Heart rate reduction by pharmacological If current inhibition.

Heart rate reduction is becoming a new strategy to treat coronary patients. The development of heart-rate-lowering drugs, with a more specific activity than Beta-blockers, coincides with the detection of the sinoatrial pacemaker I(f) current. The first selective I(f) inhibitor that has been approved for clinical use is ivabradine. Ivabradine has been shown to reduce heart rate, preserve myocardial contractility, increase diastolic filling and maintain both small and large coronary artery vasodilation, whatever the level of exercise, thus ensuring adequate endocardial blood perfusion during exercise. Furthermore ivabradine decreases myocardial oxygen consumption and improves myocardial energetics, protecting the myocardium during acute ischemic conditions and showing favorable antiremodelling properties in patients with chronic ischemic disease.

In vitro administration of ergothioneine failed to protect isolated ischaemic and reperfused rabbit heart.

Ergothioneine, a natural thiol-containing molecule, has recently been proposed to protect the heart against damage caused by ischaemia and reperfusion. We investigated the possibility that ergothioneine can have a role in maintaining the myocardial thiol/disulfide balance and consequently also a protective effect against ischaemic and reperfusion injury. We used isolated Langendorff-perfused rabbit hearts subjected to 45 min global and total ischaemia followed by 30 min reperfusion at baseline coronary flow (22 ml/min). Ergothioneine was delivered at 10(-5) M and 10(-4) M 60 min before ischaemia and during reperfusion. Myocardial damage was determined in terms of mechanical function, creatine kinase (CK) and lactate release, energy phosphate stores and the occurrence of oxidative stress. In our experimental conditions the treatment was unable to prevent myocardial damage. Ergothioneine, independently from the dosage used, failed to: (i) increase recovery of developed pressure upon reperfusion (14.4 +/- 2.3 mmHg in control hearts vs. 10.3 +/- 2.9 and 12.5 +/- 2.3 mmHg in 10(-5) M and 10(-4) M ergothioneine treated hearts, respectively); (ii) decrease the rise in diastolic pressure (44.3 +/- 4.4 mmHg in control hearts vs. 49.8 +/- 5.8 and 48.0 +/- 7.7 mmHg in treated hearts); (iii) decrease the release of CK and lactate; (iv) increase the levels of adenosine triphosphate (ATP) and creatine phosphate (CP) in tissue upon reperfusion; (v) maintain ratio between oxidized and reduced forms of adenine nucleotide coenzyme, as index of aerobic metabolism; (vi) prevent the decline of reduced glutathione (GSH), or the accumulation of oxidized glutathione (GSSG) as an index of oxidative stress.

Reduction of oxidative stress by carvedilol: role in maintenance of ischaemic myocardium viability.

OBJECTIVES: To differentiate the impact of the beta-blocking and the anti-oxidant activity of carvedilol in maintaining myocardium viability. METHODS: Isolated rabbit hearts, subjected to aerobic perfusion, or low-flow ischaemia followed by reperfusion, were treated with two doses of carvedilol, one dose (2.0 microM) with marked negative inotropic effect due to beta-blockage and the other (0.1 microM) with no beta-blockage nor negative inotropism. Carvedilol was compared with two doses of propranolol, 1.0 - without - and 5.0 microM - with negative inotropic effect. Anti-oxidant activity was measured as the capacity to counteract the occurrence of oxidative stress and myocardium viability as recovery of left ventricular function on reperfusion, membrane damage and energetic status. RESULTS: Carvedilol counteracted the ischemia and reperfusion induced oxidative stress: myocardial content of reduced glutathione, protein and non-protein sulfhydryl groups after ischaemia and particularly after reperfusion, was higher in hearts treated with carvedilol, while the myocardial content of oxidised glutathione was significantly reduced (0.30+/-0.03 and 0.21+/-0.02 vs. 0.39+/-0.03 nmol/mg prot, both P<0.01, in 0.1 and 2.0 microM). At the same time, carvedilol improved myocardium viability independently from its beta-blocking effect. On the contrary, propranolol maintained viability only at the higher dose, although to a lesser extent than carvedilol. This suggests that the effects of propranolol are dependent on energy saving due to negative inotropism. The extra-protection observed with carvedilol at both doses is likely due to its anti-oxidant effect. CONCLUSIONS: Our data show that the anti-oxidant activity of carvedilol is relevant for the maintenance of myocardium viability.

