A selective inhibitor of Na+/Ca2+ exchanger, SEA0400, preserves cardiac function and high-energy phosphates against ischemia/reperfusion injury

The Ca2+ overload by Ca2+ influx via Na+/Ca2+ exchanger (NCX) is a critical mechanism in myocardial ischemia/reperfusion injury. We investigated protective effects of a novel selective inhibitor of NCX, SEA0400, on cardiac function and energy metabolism during ischemia and reperfusion. Langendorff-perfused rat hearts were exposed to 35 minutes global ischemia and 40 minutes reperfusion. Using 31P nuclear magnetic resonance spectroscopy, cardiac phosphocreatine (PCr), ATP, and pHi were monitored. SEA0400 did not change the basic cardiac function, but improved the recovery of left ventricular developed pressure (LVDP) after reperfusion (27.6 +/- 4.9 mm Hg in control, 101.2 +/- 19.3 mm Hg in 0.1 microM, and 115.5 +/- 13.3 mm Hg in 1 microM SEA0400, means +/- SE, n = 6, P < 0.05). SEA0400 reduced left ventricular end-diastolic pressure and increased coronary flow after reperfusion. SEA0400 improved the recoveries of cardiac phosphocreatine and ATP after reperfusion, but did not affect pHi. There were significant linear correlations between left ventricular developed pressure and cardiac phosphocreatine (r = 0.79, P < 0.05), and left ventricular developed pressure and ATP (r = 0.80, P < 0.05). However, SEA0400 increased the incidence and duration of reperfusion ventricular arrhythmias. SEA0400 added only after reperfusion also improved both the contractile function and energy metabolism. It is concluded that the selective inhibition of NCX may be effective to preserve high-energy phosphates and to improve cardiac function after reperfusion, but may not be able to prevent fatal arrhythmias.     

A novel and selective Na+/Ca2+ exchange inhibitor, SEA0400, improves ischemia/reperfusion-induced renal injury

We evaluated the effects of SEA0400 (2-[4-[(2,5-difluorophenyl)methoxy]phenoxy]-5-ethoxyaniline), a novel and selective Na+/Ca2+ exchange inhibitor, on ischemic acute renal failure. Ischemic acute renal failure in rats was induced by clamping the left renal artery and vein for 45 min followed by reperfusion, 2 weeks after the contralateral nephrectomy. SEA0400 administration (0.3, 1 and 3 mg/kg, i.v.) before ischemia dose-dependently attenuated the ischemia/reperfusion-induced renal dysfunction and histological damage such as tubular necrosis. SEA0400 pretreatment at the higher dose suppressed the increment of renal endothelin-1 content after reperfusion. The ischemia/reperfusion-induced renal dysfunction was also overcome by post-ischemia treatment with SEA0400 at 3 mg/kg, i.v. In in vitro study, SEA0400 (0.2 and 1 microM) protected cultured porcine tubular cells (LLC-PK1) from hypoxia/reoxygenation-induced cell injury. These findings support the view that Ca2+ overload via the reverse mode of Na+/Ca2+ exchange, followed by endothelin-1 overproduction, plays an important role in the pathogenesis of ischemia/reperfusion-induced renal injury. The possibility exists that a selective Na+/Ca2+ exchange inhibitor such as SEA0400 is useful as effective therapeutic agent against ischemic acute renal failure in humans.     

Effects of SEA0400, a Na+/Ca2+ exchange inhibitor, on ventricular arrhythmias in the in vivo dogs

SEA0400 (2-[4-[(2,5-difluorophenyl)methoxy]phenoxy]-5-ethoxyaniline), a novel and selective inhibitor of Na+/Ca2+ exchanger, was investigated for its possible antiarrhythmic effects on arrhythmias of Ca2+ overload induced by coronary ligation/reperfusion and by digitalis in the dog. SEA0400 (1.0 mg/kg) did not change the hemodynamics but slightly prolonged the QRS duration (P<0.05). Pre-ischemic administration (10 min before coronary occlusion) of SEA0400 (1.0 mg/kg) and post-ischemic administration (1 min before reperfusion) of SEA0400 (0.3, 1.0 and 3.0 mg/kg) had no effects on the incidence of ventricular fibrillation induced by coronary ligation/reperfusion. On the other hand, SEA0400 (3.0 mg/kg) decreased the arrhythmic ratio in the digitalis arrhythmias (P<0.01). However, atrioventricular block and cardiac standstill were induced in two digitalized dogs. In conclusion, SEA0400 has no significant antiarrhythmic effect on arrhythmias induced by coronary ligation/reperfusion, but has an obvious suppressing effect on tachyarrhythmias induced by digitalis in in vivo canine models.     

