Raymond B. Darbenzio

Glyburide-reversible cardioprotective effect of BMS-180448 is independent of action potential shortening

OBJECTIVE We determine if action potential duration (APD) shortening and cardioprotection are separable phenomena in ATP-sensitive potassium channel (KATP) openers which protect ischemic myocardium via a glyburide-reversible mechanism. METHODS We determined the effect of the weakly vasodilating KATP opener, BMS-180448, and the less cardiac-selective cromakalim, on APD in normal, hypoxic or ischemic guinea pig papillary muscles or isolated hearts and compared their APD effects with their cardioprotective activity in isolated guinea pig hearts. RESULTS In isolated ischemic guinea pig hearts, cromakalim and BMS-180448 had similar cardioprotective potencies (EC25 of 3.2 and 3.3 microM, respectively, for increasing time to the onset of contracture). At 10 microM, BMS-180448 produced no APD shortening, yet was equally protective at this concentration compared to cromakalim, which produced profound APD shortening under either hypoxic or ischemic conditions. The cardioprotective effects of both compounds at 10 microM were abolished by 0.3 microM glyburide. CONCLUSIONS APD shortening is not correlated with cardioprotective activity for BMS-180448 and cromakalim while their cardioprotective effects are abolished by glyburide. These results suggest the possibility of reduced proarrhythmic activity in some KATP openers and that their cardioprotective activity is not associated with sarcolemmal KATP opening.

In vitro effects of capsaicin: antiarrhythmic and antiischemic activity

The antiarrhythmic effects of vehicle (0.1% dimethyl sulfoxide: DMSO) or capsaicin were evaluated in isolated perfused rat and guinea pig heart preparations. In the rat, capsaicin reduced ischemic ventricular tachycardia from 100% in control to 0%, and ischemic ventricular fibrillation from 60% in control to 0% at 30 microM, and diltiazem reduced the incidence of ischemic ventricular tachycardia and ventricular fibrillation to 55% and 0%, respectively. Reperfusion ventricular fibrillation was reduced from 90% to 20% and 33% for capsaicin and diltiazem, respectively, at these concentrations. In isolated perfused globally ischemic rat hearts, antiischemic efficacy was assessed as a significant extension (36% and 50%) in time to contracture with 30 microM capsaicin and 1 microM diltiazem, respectively. Capsaicin reduced left ventricular developed pressure by 35% in non-ischemic rat hearts, and increased coronary flow by 40%. The increased time to contracture for either compound was not blocked by glyburide (0.1 microM) suggesting a lack of any involvement of ATP-sensitive K+ channels. In isolated guinea pig hearts subjected to global ischemia, capsaicin and diltiazem reduced reperfusion ventricular fibrillation from 100% to 10% and 0% at 30 and 3 microM, respectively. Electrophysiologic evaluation in guinea pig papillary muscles using standard microelectrode techniques demonstrated significant (P < 0.05) action potential durations at 90% repolarization shortening at 1 Hz by 9%, 28% and 39%, and 23%, 37% and 51% at 10, 30, and 100 microM of capsaicin or diltiazem, respectively. Unlike diltiazem, no changes in action potential duration were observed with capsaicin (up to 100 microM) at faster stimulation rates (5 Hz). In conclusion, capsaicin displays both antiarrhythmic and antiischemic efficacy. These data suggest that the effects of capsaicin are mediated primarily through block of Ca2+ channels in these preparations.

Effects of Class III antiarrhythmic agents in an in vitro rabbit model of spontaneous torsades de pointe

Acquired long QT syndrome develops as a result of pharmacological interventions that prolong action potential duration. Excessive action potential prolongation may lead to torsade de pointes, a potentially fatal arrhythmia. To study this arrhythmia, in vivo models have been developed, but are difficult to interpret due to the complex nature of the intact metabolic, nervous and humoral systems. To more clearly examine the propensity of various Class III agents to elicit torsades de pointe, an in vitro model of spontaneous torsades de pointe was used in isolated perfused rabbit hearts. Male New Zealand white rabbits were anesthetized with sodium pentobarbital, and hearts isolated and perfused in a Langendorff apparatus. Electrocardiogram and epicardial monophasic action potentials were continuously recorded, and methoxamine (30 nM) and acetylcholine (0.3 microM) were given throughout the experiment. After 10 min of methoxamine and acetylcholine perfusion, Class III agents, dofetilide (0.1 to 0.7 microM), E-4031 (0.1 to 0.5 microM), D-sotalol (10 to 30 microM), or clofilium (0.1 to 0.3 microM), were given. All agents, except D-sotalol, induced torsades de pointe in 100% of hearts tested. D-Sotalol (30 microM) elicited a low incidence of torsades de pointe (25%). This could be explained by the limited prolongation of action potential duration with D-sotalol as compared to other Class III agents under these conditions. Dofetilide, a selective Class III agent, alone did not induce torsades de pointe. Nadolol (3 microM), a beta-adrenoceptor antagonist, increased the propensity of dofetilide to elicit torsades de pointe. In conclusion, increases in action potential duration (i.e., using Class III agents) in combination with a low heart rate (muscarinic receptor stimulation) and increases in intracellular Ca2+ (alpha-adrenoceptor stimulation) are needed to develop torsades de pointe in this model. Modulating these systems may provide us with new insights into preventing the initiation or maintenance of this arrhythmia.

