Possible mechanisms for reoxygenation-induced recovery of myocardial high-energy phosphates after hypoxia

Changes in several factors responsible for high-energy phosphate production and metabolism in the heart perfused under hypoxic and subsequent reoxygenated conditions were studied using rabbit heart Langendorff preparation. A marked decline in myocardial ATP and creatine phosphate contents was observed with prolonged periods of hypoxia lasting from 15 to 60 min. Upon reoxygenation after 15 or 30 min hypoxia, creatine phosphate levels were fully recovered, whereas ATP contents were partially restored. Possible mechanisms responsible for reoxygenation-induced differential recovery of high-energy phosphate contents were investigated. Mitochondrial function for generating ATP was depressed upon hypoxia for longer than 15 min hypoxia, and the decreased function was found to be irreversible upon reoxygenation even after 15 min hypoxia. However, mitochondrial ability to generate ATP in the heart receiving 60 min hypoxia was still observed to some extent. Creatine phosphokinase activity of the myocardium exposed to hypoxic solution for 60 min showed only 19% depression. A release of creatine phosphokinase from the perfused heart was observed after more than 30 min of hypoxic perfusion or during reoxygenated perfusion after 60 min hypoxia. Changes in creatine phosphokinase activities of the myocardium and of the perfusate were not associated with those in myocardial high-energy phosphate contents. Hypoxia also induced significant release of adenine nucleotide metabolites from the perfused heart in a biphasic manner. Substrates responsible for the release of the metabolites were found to be mainly inosine and partly hypoxanthine. The metabolite release was also supported by our finding of a decrease in total adenine nucleotide contents of the myocardium upon hypoxia. The present results suggested a crucial role of hypoxia-induced release of adenine nucleotide metabolites in a differential recovery of ATP and creatine phosphate upon reoxygenation.     

Effect of adrenochrome on adenine nucleotides and mitochondrial oxidative phosphorylation in rat heart

Effects of adrenochrome, an oxidation product of epinephrine, on myocardial energy production were investigated by studying changes in adenine nucleotide content and mitochondrial oxidative phosphorylation activities in the isolated rat heart. Perfusion of the heart with 50 mg/L adrenochrome induced a marked decline in contractile force within 5 min and this was associated with a rapid decline in the myocardial ATP/AMP ratio. A significant decrease in ATP and ATP/ADP ratio as well as a significant increase in ADP and AMP content was observed at 10 min of perfusion with adrenochrome. Furthermore, mitochondrial oxidative phosphorylation activities were unchanged except that an increase in state 4 respiration and a decrease in RCI value were seen in the heart perfused with adrenochrome for 10 min. Autoradiography of the sections from hearts perfused with 14C-adrenochrome revealed the localization of a significant amount of radioactivity on mitochondria. Adrenochrome at concentrations of 20 mg/L or higher was found to inhibit the oxidative phosphorylation activities of heart mitochondria under in vitro conditions. The depressant effects of adrenochrome on mitochondrial oxidative phosphorylation were additive to those seen with calcium. These data suggest that adrenochrome in the presence of excess calcium in the myocardial cell may impair the process of energy production in mitochondria and this may result in contractile failure of hearts exposed to this cardiotoxic metabolite of epinephrine.     

Pharmacological protection of mitochondrial function in hypoxic heart muscle: Effect of verapamil, propranolol, and methylprednisolone

Experiments were undertaken to determine if drugs (verapamil, propranolol, and methylprednisolone sodium saccinate) that protect the fine ultrastructure of heart muscle against damage caused by hypoxia, protect mitochondrial function. Mitochondrial function was assessed in terms of oxidative phosphorylating and Ca2 +-accumulating activities. Isolated rabbit hearts were used, and hypoxic conditions induced by reducing the perfusate PO2 from 80.8 to 0.80 kPa (600 to 6 mmHg). The drugs were either added at the start of the hypoxic perfusion or (verapamil and propranolol) the rabbits were pretreated with them. Verapamil, propranolol and, to a lesser extent, methylprednisolone sodium succinate, provided protection evidenced by the maintainance of near normal mitochondrial oxidative phosphorylating and Ca2 +-accumulating activities after 60 min hypoxic perfusion. When added directly to mitochondria isolated from hypoxic-perfused muscle, the drugs had no effect.     