Protective effects of SEA0400, a novel and selective inhibitor of the Na+/Ca2+ exchanger,on myocardial ischemia-reperfusion injuries

The Na(+)/Ca(2+) exchanger (NCX) is involved in myocardial ischemia-reperfusion injuries. We examined the effects of 2-[4-[(2,5-difluorophenyl)methoxy]phenoxy]-5-ethoxyaniline (SEA0400), a potent and selective inhibitor of NCX, on myocardial ischemia-reperfusion injury models. In canine cardiac sarcolemmal vesicles and rat cardiomyocytes, SEA0400 potently inhibited the Na(+)-dependent 45Ca(2+) uptake with an IC(50) value of 90 and 92 nM, compared with 2-[2-[4-(4-nitrobenzyloxy)phenyl]isothiourea (KB-R7943, 7.0 and 9.5 microM), respectively. In rat cardiomyocytes, SEA0400 (1 and 3 microM) attenuated the Ca(2+) paradox-induced cell death. In isolated rat Langendorff hearts, SEA0400 (0.3 and 1 microM) improved the cardiac dysfunction induced by low-pressure perfusion followed by normal perfusion. In anesthetized rats, SEA0400 (0.3 and 1 mg/kg, i.v.) reduced the incidence of ventricular fibrillation and mortality induced by occlusion of the left anterior descending coronary artery followed by reperfusion. These results suggest that SEA0400 is a most potent NCX inhibitor in the heart and that it has protective effects against myocardial ischemia-reperfusion injuries.     

Lipophilic HMG-CoA reductase inhibitors increase myocardial stunning in dogs

Pretreatment of dogs with simvastatin, a lipophilic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor, increases myocardial contractile dysfunction during reperfusion after ischemia (stunning), with reduction of tissue adenosine triphosphate (ATP). This was thought to be a consequence of prevention of ubiquinone biosynthesis by the lipophilic inhibitor in the myocardial cell. We examined whether other lipophilic HMG-CoA reductase inhibitors also influence myocardial stunning in dogs. Vehicle, atorvastatin (2 mg/ kg/day), fluvastatin (4 mg/kg/day), or cerivastatin (40 microg/kg/ day) was orally administered for 3 weeks. Hydrophilic pravastatin (4 mg/kg/day) also was given. After 3 weeks, pentobarbital-anesthetized dogs were subjected to 15-min left anterior descending coronary artery occlusion followed by 2-h reperfusion. Myocardial segment function was determined by sonomicrometry. Tissue levels of ATP were determined in 2-h reperfused hearts. All inhibitors significantly decreased serum cholesterol level. The three lipophilic inhibitors resulted in a worsening of segment function in the reperfused myocardium, as compared with the vehicle group. The levels of ATP in the atorvastatin, fluvastatin, and cerivastatin groups were significantly lower than that in the vehicle group. These results confirm that lipophilic HMG-CoA reductase inhibitors enhance myocardial stunning in association with ATP reduction after ischemia and reperfusion.     