New insights on myocardial pyridine nucleotides and thiol redox state in ischemia and reperfusion damage.

OBJECTIVE: to investigate the changes of pyridine nucleotides and thiol redox state in cardiac tissue following ischemia and reperfusion. NADH/NAD and NADPH/NADP redox couples were specifically studied and the influence of NADPH availability on cellular thiol redox was also investigated. METHODS: isolated rabbit hearts were Langendorff perfused and subjected to a protocol of ischemia and reperfusion. An improved technique for extraction and selective quantitation of pyridine nucleotides was applied. RESULTS: ischemia and reperfusion induced an increase in diastolic pressure, limited recovery in developed pressure and loss of creatine phosphokinase. Creatine phosphate and ATP were decreased by ischemia and only partially recovered during reperfusion. NADH was increased (from 0. 36+/-0.04 to 1.96+/-0.15 micromol/g dry wt. in ischemia, P<0.001), whereas NADPH decreased during ischemia (from 0.78+/-0.04 to 0. 50+/-0.06 micromol/g dry wt., P<0.01) and reperfusion (0.45+/-0.03 micromol/g dry wt.). Furthermore, we observed: (a) release of reduced (GSH) and oxidised glutathione (GSSG) during reperfusion; (b) decreased content of reduced sulfhydryl groups during ischemia and reperfusion (GSH: from 10.02+/-0.76 to 7.11+/-0.81 nmol/mg protein, P<0.05, and to 5.48+/-0.57 nmol/mg protein; protein-SH: from 280.42+/-12.16 to 135.11+/-17.00 nmol/mg protein, P<0.001, and to 190.21+/-11.98 nmol/mg protein); (c) increased content in GSSG during reperfusion (from 0.17+/-0.02 to 0.36+/-0.02 nmol/mg protein, P<0.001); (d) increased content in mixed disulphides during ischemia (from 6.14+/-0.13 to 8.31+/-0.44 nmol/mg protein, P<0.01) and reperfusion (to 9.87+/-0.82 nmol/mg protein, P<0.01). CONCLUSIONS: under severe low-flow ischemia, myocardial NADPH levels can decrease despite the accumulation of NADH. The reduced myocardial capacity to maintain NADPH/NADP redox potential can result in thiol redox state changes. These abnormalities may have important consequences on cellular function and viability.

Role of oxygen free radicals in ischemic and reperfused myocardium.

In recent years there has been considerable interest concerning the role of oxygen radicals in myocardial ischemia and reperfusion injury. The sequential univalent reduction of oxygen gives rise to very reactive intermediate products. Normally, the tissue concentration of these intermediate products of oxygen is limited and the aerobic myocardium survives because of the existence of a delicate balance between the generation of the various oxidants and the maintenance of the antioxidant defense mechanism. Several possible sources have been identified for the production of active oxygen species after ischemia and reperfusion and these sources may be mutually interactive. The ability of scavengers of oxygen free radicals, including vitamin E, to improve mechanical, mitochondrial, and sarcoplasmic reticulum function in animal models of ischemic-reperfusion injury also suggests that oxygen free radicals are partly responsible for myocardial damage in these models, although caution in the interpretation of these data is necessary.

Oxygen free radicals and myocardial damage: protective role of thiol-containing agents.

