitary pressure and its effects on flow distribution, since venting the left ventricle in additional hearts did not result in improved function during reperfusion. Although it would be tempting to conclude that BDM protected ischemic myocardium by preventing ischemic contracture, administration of BDM was also associated with reduced depletion of ATP during ischemia, perhaps related to diminished energy demand. To distinguish between the relative importance of inhibiting contracture from provision of adequate energy, the period of ischemia was extended to 120 minutes. BDM still prevented ischemic contracture (LVEDP, 10 +/- 6 mm Hg) and preserved ATP stores, but it did not prevent membrane damage (CK release, 483 +/- 254 U/g dry weight) or contractile failure during reperfusion (LVDP, 68 +/- 7% of baseline). In contrast, increasing the rate of anaerobic glycolysis during ischemia by doubling glucose and insulin in the presence of BDM markedly decreased membrane damage (CK release, 114 +/- 72 U/g dry weight; P < .05) and contractile failure during reperfusion (LVDP, 88 +/- 7% recovery of baseline; P < .01). These results suggest that insufficient energy production is primarily responsible for myocardial ischemic damage, whereas mechanical effects of ischemic contracture appear to play only a minor role.

Topics: Animals; Creatine Kinase; Diacetyl; Energy Metabolism; Glucose; Glycogen; Lactates; Lactic Acid; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Myocardial Reperfusion Injury; Phosphocreatine; Potassium; Rabbits; Ventricular Function, Left

Effects of iloprost, which is a stable prostacyclin analogue, on the ischemic myocardium were examined in the open-chest dog heart in terms of biochemical parameters. Ischemia was initiated by ligating the left anterior descending coronary artery. When the coronary artery was ligated for 3 min, the levels or glycogen, fructose-1,6-diphosphate (FDP), adenosine triphosphate and creatine phosphate decreased, and the levels of glucose-6-phosphate (G6P), fructose-6-phosphate (F6P), lactate, adenosine diphosphate and adenosine monophosphate increased. During ischemia, therefore, energy charge potential was significantly decreased from 0.89 +/- 0.01 to 0.82 +/- 0.01, and ([G6P] + [F6P])/[FDP] and [lactate]/[pyruvate] ratios were significantly increased from 1.75 +/- 0.30 to 29.05 +/- 5.70 and 13 +/- 3 to 393 +/- 112, respectively. Iloprost (0.1, 0.3, or 1 microgram.kg-1) was injected intravenously 5 min before the onset of ischemia. Iloprost (0.1, 0.3, and 1 micrograms.kg-1) reduced the ischemia-induced decrease in energy charge potential to 94, 74, and 86%, respectively, the increase in ([G6P] + [F6P])/[FDP] to 38, 29, 32%, respectively, and the increase in [lactate]/[pyruvate] to 67, 45, 65%, respectively. These results suggest that iloprost lessens the myocardial metabolic derangements produced by ischemia, and the most potent effect was obtained at the dose of 0.3 microgram.kg-1.

Topics: Adenosine Diphosphate; Animals; Dogs; Energy Metabolism; Female; Fructosediphosphates; Glucose-6-Phosphate; Glucosephosphates; Glycogen; Heart; Hemodynamics; Iloprost; Lactates; Male; Myocardial Ischemia; Phosphocreatine

We studied the effects of the potent Na/H exchange inhibitor methylisobutyl amiloride (MIA, 1 microM) on post-ischemic ventricular recovery and energy metabolic status in spontaneously contracting, isolated rat and guinea-pig hearts subjected to 45 min zero-flow ischemia followed by reperfusion. For both species, MIA was added either 15 min prior to ischemia and was present throughout reperfusion or was added at the time of reperfusion only. In control rat hearts, force recovery after 30 min of reperfusion was 25.6 +/- 6.0% of the pre-ischemic value whereas in hearts pre-treated with MIA recovery was enhanced to 55.4 +/- 9% (P < 0.05). Elevation of resting tension during the first 20 min of reperfusion was also significantly reduced by MIA pre-treatment. When MIA was added at the time of reperfusion only, recovery was generally lower than that seen with MIA pre-treatment although significantly higher values were seen through much of the reperfusion period. In rat hearts, MIA reduced the time required for return to sustained contractile recovery particularly in those hearts where the drug was added prior to ischemia (control, 11.4 +/- 2.7 min; MIA, 2.6 +/- 0.5 min, P < 0.05). Similar effects of MIA pre-treatment were seen in guinea-pig hearts in terms of contractile recovery, time to recovery and reduction in resting tension although MIA addition at the time of reperfusion was without beneficial effect either on the magnitude of contractile recovery or time required for restoration of function. In guinea-pig hearts, recovery of function was accompanied by substantial bradycardia. However, maintenance of ventricular rate through electrical pacing exerted no significant influence on the protective effects of MIA pre-treatment. There was no effect of MIA on energy metabolites in reperfused rat hearts or paced guinea-pig hearts, although in spontaneously contracting guinea-pig hearts improved recovery of function was associated with significantly higher levels of high energy phosphates. No effects of tissue metabolites were seen in ischemic non-reperfused hearts irrespective of treatment. The protective effects of MIA were not related to diminished release of creatine kinase during reperfusion. Our results demonstrate marked protective effects of MIA, on the reperfused rat and guinea-pig myocardium. These studies also demonstrate, for the first time, that the effects of amiloride analogues are not species specific and further support the concept that Na/H exch