The effect of glutamate on hypoxic newborn rabbit heart

Effects of glutamate on myocardial mechanical function and energy metabolism during 120 min of hypoxia and subsequent reoxygenation were studied in the isolated arterially perfused newborn and adult rabbit hearts. The muscle was perfused with a Krebs-Henseleit (KH) solution or KH solution which contained 1 mM glutamate. Glutamate attenuated the effects of hypoxia on mechanical function and tissue ATP concentration, and enhanced the recovery of mechanical function and tissue ATP during reoxygenation. During hypoxia, glutamate increased tissue succinate and GTP with no change in total lactate and pyruvate production. Trace studies using 14C-glutamate and the tissue homogenate showed that hypoxia increased tissue succinate and inhibited TCA cycle. Additional glutamate produced more CO2 and TCA intermediates in both oxygenated and hypoxic mediums. These data indicate that glutamate increased the rate of ATP production in the hypoxic and reoxygenated heart. This study shows that the improvement of mechanical function and ATP formation in the hypoxic myocardium by glutamate was due to an increase in both oxidative phosphorylation and substrate level phosphorylation. The effect of glutamate on the ATP and GTP production in the newborn heart was not different from the adult.     

PKCepsilon activation augments cardiac mitochondrial respiratory post-anoxic reserve--a putative mechanism in PKCepsilon cardioprotection

Modest cardiac-overexpression of constitutively active PKCepsilon (aPKCepsilon) in transgenic mice evokes cardioprotection against ischemia. As aPKCepsilon interacts with mitochondrial respiratory-chain proteins we hypothesized that aPKCepsilon modulates respiration to induce cardioprotection. Using isolated cardiac mitochondria wild-type and aPKCepsilon mice display similar basal mitochondrial respiration, rate of ATP synthesis and adenosine nucleotide translocase (ANT) functional content. Conversely, the aPKCepsilon mitochondria exhibit modest hyperpolarization of their inner mitochondrial membrane potential (DeltaPsi(m)) compared to wild-type mitochondrial by flow cytometry. To assess whether this hyperpolarization engenders resilience to simulated ischemia, anoxia-reoxygenation experiments were performed. Mitochondria were exposed to 45 min anoxia followed by reoxygenation. At reoxygenation, aPKCepsilon mitochondria recovered ADP-dependent respiration to 44 +/- 3% of baseline compared to 28 +/- 2% in WT controls (P = 0.03) in parallel with enhanced ATP synthesis. This preservation in oxidative phosphorylation is coupled to greater ANT functional content [42% > concentration of atractyloside for inhibition in the aPKCepsilon mitochondria vs. WT control (P < 0.0001)], retention of mitochondrial cytochrome c and conservation of DeltaPsi(m). These data demonstrate that mitochondria from PKCepsilon activated mice are intrinsically resilient to anoxia-reoxygenation compared to WT controls. This resilience is in part due to enhanced recovery of oxidative phosphorylation coupled to maintained ANT activity. As maintenance of ATP is a prerequisite for cellular viability we conclude that PKCepsilon activation augmented mitochondrial respiratory capacity in response to anoxia-reoxygenation may contribute to the PKCepsilon cardioprotective program.     

Role of mitochondrial oxidative phosphorylation in the maintenance of intracellular pH.

The oxidative phosphorylation of isolated rabbit heart mitochondria and the accompanying pH changes were simultaneously measured in the same sample. The nearly constant extramitochondrial proton concentration during State 4 respiration decreased considerably under conditions of oxidative phosphorylation. On the other hand, the proton concentration increased when ATP was hydrolysed by the mitochondrial ATPase. The lysis of mitochondria by Triton X-100 equilibrating the pH difference across the inner membrane did not cause significant change in the proton concentration of the sample neither before nor after State 3 respiration. The accumulation of protons during the extramitochondrial hexokinase reaction was reduced by the mitochondrial oxidative phosphorylation. It is suggested that in the normoxic myocardial cell, the protons generated by the extramitochondrial hydrolysis of ATP are utilized during mitochondrial oxidative phosphorylation.     