It has been suggested that the sudden presence of oxygen during reperfusion after a period of ischemia may be toxic for the myocardial cell. The oxygen molecule is capable of producing reactions in the cell, forming highly reactive free radicals, and inducing lipid peroxidation of membranes, altering their integrity and increasing their fluidity and permeability. The ischemic and reperfused cardiac cell is the prime candidate for this reaction sequence and may explain the molecular mechanism underlying the pathologic events related to membrane dysfunction and calcium homeostasis. However, the myocardium has a series of defense mechanisms including the enzymes superoxide dismutase (SOD), catalase, and glutathione peroxidase plus other endogenous antioxidants such as vitamin E, ascorbic acid, and cysteine to protect the cell against the cytotoxic oxygen metabolites. The prerequisite for oxygen free radical involvement in ischemia and reperfusion damage is that ischemia alters the defense mechanisms against oxygen toxicity. It is known that ischemia may impair mitochondrial SOD and, with reperfusion, oxidative stress may occur as shown by tissue accumulation and release of oxidized glutathione. This tripeptide molecule in the cofactor of glutathione peroxidase, the enzyme that removes hydrogen and lipid peroxides. Its formation and subsequent release is a reliable index of oxidative damage. In our study, we investigated the effects of N-acetylcysteine on oxidative damage in the isolated rabbit heart. N-acetylcysteine increases, in a dose-dependent manner (from 10(-7) to 10(-5) M), the myocardial glutathione content and provides an important degree of protection against ischemia and reperfusion. Oxidative stress does not occur, mitochondrial function is maintained, enzyme release is reduced, and contractile recovery is increased. Similarly, we administered N-acetylcysteine in the pulmonary artery of coronary artery disease patients undergoing coronary bypass grafting (150 mg/kg in 1 hour followed by 150 mg/kg in 4 hours). The degree of oxidative stress on reperfusion was reduced and recovery of cardiac function improved. In this article, we review the cardioprotective role of thiol-containing agents.

Effects of iloprost (ZK 36374) on glutathione status during ischaemia and reperfusion of rabbit isolated hearts.

1. Reperfusion of rabbit isolated hearts after 60 min of ischaemia resulted in poor recovery of mechanical function, release of creatine phosphokinase (CPK) and of reduced (GSH) and oxidized (GSSG) glutathione, reduction of mitochondrial superoxide dismutase (Mn SOD) activity and of tissue GSH/GSSG ratio with a shift of cellular thiol redox state toward oxidation, suggesting the occurrence of oxidative stress. 2. Pretreatment of the isolated heart with the stable prostacyclin analogue (iloprost) at 27 or 270 nM, but not at 2.7 nM, improved the functional recovery of the myocardium, reduced CPK, GSH and GSSG release, maintained Mn SOD activity and attenuated the occurrence of oxidative stress. 3. This effect of iloprost cannot be explained by a decreased demand or an enhanced delivery of oxygen during ischaemia or by a direct effect on glutathione peroxidase and reductase activity.

Role of bradykinin and eNOS in the anti-ischaemic effect of trandolapril.

1. Angiotensin converting enzyme (ACE) inhibitors are under study in ischaemic heart diseases, their mechanism of action being still unknown. 2. The anti-ischaemic effect of trandolapril and the possible involvement of a bradykinin-modulation on endothelial constitutive nitric oxide synthase (eNOS) in exerting this effect, were investigated. 3. Three doses of trandolapril, chronically administered in vivo, were studied in isolated perfused rat hearts subjected to global ischaemia followed by reperfusion. 4. Trandolapril has an anti-ischaemic effect. The dose of 0.3 mg kg(-1) exerted the best effect reducing diastolic pressure increase during ischaemia (from 33.0+/-4.5 to 14.0+/-5.2 mmHg; P<0.05 vs control) and reperfusion (from 86.1+/-9.4 to 22.2+/-4.1 mmHg; P<0.01 vs control), improving functional recovery, counteracting creatine phosphokinase release and ameliorating energy metabolism after reperfusion. 5. Trandolapril down-regulated the baseline developed pressure. 6. Trandolapril increased myocardial bradykinin content (from 31.8+/-6.1 to 54.8+/-7.5 fmol/gww; P<0.05, at baseline) and eNOS expression and activity in aortic endothelium (both P<0.01 vs control) and in cardiac myocytes (from 11.3+/-1.5 to 17.0+/-2.0 mUOD microg protein(-1) and from 0.62+/-0.05 to 0.80+/-0.06 pmol mg prot(-1) min(-1); both P<0.05 vs control). 7. HOE 140 (a bradykinin B(2) receptor antagonist) and NOS inhibitors counteracted the above-reported effects. 8. There was a negative correlation between myocyte's eNOS up-regulation and myocardial contraction down-regulation. 9. Our findings suggest that the down-regulation exerted by trandolapril on baseline cardiac contractility, through a bradykinin-mediated increase in NO production, plays a crucial role in the anti-ischaemic effect of trandolapril by reducing energy breakdown during ischaemia.