Topics: Adenine Nucleotides; Amiloride; Animals; Creatine Kinase; Glycogen; Guinea Pigs; Heart; Heart Rate; Hydrogen-Ion Concentration; In Vitro Techniques; Ion Transport; Lactates; Male; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Phosphocreatine; Rats; Rats, Sprague-Dawley; Sodium-Hydrogen Exchangers

1. The benzoylguanidine derivative Hoe 694 ((3-methylsulphonyl-4- piperidino-benzoyl) guanidine methanesulphonate) was characterized as an inhibitor of Na+/H+ exchange in rabbit erythrocytes, rat platelets and bovine endothelial cells. The potency of the compound was slightly lower or comparable to ethylisopropyl amiloride (EIPA). 2. To investigate a possible cardioprotective role of the Na+/H+ exchange inhibitor Hoe 694, rat isolated working hearts were subjected to ischaemia and reperfusion. In these experiments all untreated hearts suffered ventricular fibrillation on reperfusion. Addition of 10(-7) M Hoe 694 to the perfusate almost abolished reperfusion arrhythmias in the rat isolated working hearts. 3. Hoe 694 reduced the release of lactate dehydrogenase (LDH) and creatine kinase (CK), which are indicators of cellular damage during ischaemia, into the venous effluent of the hearts by 60% and 54%, respectively. 4. The tissue content of glycogen at the end of the experiments was increased by 60% and the high energy phosphates ATP and creatine phosphate were increased by 240% and 270% respectively in the treated hearts as compared to control hearts. 5. Antiischaemic effects of the Na+/H+ exchange inhibitor, Hoe 694, were investigated in a second experiment in anaesthetized rats undergoing coronary artery ligation. In these animals, pretreatment with Hoe 694 caused a dose-dependent reduction of ventricular premature beats and ventricular tachycardia as well as a complete suppression of ventricular fibrillation down to doses of 0.1 mg kg-1, i.v. Blood pressure and heart rate remained unchanged. 6. We conclude that the new Na+/H+ exchange inhibitor, Hoe 694, shows cardioprotective and antiarrhythmic effects in ischaemia and reperfusion in rat isolated hearts and in anaesthetized rats. In view of the role which Na+/H+ exchange seems to play in the pathophysiology of cardiac ischaemia these effects could probably be attributed to Na+/H+ exchange inhibition.

Topics: Adenosine Triphosphate; Amiloride; Anesthesia; Animals; Blood Platelets; Cattle; Endothelium, Vascular; Erythrocyte Membrane; Female; Glycogen; Guanidines; In Vitro Techniques; Ion Exchange; Male; Myocardial Ischemia; Phosphocreatine; Potassium; Rabbits; Rats; Rats, Sprague-Dawley; Rats, Wistar; Sodium; Sulfones