Cardioprotective effect of SEA0400, a selective inhibitor of the Na(+)/Ca(2+) exchanger, on myocardial ischemia-reperfusion injury in rats

In this study, we investigated whether the Na(+)/Ca(2+) exchanger (NCX) inhibitor SEA0400 (2-[4-[(2,5-difluorophenyl)methoxy]phenoxy-5-ethoxyaniline) might have a protective effect against myocardial ischemia-reperfusion injury in rats. In particular, we focused on cardiac function using Doppler echocardiography and cardiac gene expression. We intravenously administered either SEA0400 and delivery vehicle or only the vehicle (as a control) to Wistar rats 5 min before ischemia was induced. Reperfusion was performed after 30 min of ischemia. At 1 week after ischemia-reperfusion injury, we assessed hemodynamics by inserting a polyethylene-tubing catheter, cardiac function by Doppler echocardiography, and myocardial mRNA expression was determined by Northern blot analysis. Left ventricular (LV) end-diastolic dimensions (LVDd) and LV end-diastolic volume (LVEDV) were significantly increased in the ischemia-reperfusion rat model group compared to the control group. The SEA0400-treated group had a significantly attenuated LVDd (P<0.05) and LVEDV (P<0.01) increase, compared to the vehicle-treated group. A decrease in the LV ejection fraction (P<0.05) was significantly prevented in the SEA0400-treated group compared to the vehicle-treated group. Moreover, mRNA expression of plasminogen activator inhibitor-1 in the non-infarcted LV of the SEA0400-treated group was significantly lower than in the vehicle-treated group (P<0.05). This study demonstrates that the NCX is an important mechanism for cell death in myocardial ischemia and reperfusion in rats. SEA0400 may prove to be a promising new drug in the clinical treatment of myocardial ischemia and reperfusion.     

Nicorandil attenuates myocardial dysfunction associated with transient ischemia by opening ATP-dependent potassium channels.

The objective of this study was to determine whether ATP-dependent potassium channel activation is involved in the mechanism by which nicorandil reduces postischemic contractile dysfunction produced by a brief period of ischemia (myocardial stunning). Barbital-anesthetized dogs were subjected to 15-min left anterior descending (LAD) coronary artery occlusion followed by 3-h reperfusion. Saline or nicorandil (100 micrograms/kg + 25 micrograms/kg/min) were infused 15 min before and throughout occlusion with or without addition of the KATP channel antagonist, glibenclamide 0.3 mg/kg as an intravenous (i.v.) bolus. Regional myocardial blood flow was measured by radioactive microspheres, and left ventricular (LV) segment function was measured by sonomicrometry. There were no significant differences between the groups in area-at-risk size or collateral blood flow. In contrast, nicorandil significantly reduced mean aortic blood pressure (BP) and the rate-pressure product (RPP) which persisted throughout the occlusion period. In addition, nicorandil markedly accelerated recovery of segment shortening in the ischemic/reperfused region as compared with control dogs. Pretreatment of dogs with glibenclamide blocked none of the hemodynamic effects of nicorandil, but it did prevent improvement in reperfusion segment function. The small dose of glibenclamide used had no effect on hemodynamics or the degree of stunning. Thus, these results suggest that nicorandil attenuates stunning in anesthetized dogs by a direct cardioprotective effect as a result of KATP channel activation in ischemic myocardium.     

SEA0400: a novel sodium-calcium exchange inhibitor with cardioprotective properties

The cardiac sodium-calcium exchanger (NCX) plays an important role in calcium homeostasis. It is the primary mechanism for removing calcium ions that enter myocytes through L-type calcium channels on a beat-to-beat basis. Its direction of transport is determined by the membrane potential and the ionic concentrations of Na+ and Ca2+, with the forward (or Ca2+-efflux) mode of transport being the dominant mode under physiological conditions. In contrast, the Ca2+-influx mode (or reverse mode) of NCX becomes important in certain pathophysiological conditions, such as myocardial ischemia and reperfusion. Recent discovery of compounds that inhibit the Ca2+-influx mode (or reverse mode) of NCX has generated intense research interest in the pharmacology of NCX. Among the newer NCX inhibitors described to date, 2-[4-[(2,5-difluorophenyl)methoxy]-phenoxy]-5-ethoxyaniline (SEA0400) appears particularly promising in attenuating cardiac, renal, and cerebral ischemia/reperfusion injuries in various experimental models. Moreover, the mixed results that have emerged from clinical trials evaluating the efficacy and safety of inhibitors of the sodium-hydrogen exchanger (an upstream target in relation to the sodium-calcium exchanger) in myocardial protection stimulated interest in evaluating NCX as an alternative therapeutic target. This article reviews the pharmacological profile of SEA0400, as presented in the published literature, and discusses the therapeutic potential of this compound in attenuating myocardial ischemia/reperfusion injury.     