Changes in oxidative stress and cellular redox potential during myocardial storage for transplantation: experimental studies.

BACKGROUND: Cardioplegic solutions assure only a sub-optimal myocardial protection during prolonged storage for transplantation. The ultimate cause of myocardial damage during storage is unknown, but oxygen free radicals might be involved. We evaluated the occurrence of oxidative stress and changes in cellular redox potential after different periods of hypothermic storage. METHODS: Langendorff-perfused rabbit hearts were subjected to a protocol mimicking each stage of a cardiac transplantation procedure: explantation, storage and reperfusion. Three periods of storage were considered: Group A = 5 hours, Group B = 15 hours, and Group C = 24 hours. Oxidative stress was determined in terms of myocardial content and release of reduced (GSH) and oxidized (GSSG) glutathione, and cellular redox potential as oxidized and reduced pyridine nucleotides ratio (NAD/NADH). Data on mechanical function, cellular integrity and myocardial energetic status were collected. RESULTS: At the end of reperfusion, despite the different timings of storage, recovery of left ventricular developed pressure (46.1+/-7.0, 54.7+/-6.7, and 45.7+/-7.4% of the baseline pre-ischaemic value), energy charge (0.81+/-0.02, 0.81+/-0.02, and 0.77+/-0.01) and NAD/NADH ratio (8.87+/-1.08, 9.39+/-1.72, and 10.26+/-1.98) were similar in all groups (A, B and C). On the contrary, the rise in left ventricular resting pressure (LVRP) and GSH/GSSG ratio were significantly different between Group C, and Groups A and B (p<0.0001, analyzed by Generalized Estimating Equations model for repeated measures, and p<0.05, respectively). CONCLUSIONS: The pathophysiology of myocardial damage during hypothermic storage cannot be considered as a normothermic ischaemic injury and parameters other than energetic metabolism, such as thiolic redox state, are more predictive of functional recovery upon reperfusion.

Effects of anipamil on myocardial sarcolemmal and mitochondrial calcium transport, comparison with verapamil and nifedipine.

The calcium antagonists anipamil, verapamil and nifedipine inhibited, dose dependently, passive and ATP-driven 45Ca2(+)-uptake in purified rabbit ventricular sarcolemmal vesicles exposed to a wide range of free calcium concentration (from 0 to 200 microM). The IC50 values for passive binding were virtually identical for all calcium antagonists and the inhibition was relatively independent of the amount of free calcium employed. On the contrary, the order of potency for inhibition of the ATP-driven calcium uptake was: anipamil greater than verapamil greater than nifedipine. The inhibition of nifedipine, at free calcium concentrations lower than 80 microM, was preceded by a slight stimulation. The inhibitory effects of anipamil and verapamil, but not those of nifedipine, on the ATP-driven calcium uptake were more evident with increasing external calcium concentration. Verapamil and nifedipine failed to modify the initial rate of mitochondrial calcium transport either in the presence or in the absence of ADP; on the contrary, anipamil induced a dose-dependent inhibition of mitochondrial calcium transport. The inhibition occurred over the whole range of calcium concentrations tested, independent of the presence of ADP. The effects of anipamil, but not those of verapamil and nifedipine, on sarcolemmal and mitochondrial calcium transport were long lasting and survived membrane isolation.

Role of timing of administration in the cardioprotective effect of iloprost, a stable prostacyclin mimetic.