Oxygen-induced enzyme release. Assessment of mitochondrial function in anoxic myocardial injury and effects of the mitochondrial uncoupling agent 2,4-dinitrophenol (DNP)

Hypoxic injury to isolated-perfused rat hearts resulted in progressive loss of mitochondrial function, as assessed by polarographic techniques. Decreased mitochondrial respiration could be detected as early as 15 min after beginning hypoxic-substrate-free perfusion and by 75 min little or no functional capacity could be detected. Control studies using mixtures of mitochondria from control and damaged hearts suggested that polarographic techniques tend to overestimate mitochondrial injury. In hearts injured by hypoxia, mitochondrial damage was heterogenous and there was a good linear correlation between the respiratory control ratios of mitochondrial suspensions and the percent damaged mitochondria in the suspensions, as determined by electron microscopy. Respiratory control ratios approached one when only 60 to 70% of mitochondria appeared morphologically damaged. The evidence suggested that significant mitochondrial function was retained in hearts after 60 min of hypoxic perfusion, a time when oxygen-induced enzyme release was nearly maximal.In similarly perfused hearts, both oxygen-induced enzyme release and associated anomalous cellular contracture were reduced or completely inhibited by 0.1 or 1 mm 2,4-dinitrophenol (DNP), but not by 0.04 mm DNP. In intact hearts, 1 mm DNP was required to produce biological effects on cellular morphology and enzyme release equivalent to those produced by either hypoxia or cyanide. These results provide evidence that both oxygen-induced enzyme release and cellular contractures are energy dependent events mediated by resumption of mitochondrial oxidative phosphorylation.     

Mitochondrial oxidative phosphorylation and respiratory chain: Review

Basic events concerning oxidative phosphorylation, i.e. the synthesis of ATP at the expense of respired oxygen at the level of mitochondria are described. Our knowledge concerning the functioning of respiratory chain, its structure, organization and topology inside the inner membrane of mitochondria has considerably improved in recent years. A central question — how does the respiratory chain cooperate with ATP-synthetase, also embedded in the inner membrane, to bring about the oxidative phosphorylation of ADP to ATP — has been one of the most challenging and difficult problems in biochemical research. The chemiosmotic hypothesis proposed by the British biochemist Peter Mitchell appears best in describing the basic events of the recovery of the redox energy liberated along the respiratory chain to synthesize ATP through a membrane process. Moreover the chemiosmotic hypothesis is not restricted to mitochondrial oxidative phosphorylation but appears to provide a general explanation to the synthesis of ATP in all transducing membranes: inner mitochondrial membrane, bacterial plasma membrane, thylakoid membranes in chloroplasts of green plants.     

Deleterious effects of endogenous catecholamines on hypoxic myocardial cells following reoxygenation.

The ability of endogenous myocardial catecholamines to modify biochemical alterations during conditions of severe myocardial hypoxia and subsequent reoxygenation was studied in isolated, Langendorff-perfused, potassium-arrested, rat heart preparations. Hearts depleted of endogenous catecholamines (induced by reserpine pretreatment) exhibited higher levels of ATP, creatine phosphate and glycogen and a lesser amount of lactate dehydrogenase release following 100 min of hypoxia (95% N₂: 5% CO₂). The large amount of myocardial lactate dehydrogenase release normally present during reoxygenation of the hypoxic heart (oxygen paradox) was abolished by reserpine pretreatment. These biochemical changes in reserpinized hearts reflect a diminution of myocardial cell damage. Glucagon administration reversed all the beneficial effects of catecholamine depletion in conditions of both severe hypoxia and reoxygenation.It is concluded that endogenous myocardial catecholamines exert harmful effects on the evolution of hypoxic myocardial cell damage in the rat.     