TY-12533, a novel Na(+)/H(+) exchange inhibitor, prevents myocardial stunning in dogs

Anesthetized open-chest dogs were subjected to 15-min myocardial ischemia followed by 2-h reperfusion to induce myocardial stunning. A novel Na(+)/H(+) exchange inhibitor 6,7,8,9-tetrahydro-2-methyl-5H-cyclohepta[b]pyridine-3-carbonylguanidine maleate (TY-12533), administered 10 min before or 10 min after start of ischemia (3 mg/kg/10 min, i.v.), did not affect reductions in regional myocardial wall thickening, blood flow and pH during ischemia, but it significantly improved recovery of the wall thickening and blood flow after reperfusion. These results indicate that TY-12533, even when administered during ischemia, could prevent myocardial stunning without affecting myocardial dysfunction or acidosis induced by brief ischemia.     

Preservation of mitochondrial function may contribute to cardioprotective effects of Na+/Ca2+ exchanger inhibitors in ischaemic/reperfused rat hearts

BACKGROUND AND PURPOSE: Na+/Ca2+ exchanger (NCX) inhibitors are known to attenuate myocardial reperfusion injury. However, the exact mechanisms for the cardioprotection remain unclear. The present study was undertaken to examine the mechanism underlying the cardioprotection by NCX inhibitors against ischaemia/reperfusion injury. EXPERIMENTAL APPROACH: Isolated rat hearts were subjected to 35-min ischaemia/60-min reperfusion or 20-min ischaemia/60-min reperfusion. NCX inhibitors (3-30 microM KB-R7943 (KBR) or 0.3-1 microM SEA0400 (SEA)) were given for 5 min prior to ischaemia (pre-ischaemic treatment) or for 10 min after the onset of reperfusion (post-ischaemic treatment). KEY RESULTS: With 35-min ischaemia/60-min reperfusion, pre- or post-ischaemic treatment with KBR or SEA neither enhanced post-ischaemic contractile recovery nor attenuated ischaemia- or reperfusion-induced Na+ accumulation and damage to mitochondrial respiratory function. With the milder model (20-min ischaemia/reperfusion), pre- or post-ischaemic treatment with 10 microM KBR or 1 microM SEA significantly enhanced the post-ischaemic contractile recovery, associated with reductions in reperfusion-induced Ca2+ accumulation, damage to mitochondrial function, and decrease in myocardial high-energy phosphates. Furthermore, Na+ influx to mitochondria in vitro was enhanced by increased concentrations of NaCl. KBR (10 microM) and 1 microM SEA partially decreased the Na+ influx. CONCLUSIONS AND IMPLICATIONS: The NCX inhibitors exerted cardioprotective effects during relatively mild ischaemia. The mechanism may be attributable to prevention of mitochondrial damage, possibly mediated by attenuation of Na+ overload in cardiac mitochondria during ischaemia and/or Ca2+ overload via the reverse mode of NCX during reperfusion.     

Effect of diltiazem on extent of ultimate myocardial injury resulting from temporary coronary artery occlusion in dogs