We administered iloprost, a stable prostacyclin mimetic, 27 nM, to isolated and perfused rabbit hearts submitted, after 60 min of equilibration, to an ischaemic period (60 min at a coronary flow of 1 ml/min) followed by a period of reperfusion (30 min at a coronary flow of 25 ml/min). Iloprost was delivered at different times during the experimental protocol: 60 min before ischaemia, at the onset and after 30 min of ischaemia and only during reperfusion. The iloprost cardioprotective effect was evaluated in terms of recovery of left ventricular pressure developed during reperfusion, creatine phosphokinase (CPK) and noradrenaline release, mitochondrial function (expressed as yield, RCI (respiratory control index), QO2, ADP/O), ATP and creatine phosphate (CP) tissue contents, calcium homeostasis and by measuring several parameters of oxidative stress: reduced and oxidized glutathione release and tissue contents, Mn and Cu-Zn superoxide dismutase activities; glutathione reductase and peroxidase activities. Our data show that the cytoprotective action of iloprost is closely related to the time of administration. Optimal myocardial preservation was achieved when it was given before or at the onset of ischaemia. Iloprost administration 30 min after the onset of ischaemia was still beneficial, although to a lesser extent. Iloprost lost its protective effect when given only on reperfusion. The data suggest that the iloprost cardioprotective effect is related to maintainance of membrane integrity.

Time course of human atrial natriuretic factor release during cardiopulmonary bypass in mitral valve and coronary artery diseased patients.

We determined the time-course of the release of atrial natriuretic factor (ANF) during cardiopulmonary bypass (CPB) in six patients undergoing coronary artery bypass (CAD) and eight patients undergoing valve replacement for mitral stenosis (MS). Before CPB, the arterial ANF was significantly higher in MS patients than in CAD patients (243 +/- 38 and 29 +/- 5.8 pg/ml respectively, P less than 0.01). With the onset of CPB, the acute pressure unloading of the atria induced a significant, rapid decrease of ANF only in MS patients (-64% of pre-CPB value at 5 min) and no major changes in CAD patients. Clamping of the aorta induced a further progressive reduction of ANF release to almost zero in both groups. Readmission of coronary flow to the empty atria with declamping resulted in an increase in the plasma level of ANF in both groups to reach the concentration present in MS patients before CPB. After CPB, the ANF levels decreased in CAD patients while remaining elevated in MS patients. These data suggest that ANF release from human atria depends on atrial filling pressure and other unknown factors.

Recognized molecular mechanisms of heart failure: approaches to treatment.

Abnormalities of cytosolic calcium handling and myocyte energetics appear to play an important role in mediating contractile dysfunction in heart failure. Systolic and diastolic dysfunction in the failing heart are related to abnormalities of the excitation-contraction mechanism as well as myofilament calcium sensitivity. These abnormalities can be viewed as a compensatory mechanism as the myocytes by down regulating its function and metabolic activity preserve energy consumption and allow better maintenance of basal cellular homeostasis. The end point of myocyte dysfunction, however, is a reduced contraction, which, in turn, might cause a reduced cardiac output and a threatening of arterial pressure. This causes a second level of adaptation, which implies a neuroendocrine response of the whole organism. Consequently, the syndrome of congestive heart failure is characterized not only by impaired ventricular function, but also by an increase in some endogenous substances leading to vasoconstriction and water and salt retention. Although activation of the systems that release these substances is presumed to be compensatory, the sympathetic nervous system and renin-angiotensin-aldosterone system as well as the endothelins may contribute to the pathogenesis of the syndrome. Opposite to the effects of these systems are those evoked by the release of atrial natriuretic peptides. The peptides exert a potent direct vasodilatation and natriuresis. In addition, atrial natriuretic peptides inhibit the release of norepinephrine from nerve terminals and suppress the formation of renin. However, the natriuretic and vasodilator effects of these peptides in patients with congestive heart failure are outweighed by the sodium retention and vasoconstriction caused by sympathetic stimulation and activation of the renin-angiotensin-aldosterone system. The reasons for this are not entirely known. The atrial stretch receptors that are responsible for the release of the atrial peptides become impaired, and it has been suggested that patient with heart failure may adapt to the physiologic effects of atrial natriuretic peptides. The possibility that congestive heart failure is in part a humoral disease is reviewed here and consequently pharmacologic treatment aimed at reducing the effects of the neuroendocrine response as to be advantageous for patients with heart failure.

Therapeutic modulation of the nitric oxide: all ace inhibitors are not equivalent.