1. In isolated Langendorff-perfused, electrically-paced, hearts of guinea-pigs, global low-flow-ischaemia (LFI; at 0.7 ml min-1) resulted in marked increases in the rates of release of lactate, lactate dehydrogenase (LDH) and creatine kinase (CK) over a 30 min period. At the end of the LFI period, tissue ATP content was significantly reduced from a control value of 11.8 +/- 0.8 (5) to 5.6 +/- 0.8 (5) mumol g-1 dry weight. 2. The presence of ranolazine [(+/-)-N-(2,6-dimethyl-phenyl)-4[2-hydroxy-3-(2-methoxy-phenoxyl)- propyl] - l-piperazine acetamide dihydro-chloride; RS-43285-193] at 10 microM, from 20 min prior to and during LFI, resulted in significant reductions in the release of lactate, LDH and CK during the ischaemic period and a significant preservation of tissue ATP (9.0 +/- 1.1 (6) mumol g-1 dry wt.). Ranolazine did not prevent the reductions in creatine phosphate or glycogen observed in LFI, nor did it have any significant effects on any contractile parameters before or during the LFI period. 3. Neither ranolazine nor LFI affected the total amounts of tissue pyruvate dehydrogenase (PDH) activity; however, the significant reduction in the amount of active, non-phosphorylated PDH caused by LFI (from 88.2 +/- 5.5 to 44.2 +/- 3.2% of total activity) was partially but significantly prevented by ranolazine (67.2 +/- 6.8%). This effect of ranolazine on PDH may be part of the mechanism whereby the compound reduces lactate release and preserves tissue ATP during ischaemia.

Topics: Acetanilides; Adenosine Triphosphate; Angina Pectoris; Animals; Creatine Kinase; Female; Glycogen; Guinea Pigs; Heart; In Vitro Techniques; L-Lactate Dehydrogenase; Lactates; Lactic Acid; Myocardial Ischemia; Myocardium; Perfusion; Phosphocreatine; Piperazines; Pyruvate Dehydrogenase Complex; Ranolazine

Although previous investigators have demonstrated that myocardial preconditioning reduces infarct size, the mechanisms of cardioprotection associated with preconditioning are not completely understood.. To test the hypothesis that preconditioning (four 5-minute episodes of ischemia each followed by 5 minutes of reperfusion) reduces infarct size by depleting cardiac glycogen stores and attenuating the degree of intracellular acidosis during subsequent prolonged left coronary artery occlusion, preconditioned and control rats were subjected to 45 minutes of left coronary artery occlusion and 120 minutes of reflow immediately after preconditioning (groups 1P and 1C, respectively) or after 30 minutes (groups 2P+30m and 2C), 1 hour (groups 3P+60m and 3C), or 6 hours (groups 4P+360m and 4C) of nonischemic recovery after preconditioning but before prolonged ischemia. In each group, cardiectomy was performed in selected rats immediately before prolonged ischemia for cardiac glycogen assay. In selected animals, 31P magnetic resonance spectroscopy was performed to monitor intracellular pH and measure high-energy phosphate levels during ischemia and reperfusion. Group 1P rats demonstrated marked glycogen depletion after preconditioning compared with controls (0.72 +/- 0.39 [n = 9] versus 5.67 +/- 1.73 [n = 12] mg glucose/g wet wt; p < 0.001 versus group 1C) that was associated with attenuation of intracellular acidosis during ischemia, as measured by 31P magnetic resonance spectroscopy (6.8 +/- 0.3 [n = 11] versus 6.2 +/- 0.3 [n = 9] pH units; p < 0.01), and marked infarct size reduction (0.3 +/- 0.6% [n = 7] versus 38.1 +/- 11.3% [n = 7], infarct size divided by risk area; p < 0.0001). During ischemia, there were no differences in myocardial ATP or phosphocreatine levels or in any hemodynamic determinant of myocardial oxygen demand between groups 1P and 1C. In preconditioned rats that were allowed to recover before ischemia (groups 2P+30m, 3P+60m, and 4P+360m), the time course of glycogen repletion paralleled the loss of protection from ischemic injury.. Glycogen depletion and the attenuation of intracellular acidosis during ischemia appear to be important factors in delaying irreversible injury and reducing infarct size in this animal model of myocardial preconditioning.

Topics: Animals; Glycogen; Hemodynamics; Hydrogen-Ion Concentration; Intracellular Membranes; Magnetic Resonance Imaging; Myocardial Infarction; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Phosphates; Rats; Rats, Sprague-Dawley; Recurrence