The calcium antagonist, diltiazem, was evaluated for its ability to reduce the extent of myocardial injury resulting from 90 min of left circumflex (LCX) coronary artery occlusion in anesthetized dogs. Administration of diltiazem (0.75 mg/kg over 10 min, followed by 600 microgram/kg/h for 4 h) was initiated 30 min prior to LCX occlusion. Regional myocardial blood flow (RMBF) was measured with radioactive microspheres 30 min after LCX occlusion, and at 45 min and 24 h after reperfusion. At 24 h, after obtaining hemodynamic and RMBF measurements, excised hearts were processed by perfusion staining to determine the percent of left ventricle (LV) perfused by LCX (area at risk) and infarct size, with triphenyltetrazolium chloride. Infarct size, expressed as a percentage of the area at risk, was significantly lower in the diltiazem-treated group compared to the control group (27 +/- 4 vs. 42 +/- 5%, respectively). The area at risk, expressed as a percentage of left ventricular mass, was similar in both groups [41 +/- 2 and 44 +/- 3% (area at risk-LV)]. In addition, the marked elevation of tissue Ca2+ content in noninfarcted and infarcted myocardium within the area at risk (18 +/- 2 and 42 +/- 8 mumol Ca2+/g) in control animals was attenuated by diltiazem (6 +/- 3 and 18 +/- 8 mumol Ca2+/g). Diltiazem did not increase blood flow to ischemic myocardium during LCX occlusion. However, reflow to the inner layers of formerly ischemic myocardium during reperfusion was significantly greater in diltiazem-treated dogs. Both arterial blood pressure and heart rate were significantly lower in the diltiazem -treated group. In addition, mortality (1 vs. 4) and occurrence of ventricular arrhythmias during reperfusion were lower in diltiazem-treated dogs. The data suggest that diltiazem reduces myocardial ischemic injury by lowering myocardial oxygen demands indirectly via favorable hemodynamic alterations, and directly by limiting transmembrane Ca2+ fluxes during ischemia and reperfusion.     

Involvement of the Na+/Ca2+ exchanger in ouabain-induced inotropy and arrhythmogenesis in guinea-pig myocardium as revealed by SEA0400

Involvement of the Na+/Ca2+ exchanger in ouabain-induced inotropy and arrhythmogenesis was examined with a specific inhibitor, SEA0400. In right ventricular papillary muscle isolated from guinea-pig ventricle, 1 microM SEA0400, which specifically inhibits the Na+/Ca2+ exchanger by 80%, reduced the ouabain (1 microM)-induced positive inotropy by 40%, but had no effect on the inotropy induced by 100 microM isobutyl methylxantine. SEA0400 significantly inhibited the contracture induced by low Na+ solution. In HEK293 cells expressing the Na+/Ca2+ exchanger, 1 microM ouabain induced an increase in intracellular Ca2+, which was inhibited by SEA0400. The arrhythmic contractions induced by 3 microM ouabain were significantly reduced by SEA0400. These results provide pharmacological evidence that the Na+/Ca2+ exchanger is involved in ouabain-induced inotropy and arrhythmogenesis.     

Cicletanine and reperfusion injury: is there any correlation between arrhythmias, 6-keto-PGF1 alpha, thromboxane B2, and myocardial ion shifts (Na+, K+, Ca2+, and Mg2+) induced by ischemia/reperfusion in isolated rat heart.

We studied the effects of cicletanine, an anti-hypertensive drug, on reperfusion arrhythmias in relation to 6-keto-PGF1 alpha, thromboxane B2 (TXB2), and ion shifts (Na+, K+, Ca2+, and Mg2+) induced by ischemia and reperfusion in hearts isolated from spontaneously hypertensive rats. Hearts were subjected to 30-min global ischemia followed by 10-min reperfusion. 6-keto-PGF1 alpha and TXB2 concentrations were determined by radioimmunoassay (RIA) from coronary effluents, and myocardial Na+, K+, Ca2+, and Mg2+ contents were measured by atomic absorption spectrophotometry from myocardial tissue. Two basic protocols were used: (a) acute administration when 3, 10, 30, or 100 mg/L cicletanine was included in the perfusion buffer; and (b) chronic application, in which rats received cicletanine 3, 10, 30, or 100 mg/kg/day orally for 14 days. Acute administration of the drug in low concentrations (3 or 10 mg/L), significantly increased endogenous 6-keto-PGF1 alpha production before ischemia and during reperfusion, whereas higher doses of cicletanine (30 or 100 mg/L) as well as chronic application of the drug failed to increase production of 6-keto-PGF1 alpha in the myocardium. TXB2 production was not influenced by either acute or chronic treatment with the drug. Neither treatment changed myocardial ion contents in comparison with control values (Na+ = 45 +/- 4, K+ = 252 +/- 7, Ca2+ = 1.4 +/- 0.1, and Mg2+ = 12.5 +/- 0.3 mmol/kg dry weight) in nonischemic hearts. Thirty-minute ischemia resulted in a two- and fourfold accumulation of myocardial Na+ and Ca2+ and a 50% decrease in both K+ and Mg2+.(ABSTRACT TRUNCATED AT 250 WORDS)     