The properties of the angiotensin-converting enzyme (ACE) inhibitors have largely been attributed to a class effect. However, this opinion is now increasingly challenged in view of the findings from recent clinical trials, which have demonstrated differential effects of ACE inhibitors, in particular with respect to secondary cardiovascular prevention outcomes. In this experimental study, Sprague-Dawley rats were treated with five different ACE inhibitors (enalapril, perindopril, quinapril, ramipril, and trandolapril) at equihypotensive doses. All ACE inhibitors increased endothelial nitric oxide synthase (eNOS) protein expression and activity in the aorta (both P<0.0001 versus vehicle) and in cardiac myocytes (both P<0.05 versus vehicle). A highly significant effect was observed with perindopril when compared with vehicle in the modulation of eNOS protein expression and activity in aorta (22.52+/-1.09 versus 9.12+/-0.57 AU microg(-1) protein and 1.59+/-0.03 versus 0.77+/-0.02 pmol l(-1) citrulline min(-1)mg protein(-1), respectively) and in cardiac myocytes (17.64+/-0.94 versus 11.30+/-0.59 AU microg(-1) protein and 0.93+/-0.02 versus 0.62+/-0.03 pmol l(-1) citrulline min(-1)mg protein(-1), respectively). On the basis of the eNOS protein expression in the rat aorta, the other ACE inhibitors had similar, but lower effects. Indeed, the rank of potency - based both on eNOS protein expression and activity - was perindopril>trandolapril approximately quinapril approximately ramipril approximately enalapril (P<0.05 perindopril versus trandolapril and ramipril and P<0.01 perindopril versus enalapril, respectively). Levels of circulating nitrite/nitrate, the end-metabolites of nitric oxide, were also significantly affected by ACE inhibition, with the same order of potency. Our findings provide further evidence in favor of differential effects associated with ACE inhibitor therapy and suggest that the clinical benefits associated with these drugs may not solely reflect a class effect extending their benefit beyond blood pressure-lowering effect.

Effect of lacidipine on ischaemic and reperfused isolated rabbit hearts.

Lacidipine is a new developed dihydropyridine calcium-antagonist, showing a slow onset and long lasting-selective activity. To assess whether the administration of lacidipine protects the myocardium in a dose-dependent manner against ischaemia and reperfusion, isolated rabbit heart were infused with three different concentrations of lacidipine: 10(-10); 10(-9); 10(-8) M. Diastolic and developed pressures were monitored; coronary effluent was collected and assayed for CPK activity and for noradrenaline concentration; mitochondria were harvested and assayed for respiratory activity, ATP production and calcium content and tissue concentration of ATP, creatine phosphate (CP) and calcium were determined. Occurrence of oxidative stress during ischaemia and reperfusion was also monitored in terms of tissue content and release of reduced (GSH) and oxidized (GSSG) glutathione. Treatment with lacidipine at 10(-10) and 10(-9) M had no effects on the hearts when perfused under aerobic condition, whilst the higher dose reduced developed pressure of 36%. The ischaemic-induced deterioration of mitochondrial function was attenuated. On reperfusion treated hearts recovered better than the untreated hearts with respect to left ventricular performance, replenishment of ATP and CP stores and mitochondrial function. The reperfusion-induced tissue and mitochondrial calcium overload, release of CPK and of noradrenaline and oxidative stress were also significantly reduced. The effects of lacidipine were dose-dependent. The lower concentration (10(-10) M) failed to modify ischaemic and reperfusion damage. The dose of 10(-9) M was cardioprotective, but the best effect was found at 10(-8) M. It is concluded that lacidipine infusion provides a dose dependent protection of the heart against ischaemia and reperfusion. Because this protection occurred also at 10(-9) M, in the absence of negative inotropic effect during normoxia and of a coronary dilatory effect during ischaemia, it cannot be attributed to an energy sparing effect or to improvement of oxygen delivery. From our data we can envisage two other major mechanism: -1) membrane protection -2) reduction of oxygen toxicity. The ATP sparing effect occurring at 10(-8) M is likely to be responsable for the further protection.

Cardioprotection by nisoldipine: role of timing of administration.