The relationship of regional coronary blood flow to mitochondrial function during reperfusion of the ischemic myocardium

The relationship of changes in regional coronary flow to the nature and degree of biochemical disturbances during occlusion of branches of the left anterior descending coronary artery and following reestablishment of flow was investigated in two groups of dogs: group I, moderate ischemia before reflow, and group II, severe ischemia prior to reflow. Regional coronary blood flow was determined before ligation, after 60 min of ischemia and after 15 min of reflow using labelled microspheres. Hearts made ischemic for 60 min but not reperfused served as controls. Groups I and II were distinguished by the following features. Group II showed a marked exacerbation of biochemical damage on reperfusion of the ischemic region (reduced levels of ATP, impairment of mitochondrial oxygen consumption and mitochondrial calcium binding). This was accompanied by significant subendocaridial hyperemia. Reperfusion in group I, on the otherhand, partially reversed these changes (increased level of ATP in the ischemic-reperfused region, improved mitochondrial oxygen consumption and calcium binding). Mitochondrial calcium uptake and oxidative phosphorylation (ADP/O ratio) were not affected in any group. These data illustrate that the degree of biochemical damage following reperfusion of the ischemic myocardium is determined by the degree of ischemia, and suggest that interference with ATP production by the mitochondria is not responsible for the damage.     

Impairment of mitochondrial and sarcoplasmic reticular functions during the development of heart failure in cardiomyopathic (UM-X7.1) hamsters

The oxidative phosphorylation as well as calcium transporting properties of heart mitochondria and calcium transport activities of the fragments of the sarcoplasmic reticulum (microsomes) were studied during the life span of cardiomyopathic hamsters (UM-X7.1). Control healthy hamsters of the same age group were used for comparison. No changes in the oxidative phosphorylation ability of cardiomyopathic mitochondria were seen at early and moderate stages of heart failure; however, at severe stages, mitochondrial respiratory functions, but not the ADP:0 ratio, were impaired. Both creatine phosphate and ATP contents were decreased without any significant changes in the ATPase activities of myofibrils from the failing hearts. Heart mitochondria from cardiomyopathic animals at severe stages of failure exhibited less calcium binding and uptake activities in comparison with the control values whereas no changes in the mitochondrial calcium binding and uptake were seen in cardiomyopathic hamsters which showed no clinical signs of heart failure. Although mitochondrial calcium binding in cardiomyopathic hearts at early and moderate stages of failure was decreased, mitochondrial calcium uptake was not significantly different from the control. Microsomal calcium binding activity, unlike calcium uptake activity, was decreased in the hearts of cardiomyopathic hamsters without any signs of heart failure. Both calcium binding and calcium uptake activities of microsomes from animals with early, moderate and severe heart failure were less in comparison with the control values but were not associated with any changes in the Ca2+-stimulated ATPase activity. These results suggest that changes in the process of mitochondrial energy production and mitochondrial Ca2+-transport may be secondary to other factors whereas alterations in the sarcoplasmic reticular Ca2+-transport may lead to the development of heart failure in the cardiomyopathic hamsters.     

Adenine nucleotide metabolites are beneficial for recovery of cardiac contractile force after hypoxia

In a previous study, we demonstrated a significant release of adenosine, inosine and hypoxanthine during hypoxia and subsequent reoxygenation. The present study was designed to determine whether or not exogenous adenosine, inosine and hypoxanthine are beneficial for the recovery of hypoxia-induced loss of cardiac contractile force. Hearts were perfused for 20 min under hypoxic conditions, followed by 45 min-perfusion under reoxygenated conditions, and changes in contractile force, resting tension and metabolic parameters of the perfused heart were examined. When either adenosine, inosine or hypoxanthine were exogenously infused during hypoxia at the rate of 3 mumol/min, remarkable recovery (61 to 68%) of cardiac contractile force was observed upon reoxygenation. The recovery was accompanied by a significant restoration of myocardial ATP (90 to 100%) and CP contents (80 to 86%), suggesting that exogenous metabolites are utilized for the restoration of myocardial ATP during reoxygenation, which may lead to a beneficial recovery of hypoxia-induced loss of cardiac contractile force upon reoxygenation. Infusion of exogenous metabolites also resulted in an almost complete inhibition of hypoxia- and reoxygenation-induced release of creatine phosphokinase from the perfused heart as well as a significant depression of hypoxia-induced calcium accumulation in the cardiac tissue. Since these phenomena are considered to represent increases in cell membrane permeability, protection of the myocardium against hypoxia- and reoxygenation-induced changes in cell membrane permeability may be an alternative mechanism for the beneficial effect of adenosine, inosine and hypoxanthine on the hypoxic myocardium.     