SEA0400, a specific inhibitor of the Na+-Ca2+ exchanger, attenuates sodium nitroprusside-induced apoptosis in cultured rat microglia

1. Using SEA0400, a potent and selective inhibitor of the Na+-Ca2+ exchanger (NCX), we examined whether NCX is involved in nitric oxide (NO)-induced disturbance of endoplasmic reticulum (ER) Ca2+ homeostasis followed by apoptosis in cultured rat microglia. 2. Sodium nitroprusside (SNP), an NO donor, decreased cell viability in a dose- and time-dependent manner with apoptotic cell death in cultured microglia. 3. Treatment with SNP decreased the ER Ca2+ levels as evaluated by measuring the increase in cytosolic Ca2+ level induced by exposing cells to thapsigargin, an irreversible inhibitor of ER Ca2+-ATPase. 4. The treatment with SNP also increased mRNA expression of CHOP and GPR78, makers of ER stress. 5. SEA0400 at 0.3-1.0 microM protected microglia against SNP-induced apoptosis. 6. SEA0400 blocked not only the SNP-induced decrease in ER Ca2+ levels but also SNP-induced increase in CHOP and GRP78 mRNAs. 7. SEA0400 did not affect capacitative Ca2+ entry in the presence and absence of SNP. 8. SNP increased Na+-dependent 45Ca2+ uptake and this increase was blocked by SEA0400. 9. These results suggest that SNP induces apoptosis via the ER stress pathway and SEA0400 attenuates SNP-induced apoptosis via suppression of the ER stress in cultured microglia. Our findings imply that NCX plays a role in ER Ca2+ depletion under pathological conditions.     

Probucol reduces myocardial dysfunction during reperfusion after short-term ischemia in rabbit heart.

The effects of probucol, a lipophilic antioxidant, on the myocardial dysfunction (stunning) observed during reperfusion after 15-min ischemia in rabbit heart were studied. Rabbits received food with or without 1% probucol for 3 weeks. They were then anesthetized and prepared for recording of myocardial segment shortening, arterial blood pressure (BP), left ventricular pressure (LVP), rate of development of LVP (dP/dt), and a lead II ECG. Regional myocardial ischemia was produced by acute occlusion of the first marginal branch of the left coronary artery. Myocardial segment shortening was depressed after reperfusion in control rabbits. In comparison, myocardial segment shortening was significantly greater in probucol-treated rabbits than in control rabbits during reperfusion, indicating a beneficial effect. No hemodynamic or ECG changes measured could explain this difference. The number of premature ventricular contractions was reduced in the probucol-treated group, although the incidence of ventricular tachycardia (VT) and ventricular fibrillation (VF) were not. Concentrations of probucol in serum and heart of five rabbits were 15.0 +/- 1.2 micrograms/ml and 17.5 +/- 2.5 micrograms/g (mean +/- SEM), respectively. Only probucol concentrations in the serum were positively correlated with the improvement in myocardial segment shortening (r = 0.91, p = 0.03). We conclude that a clinically relevant serum concentration of probucol reduces ischemia-induced myocardial stunning in the rabbit.     

SM-20550, a new Na+/H+ exchange inhibitor and its cardioprotective effect in ischemic/reperfused isolated rat hearts by preventing Ca2+-overload