Nisoldipine was administered at 10(-9) M, a dose lacking negative inotropism, to isolated and perfused rabbit hearts submitted to 60 min ischaemia (1 ml.min-1) followed by 30 min reperfusion. The drug was delivered either 30 min before ischaemia, at the onset and after 30 min of ischaemia and during reperfusion only. Cardiac protection was evaluated in terms of recovery of left ventricular pressure during reperfusion, release of creatine phosphokinase (CPK), mitochondrial function, tissue content of adenosine triphosphate (ATP) and creatine phosphate (CP), calcium homeostasis and the occurrence of oxidative stress, established measuring content and release of reduced and oxidized glutathione. The cytoprotective action of nisoldipine occurs in the absence of negative inotropism and is closely related to the time of administration. Optimal myocardial preservation is achieved when nisoldipine is given before or at the onset of ischaemia. Prophylactic administration of nisoldipine improved the recovery of the developed pressure from 15.9 +/- 1.0 (SE) mmHg to 47.8 +/- 1.9 mmHg, P < 0.01 and reduced the release of CPK from 830 +/- 29 to 229 +/- 27 mU.min-1 g-1 wet wt, P < 0.01. The accumulation of tissue and mitochondrial calcium was reduced from 58 +/- 11 and 49 +/- 9 to 14 +/- 6 and 10 +/- 4 mmol.kg-1 dry wt respectively, P < 0.01. This resulted in a significant (P < 0.01) preservation of all indices of mitochondrial function, allowing a higher recovery of ATP and CP after reperfusion (from 4.1 +/- 0.7 and 10.0 +/- 0.6 to 16.1 +/- 1.0 and 29.9 +/- 0.2 mumol.g-1 dry wt respectively, P < 0.001). Reperfusion-induced myocardial accumulation and release of oxidized glutathione were reduced from 0.493 +/- 0.07 nmol.mg-1 protein and 0.768 +/- 0.063 nmol.min-1 g-1 wet wt to 0.225 +/- 0.07 and 0.157 +/- 0.038 respectively, P < 0.01. Similar data were obtained when nisoldipine was given at the time of ischaemia, while administration 30 min after the onset of ischaemia showed only a trend towards protection. Nisoldipine lost its protective effect when given on reperfusion. A multifactorial analysis of the data suggest that the cardioprotective effect of nisoldipine is related to the maintenance of membrane integrity, possibly since nisoldipine is highly lipophilic.

Protective effect of a prostacyclin-mimetic on the ischaemic-reperfused rabbit myocardium.

To assess whether the administration of the stable prostacyclin-mimetic ZK 36374 (iloprost) protects the myocardium in a dose-dependent manner against ischaemia and reperfusion, isolated rabbit hearts were infused with three different concentrations of iloprost: 2.7, 27 and 270 nM. Diastolic and developed pressures were monitored; coronary effluent was collected and assayed for creatine phosphokinase (CPK) activity and for noradrenaline concentration; mitochondria were harvested and assayed for respiratory activity; ATP production and calcium content and tissue concentration of adenosine triphosphate (ATP) and creatine phosphate (CP) were determined. Treatment with iloprost altered neither developed pressure under normoxic conditions nor the rate and extent of depletion of ATP and CP during ischaemia. The ischaemic-induced deterioration of mitochondrial function, however, was attenuated. On reperfusion, hearts treated with iloprost recovered better than the untreated hearts with respect to left ventricular performance, replenishment of ATP and CP stores and mitochondrial function. The reperfusion-induced mitochondrial calcium overload and release of CPK and of noradrenaline were also significantly reduced. The effect of iloprost was dose-dependent. The lower concentration (2.7 nM) failed to modify ischaemic and reperfusion damage. The best protective effect was found at 27 nM. An increase of the dose to 270 nM did not result in further protection. It is concluded that iloprost infusion provides a dose-dependent protection of the heart against some of the deleterious effects of ischaemia and reperfusion and, in particular, prevents mitochondrial calcium overload and maintains mitochondrial function. Because this protection occurred in the absence of negative inotropic effect during normoxia or of a coronary dilatory effect during ischaemia, it cannot be attributed to an energy sparing effect or to improvement of oxygen delivery. Therefore, alternative mechanisms of action are to be considered.