Cardiac mitochondria in heart failure: decrease in respirasomes and oxidative phosphorylation

AIMS: Mitochondrial dysfunction is a major factor in heart failure (HF). A pronounced variability of mitochondrial electron transport chain (ETC) defects is reported to occur in severe acquired cardiomyopathies without a consistent trend for depressed activity or expression. The aim of this study was to define the defect in the integrative function of cardiac mitochondria in coronary microembolization-induced HF. METHODS AND RESULTS: Studies were performed in the canine coronary microembolization-induced HF model of moderate severity. Oxidative phosphorylation was assessed as the integrative function of mitochondria, using a comprehensive variety of substrates in order to investigate mitochondrial membrane transport, dehydrogenase activity and electron-transport coupled to ATP synthesis. The supramolecular organization of the mitochondrial ETC also was investigated by native gel electrophoresis. We found a dramatic decrease in ADP-stimulated respiration that was not relieved by an uncoupler. Moreover, the ADP/O ratio was normal, indicating no defect in the phosphorylation apparatus. The data point to a defect in oxidative phosphorylation within the ETC. However, the individual activities of ETC complexes were normal. The amount of the supercomplex consisting of complex I/complex III dimer/complex IV, the major form of respirasome considered essential for oxidative phosphorylation, was decreased. CONCLUSIONS: We propose that the mitochondrial defect lies in the supermolecular assembly rather than in the individual components of the ETC.     

Hypoxia/reoxygenation of isolated rat heart mitochondria causes cytochrome c release and oxidative stress; evidence for involvement of mitochondrial nitric oxide synthase

The objective of the present study was to delineate the molecular mechanisms for mitochondrial contribution to oxidative stress induced by hypoxia and reoxygenation in the heart. The present study introduces a novel model allowing real-time study of mitochondria under hypoxia and reoxygenation, and describes the significance of intramitochondrial calcium homeostasis and mitochondrial nitric oxide synthase (mtNOS) for oxidative stress. The present study shows that incubating isolated rat heart mitochondria under hypoxia followed by reoxygenation, but not hypoxia per se, causes cytochrome c release from the mitochondria, oxidative modification of mitochondrial lipids and proteins, and inactivation of mitochondrial enzymes susceptible to inactivation by peroxynitrite. These alterations were prevented when mtNOS was inhibited or mitochondria were supplemented with antioxidant peroxynitrite scavengers. The present study shows mitochondria independent of other cellular components respond to hypoxia/reoxygenation by elevating intramitochondrial ionized calcium and stimulating mtNOS. The present study proposes a crucial role for heart mitochondrial calcium homeostasis and mtNOS in oxidative stress induced by hypoxia/reoxygenation.     

Arrhythmia after the release of inhibited oxidative phosphorylation

Automaticities due to delayed afterdepolarizations, elicited upon reoxygenation, are thought to be caused by Ca overload. Since tissue Ca uptake upon reoxygenation has been reported to be closely related to metabolic inhibition during hypoxic perfusion, the relationship between the degree of metabolic inhibition during hypoxia and reoxygenation-induced arrhythmias was investigated in guinea pig papillary muscles. (1) Arrhythmias occurred after 60 min substrate-free hypoxia, but not after 30 min hypoxia. The chance of automaticities was closely related with the increase in resting tension achieved during hypoxic period. The incidence of arrhythmias was, however, lower after 90 or 120 min hypoxia. (2) There were arrhythmias after 20-30 min hypoxia with glycolytic inhibition (20 mM 2-deoxyglucose + 5 mM acetate). On the other hand, 60-min hypoxia in the presence of 5 mM glucose did not elicit arrhythmias on reoxygenation. (3) Stimulation of glycolysis (50 mM glucose) during substrate-free hypoxia prolonged the action potential duration, but did not cause arrhythmias. Washout of cyanide (1 mM) after 60 min perfusion in the presence of oxygen, caused arrhythmias and aftercontractions. These results suggest that the degree of metabolic inhibition during hypoxia is closely related to Ca overload and the resultant arrhythmias upon reoxygenation. The release of inhibited oxidative phosphorylation, rather than the reintroduction of oxygen per se, was thought to be the key mechanism of reoxygenation-induced arrhythmias.     