The structural correlates of 'chronic hibernating myocardium' in man consist of myocardial cells which transformed from a functional state (rich in contractile material) to a surviving state (poor in contractile material, rich in glycogen). Since the calcium-handling organelles such as SR, sarcolemma and mitochondria underwent structural changes in cells so affected, the distribution of calcium was investigated in biopsies obtained from 'hibernating' areas. The material was processed for microscopic localization of total calcium (laser microprobe mass analysis, LAMMA) and of exchangeable calcium (phosphate-pyroantimonate precipitation method, PPA). Subcellular distribution of total calcium as assessed by LAMMA revealed that in the structurally affected cells the areas in which sarcomeres were replaced by glycogen contained significantly more calcium than all other areas probed such as mitochondria, remaining sarcomeres at the cell periphery and subcellular areas of normally structured cells. Calcium precipitate, obtained after PPA assessment, was localized at the sarcolemma but was virtually absent in the mitochondria of affected cells. The high calcium content in the myolytic areas of affected cells most probably belongs to a pool of bound calcium. The observations that calcium is retained at the sarcolemma and that mitochondria are devoid of precipitate favour the hypothesis that cells structurally affected as such are not ischaemic and are still able to regulate their calcium homeostasis.

Topics: Calcium; Glycogen; Humans; Microscopy, Electron; Mitochondria, Heart; Myocardial Ischemia; Myocardium; Sarcoplasmic Reticulum

Topics: Animals; Cardiopulmonary Bypass; Glucose; Glycogen; Heart; Heart Arrest, Induced; Humans; Insulin; Isotonic Solutions; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Potassium; Premedication

Repeated brief episodes of ischaemia "precondition" the myocardium and protect it during a subsequent period of sustained ischaemia. We subjected isolated rat hearts to sustained ischaemia with or without reperfusion after different schedules of preconditioning. We demonstrated that preconditioning with three 5 min periods of ischaemia separated by 10 min periods of reperfusion permits better functional recovery than preconditioning with three 2 min ischaemic periods separated by 10 min of reperfusion. Preconditioned hearts had creatine phosphate and adenine nucleotide levels comparable to those in the aerobically perfused controls, and showed good functional recovery. Although the mechanisms by which preconditioning protects the heart from subsequent ischaemic damage are unclear, we speculate that preservation of mitochondrial function and oxidative energy production is involved.

Topics: Adenine Nucleotides; Animals; Creatine Kinase; Energy Metabolism; Glycogen; Heart; In Vitro Techniques; L-Lactate Dehydrogenase; Lactates; Lactic Acid; Male; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Phosphocreatine; Rats; Rats, Sprague-Dawley; Time Factors

It was examined whether lactate influences postischaemic hemodynamic recovery as a function of the duration of ischaemia and whether changes in high-energy phosphate metabolism under ischaemic and reperfused conditions could be held responsible for impairment of cardiac function. To this end, isolated working rat hearts were perfused with either glucose (11 mM), glucose (11 mM) plus lactate (5 mM) or glucose (11 mM) plus pyruvate (5 mM). The extent of ischaemic injury was varied by changing the intervals of ischaemia, i.e. 15, 30 and 45 min. Perfusion by lactate evoked marked depression of functional recovery after 30 min of ischaemia. Perfusion by pyruvate resulted in marked decline of cardiac function after 45 min of ischaemia, while in glucose perfused hearts hemodynamic performance was still recovered to some extent after 45 min of ischaemia. Hence, lactate accelerates postischaemic hemodynamic impairment compared to glucose and pyruvate. The marked decline in functional recovery of the lactate perfused hearts cannot be ascribed to the extent of degradation of high-energy phosphates during ischaemia as compared to glucose and pyruvate perfused hearts. Glycolytic ATP formation (evaluated by the rate of lactate production) can neither be responsible for loss of cardiac function in the lactate perfused hearts. Moreover, failure of reenergization during reperfusion, the amount of nucleosides and oxypurines lost or the level of high-energy phosphates at the end of reperfusion cannot explain lactate-induced impairment. Alternatively, the accumulation of endogenous lactate may have contributed to ischaemic damage in the lactate perfused hearts after 30 min of ischaemia as it was higher in the lactate than in the glucose or pyruvate perfused hearts. It cannot be excluded that possible beneficial effects of the elevated glycolytic ATP formation during 15 to 30 min of ischaemia in the lactate perfused hearts are counterbalanced by the detrimental effects of lactate accumulation.

Topics: Adenosine; Adenosine Triphosphate; Animals; Energy Metabolism; Glucose; Glycogen; Guanosine Triphosphate; Heart; Hypoxanthine; Hypoxanthines; In Vitro Techniques; Inosine; Inosine Monophosphate; Kinetics; Lactates; Male; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Phosphocreatine; Pyruvates; Rats; Rats, Inbred Lew; Time Factors; Xanthine; Xanthines