We investigated the effect of a newly synthesized compound, SM-20550 [N-(aminoiminomethyl)-1,4-dimethyl-1H-indole-2-carboxamide methanesulfonic acid] on Na+/H+ or Na+/Ca2+ exchange activity in rat cardiomyocytes, and on radioligand binding with several channels or receptors in membrane preparations, and ischemia/reperfusion injury in isolated perfused rat hearts. In myocytes, SM-20550 concentration-dependently inhibited the recovery from acidosis induced by an NH4Cl prepulse in HCO3(-)-free solution. Its IC50 was 10(-8) M, which was 10 times lower than that of ethylisopropyl amiloride (EIPA). SM-20550 (10(-6) M) did not affect the Na+-dependent Ca2+ influx (Na+/Ca2+ exchange activity) in cardiomyocytes. In the radioligand binding assay, SM-20550 did not have affinity for K+ channel, beta-adrenoceptor, adenosine, angiotensin, or endothelin receptors, and had low affinity for Na+ and Ca2+ channels and alpha-adrenoceptors, only at the concentrations of 10(-6)-10(-5) M. In perfused hearts exposed to 40 min of global ischemia and 20 min of reperfusion, SM-20550 (10(-8)-10(-7) M) significantly reduced the elevation of left ventricular end-diastolic pressure during reperfusion, improved the postischemic recovery of developed pressure, and prevented coronary perfusion pressure increase after reperfusion. Furthermore, SM-20550 reduced creatine phosphokinase release during reperfusion and prevented the abnormal gain of tissue Na+ and Ca2+ at the end of reperfusion. These results suggest that SM-20550 is a potent, highly specific Na+/H+ exchange inhibitor, which exerts a protective effect against myocardial ischemia/reperfusion injury. In addition, our data strongly support the hypothesis that Na+/H+ exchange plays an important role in the development of postischemic cardiac dysfunction, most likely by inducing Na+ and Ca2+ overload.     

Functional proteins involved in regulation of intracellular Ca(2+) for drug development: pharmacology of SEA0400, a specific inhibitor of the Na(+)-Ca(2+) exchanger

The Na(+)-Ca(2+) exchanger (NCX) is involved in regulation of intracellular Ca(2+) concentration. A specific inhibitor of NCX has been required for clarification of the physiological and pathological roles of NCX. We have developed 2-[4-[(2,5-difluorophenyl)methoxy]phenoxy]-5-ethoxyaniline (SEA0400), a highly potent and selective inhibitor of NCX. SEA0400 in the concentration range that inhibits NCX exhibits negligible affinities for the Ca(2+) channels, Na(+) channels, K(+) channels, noradrenaline transporter, and 14 receptors; and it does not affect the activities of the store-operated Ca(2+) channel, Na(+)-H(+) exchanger, and several enzymes including Na(+),K(+)-ATPase and Ca(2+)-ATPase. Furthermore, recent studies show that SEA0400 attenuates ischemia-reperfusion injury in the brain, heart, and kidney and radiofrequency lesion-induced edema in rat brain. These findings suggest that NCX plays a key role in ischemia-reperfusion injury and may be a target molecule for treatment of reperfusion injury-related diseases.     

Radical scavenging properties of novel benzopyran derivatives, TA248 and TA276, and effects of the compounds on ischemic/reperfused myocardium in dogs

Characteristics of novel benzopyran derivatives, TA248 and TA276, and their effects on myocardial contraction in ischemic/reperfused hearts in dogs were examined. TA248 and TA276 inhibited NADPH-dependent lipid peroxidation induced by Fe(3+) in the rat brain homogenate. Both compounds reduced *O(2-) produced by xanthine-xanthine oxidase system in a dose-dependent manner. TA276 scavenged.OH generated by Fenton reaction in a dose-dependent manner. TA248 also inhibited the.OH production, but the effect was neither complete nor dose dependent. Myocardial contraction was assessed as segment shortening of the left ventricular wall in pentobarbital-anesthetized open-chest dogs. The segment shortening was decreased by the left anterior descending coronary artery ligation (ischemia) and returned by release of the ligated artery (reperfusion). The segment shortening did not recover fully during reperfusion. Either TA248 or TA276 injected 10 min before ischemia improved the recovery of myocardial contraction during reperfusion. Both compounds preserved the level of ATP in the 60-min reperfused myocardium. However, the level of lipid peroxides was not changed by TA248 and TA276. TA248 and TA276 may protect myocardium against ischemic/reperfusion insult, partly because of their free radical scavenging activity, but no significant change in myocardial lipid peroxide level was observed.     