Increased mitochondrial K(ATP) channel activity during chronic myocardial hypoxia: is cardioprotection mediated by improved bioenergetics?

Increased resistance to myocardial ischemia in chronically hypoxic immature rabbit hearts is associated with activation of ATP-sensitive K(+) (K(ATP)) channels. We determined whether chronic hypoxia from birth alters the function of the mitochondrial K(ATP) channel. The K(ATP) channel opener bimakalim (1 micromol/L) increased postischemic recovery of left ventricular developed pressure in isolated normoxic (FIO(2)=0.21) hearts to values (42+/-4% to 67+/-5% ) not different from those of hypoxic controls but did not alter postischemic recovery of developed pressure in isolated chronically hypoxic (FIO(2)=0.12) hearts (69+/-5% to 72+/-5%). Conversely, the K(ATP) channel blockers glibenclamide (1 micromol/L) and 5-hydroxydecanoate (5-HD, 300 micromol/L) attenuated the cardioprotective effect of hypoxia but had no effect on postischemic recovery of function in normoxic hearts. ATP synthesis rates in hypoxic heart mitochondria (3.92+/-0.23 micromol ATP. min(-1). mg mitochondrial protein(-1)) were significantly greater than rates in normoxic hearts (2.95+/-0.08 micromol ATP. min(-1). mg mitochondrial protein(-1)). Bimakalim (1 micromol/L) decreased the rate of ATP synthesis in normoxic heart mitochondria consistent with mitochondrial K(ATP) channel activation and mitochondrial depolarization. The effect of bimakalim on ATP synthesis was antagonized by the K(ATP) channel blockers glibenclamide (1 micromol/L) and 5-HD (300 micromol/L) in normoxic heart mitochondria, whereas glibenclamide and 5-HD alone had no effect. In hypoxic heart mitochondria, the rate of ATP synthesis was not affected by bimakalim but was attenuated by glibenclamide and 5-HD. We conclude that mitochondrial K(ATP) channels are activated in chronically hypoxic rabbit hearts and implicate activation of this channel in the improved mitochondrial bioenergetics and cardioprotection observed.     

Direct thyroid hormone activation of mitochondria: the role of adenine nucleotide translocase

A presumptive mitochondrial T3 receptor previously reported from this and other laboratories appears capable of accounting for the activation of liver mitochondrial oxidative phosphorylation within 30 min after iv bolus injection of nanogram doses of T3 into hypothyroid rats. The inner mitochondrial membrane carrier adenine nucleotide translocase (AdNT) catalyzes the exchange between the extra- and intramitochondrial ADP and ATP, and has been shown by measurements of flux control coefficients to exert a significant measure of control over the rate of mitochondrial oxidative phosphorylation. The activity of this carrier had been reported to be depressed below normal in hypothyroid rats and restored to normal by hormone replacement. Preparations of AdNT from beef heart mitochondria were found to exhibit high affinity, low capacity binding of [125I]T3. The findings make the mitochondrial carrier AdNT a strong candidate for the initiating site for thyroid hormone stimulation in mammalian species.     

Functional and subcellular changes in the isolated rat heart perfused with oxidized isoproterenol.

The isolated rat heart failed to generate contractile force within 10, 15 and 60 min upon perfusion with medium containing 100, 50 and 20 mg/l oxidized isoproterenol respectively, whereas the contractile force was depressed by about 85% of control following a 90 min perfusion with 10 mg/l oxidized isoproterenol. Swelling of mitochondria and sarcoplasmic reticulum, and disruption of the contractile proteins were seen in all hearts failing due to oxidized isoproterenol. Furthermore, calcium uptake activity, but not calcium binding of the microsomal fraction from hearts perfused with oxidized isoproterenol was depressed, whereas mitochondrial calcium binding and uptake activities were unaffected. Perfusion of the hearts with oxidized isoproterenol did not change the mitochondrial or microsomal ATPase activities; however, mitochondrial phosphorylation rate, state 3 respiration and RCI values were significantly depressed. These results indicate changes in subcellular mechanisms during the induction of myocardial necrosis and contractile failure due to oxidized isoproterenol.     

beta-agonistic bronchodilators: comparison of dose/response in working rat hearts

STUDY OBJECTIVES: Different beta-agonists are compared with regard to their cardiodepressive side effects. DESIGN: The metaphenolic bronchodilators reproterol, salbutamol, fenoterol, and terbutaline were introduced at a dosage of 0.0005 micromol to a maximum of 10 micromol per gram of heart tissue into the isolated working rat heart under hypoxic conditions, and the response was observed during subsequent reoxygenation. As an index of external heart work, aortic flow was measured. Heart rate, coronary flow, and developed pressure were recorded. At the end of heart perfusion, mitochondria were isolated and analyzed for adenosine triphosphatase activity, adenosine triphosphate (ATP) synthesis, and membrane fluidity. Moreover, intact mitochondria and lipid peroxidation were investigated using a model system. Measurements and results: Compared to controls, reproterol gave the most favorable results, with an increase of 25 to 30% of aortic flow during reoxygenation at a concentration of 10 micromol/g heart tissue. In contrast, both fenoterol and salbutamol at a concentration of 1 micromol/g heart tissue decreased aortic flow during reoxygenation, whereas terbutaline had a negative influence on aortic flow at 0.01 to 0.1 micromol/g heart tissue. Mitochondria of these hearts were isolated at the end of the experiment. Mitochondrial ATP synthesis was increased above controls at nearly all concentrations of reproterol. ATP synthesis was decreased at 1 micromol and 10 micromol fenoterol. As little as 0.0005 micromol terbutaline decreased ATP synthesis by 50%. In intact mitochondria, adenosine diphosphate (ADP) to oxygen ratios were found to be increased with terbutaline and fenoterol, indicating ADP consumption by myokinase activation. Lipid peroxidation was increased in a model system between concentrations of 0.002 micromol/mg and 0.04 micromol/mg phosphatidylcholine by fenoterol and terbutaline, whereas a decrease was noted with reproterol. Membrane fluidity was found increased after addition of reproterol, which supports the evidence of efficient ATP synthesis by this compound. CONCLUSIONS: Cardiodepressive side effects and greater toxicity of fenoterol and terbutaline were found under the conditions of our experiment. Salbutamol and, in particular, reproterol appear much better tolerated. In addition to partial beta-adrenergic agonism, reproterol may exert an inhibitory influence on adenosine receptor sites and phosphodiesterase, which could result in membrane stabilization by saving cyclic adenosine monophosphate or ATP.     

Heart mitochondria: gates of life and death

Mitochondria are important generators of energy, providing ATP through oxidative phosphorylation. However, mitochondria also monitor complex information from the environment and intracellular milieu, including the presence or absence of growth factors, oxygen, reactive oxygen species, and DNA damage. Mitochondria have been implicated in the loss of cells in various cardiac pathologies, including ischaemia/reperfusion injury, cardiomyopathy, and congestive heart failure. The release of factors such as cytochrome c, Smac, Omi/Htr2A, and AIF from mitochondria serves to activate a highly complex and regulated cell death program. Furthermore, mitochondrial calcium overload might trigger opening of the mitochondrial permeability transition pore, causing uncoupling of oxidative phosphorylation, swelling of the mitochondria due to influx of water, and rupture of the mitochondrial outer membrane. In this review, we discuss the role of mitochondria in the control of cell death in cardiac myocytes.