粉防己碱对缺血顿抑大鼠心肌功能和ATP酶活力的影响

目的：观察粉防己碱（ｔｅｔｒａｎｄｒｉｎｅ，Ｔｅｔ）对缺血顿抑心肌的保护作用并分析与心肌ＡＴＰ酶活力的关系。方法：离体大鼠工作心脏，全心缺血３０ｍｉｎ再灌４０ｍｉｎ，无机磷显色法分析ＡＴＰ酶活力。结果：顿抑心肌收缩舒张功能均明显降低，再灌期末ＬＶＳＰ×ＨＲ仅恢复５２％±８％，ＬＶＥＤＰ抬高２９８％±６４％，ＣＯ仅恢复４０％±８％，心肌细胞膜和线粒体Ｎａ＋，Ｋ＋－ＡＴＰ酶和Ｃａ２＋，Ｍｇ２＋－ＡＴＰ酶活力均下降，Ｔｅｔ（３０ｍｇ·ｋｇ－１·ｄ－１，ｉｐ，３ｄ，１０μｍｏｌ·Ｌ－１灌流缺血全程）可使ＬＶＳＰ×ＨＲ恢复８５％±１２％，ＬＶＥＤＰ仅抬高１６６％±４４％，ＣＯ恢复７５％±１１％，心肌细胞膜及线粒体Ｎａ＋，Ｋ＋－ＡＴＰ酶和Ｃａ２＋，Ｍｇ２＋－ＡＴＰ酶活力增高。结论：Ｔｅｔ对心肌缺血后顿抑损伤有一定保护作用，这一作用与维护顿抑期心肌细胞ＡＴＰ酶活力有关。     

Ca(2+) sensitization and diastolic function of normal and stunned porcine myocardium

Ca(2+) sensitizers prolong myofibrillar force development in vitro and might therefore aggravate relaxation abnormalities of stunned myocardium. This is the first in vivo study of the effects of the thiadiazinone derivative EMD 60263 ((+)-5-(l-(alpha-ethylimino-3, 4-dimethoxybenzyl)-1,2,3,4-tetrahydroquinoline-6-yl)-6-methyl-3, 6-dihydro-2H-1,3,4-thiadiazine-2-on), a Ca(2+)-sensitizing agent with negligible phosphodiesterase III inhibitory activity, on diastolic function of regionally stunned myocardium. After producing stunning by two sequences of 10-min coronary artery occlusion and 30 min of reperfusion, anaesthetised pigs received either saline (n=7) or 1.5 and 3.0 mg/kg of EMD 60263 (n=8) or its enantiomer EMD 60264 (n=6), which lacks the Ca(2+)-sensitizing properties but shares the bradycardiac action via inhibition of the delayed inward rectifier K(+) current. In stunned myocardium, systolic shortening was reduced to 46+/-4% of baseline (P<0.05) and mean rate of half end-diastolic segment lengthening, an index for diastolic function, to 35+/-4%; systolic shortening and mean rate of half end-diastolic lengthening of remote normal myocardium remained unchanged. Saline did not affect these parameters in stunned or normal myocardium. EMD 60264 did not affect systolic shortening but decreased mean rate of half end-diastolic lengthening in normal myocardium to 61+/-8% and in stunned myocardium to 16+/-5% of baseline. During saline and EMD 60264, normal and stunned segments started to lengthen immediately after minimal segment length was reached (DeltaT=0). Low dose EMD 60263 restored systolic shortening of the stunned region with no effect on DeltaT. The high dose increased systolic shortening above baseline and DeltaT to 210+/-30 ms in both regions. Consequently, mean rate of half end-diastolic lengthening increased to 66+/-11% in stunned, while decreasing to 55+/-3% in normal myocardium. After elimination of bradycardia, DeltaT and hence mean rate of half end-diastolic lengthening recovered in stunned myocardium, but in normal myocardium the latter remained depressed because DeltaT persisted. In conclusion, both doses of EMD 60263 improved systolic as well as diastolic function of stunned myocardium. The high dose delayed relaxation of normal myocardium without adversely affecting systolic function.