Effects of cromakalim or pinacidil on pacing- and ischemia-induced ventricular fibrillation in the anesthetized pig

SummaryThe effects of the potassium channel openers (KCO), cromakalim or pinacidil, were evaluated in an anesthetized porcine model of pacing- and ischemia-induced ventricular fibrillation (VF). Hearts were paced at 180 bpm and the left anterior descending coronary artery was occluded until VF was induced.Reproducible times to VF (in seconds) were obtained allowing at least 20 min recovery following defibrillation. Cromakalim (0.3 mg/kg) or pinacidil (3 mg/kg) produced equivalent drops in mean arterial blood pressure. At these doses, cromakalim reduced monophasic action potential duration measured at 90% repolarization (APD90). Although time to VF in the cromakalim group was significantly greater than the vehicle treated group, it was not significantly different from its predrug value. In contrast, pinacidil reduced APD90, and significantly increased time to VF from 134±5 to 322±62 s (p<0.05). Neither cromakalim nor pinacidil affected whole-cell calcium currents recorded in guinea pig myocytes. During ischemia, cromakalim or pinacidil further reduced APD90; however, pinacidil had a two-fold greater effect than did cromakalim. The Class III antiarrhythmic agent, dofetilide, prolonged APD90, but did not increase time to VF.In conclusion, the increased time to VF observed with pinacidil coincides with its ability to shorten APD, and is consistent with activation of ATP-sensitive K+ channels (K+ATP). It is suggested that indirect reduction of calcium influx through K+ATP activation and APD shortening is sufficient to increase time to VF in this model. However, the inability of dofetilide to be effective suggests that this model would not be useful to test for Class III antiarrhythmic agents.

Hemodynamic and cardiac effects of BMS-180448, a novel K+ATP opener, in anesthetized dogs and isolated rat hearts.

Hemodynamic and cardiac effects of BMS-180448 (0.3-10 mg/kg i.v.) or cromakalim (0.01-0.3 mg/kg i.v.) were evaluated in anesthetized open-chest dogs and isolated perfused rat hearts. In the canine studies, heart rate (HR), mean arterial pressure and left ventricular pressure were measured as well as electromagnetic blood flows recorded from the aortic, renal, coronary and femoral vascular beds. BMS-180448 was 187-fold less potent than cromakalim in lowering blood pressure (ED-20 values of 7.84, and 0.042 mg/kg for BMS-180448 and cromakalim, respectively). Both compounds increased HR. Effects of BMS-180448 occurred at doses higher than those of cromakalim, but at doses slightly lower than those needed to cause hypotension (ED(HR)/ED(MABP) ratio of 0.18 for BMS-180448). BMS-180448 had no effect on myocardial contractility or relaxation over the doses studied, whereas cromakalim significantly increased +dP/dt and lowered -dP/dt. Effects on +dP/dt were associated with a decrease in blood pressure. Although BMS-180448 reduced total peripheral resistance (ED-25 = 5.75 mg/kg), it had little effect on specific vascular beds, with the exception of the coronary bed. BMS-180448, unlike cromakalim which caused more general vasodilating effects, appeared to be relatively selective in dilating the coronary vascular bed. In isolated perfused rat hearts, BMS-180448, 10-fold more potent as a cardioprotectant (EC25 = 2.7 microM) than as a cardiodepressant (ED-25 = 27.8 microM), had no effect on HR, suggesting a lack of effect of BMS-180448 on myocardial conduction. In conclusion, BMS-180448, a recently developed K+ATP opener, exerted less hypotensive and more selective vascular effects than did cromakalim. These results suggest that BMS-180448, at doses previously reported to give cardioprotection, should have a safe hemodynamic profile.

Effects of dofetilide on electrical dispersion and arrhythmias in post-infarcted anesthetized dogs

An increase in dispersion of myocardial refractoriness has been shown to coincide with a greater risk of inducible ventricular arrhythmias. We compared the dispersion of electrophysiologic parameters and antiarrhythmic effects of dofetilide (0.03, 0.1, 0.3 and 1 mg/kg i.v.) in postinfarcted anesthetized dogs. Animals were tested for inducibility of arrhythmias using a programmed electrical stimulation (PES) protocol, and divided into inducible (1) and non-inducible (NI) groups. In addition, myocardial vulnerability was measured using ventricular fibrillation thresholds (VFT), as well as susceptibility to sudden cardiac death (SCD). Dofetilide significantly increased ventricular effective refractory periods (ERP) and monophasic action potential durations (APD) in a dose-dependent manner. The standard deviation of ERP, which was used as an index of dispersion of refractoriness, increased from sham (control value of 5.4±sd2.5 ms), non-inducible (control value of 11.0 ±5.5 and 8.0±3.7 ms for vehicle and dofetilide groups, respectively) and inducible states (control value of 17.3±6.2 and 21.6±7.1 ms for vehicle and dofetilide groups, respectively). However, dofetilide treatment did not alter dispersion of refractoriness over the dose range studied. Dofetilide did not significantly increase inducibility in the NI group (2 out of 8[25%] compared to 0 out of 9 [0 %] in vehicle treated animals). In the I group, dofetilide (0.3 mg/kg) treated animals converted 2 out of 7 (29 %) to NI, and 5 out of 7 (71 %; significant at p<0.05) to a NI or non sustained ventricular tachycardia. There were no significant changes in VFT following the last dose of dofetilide given. Dofetilide did not significantly affect SCD survival (88 % and 29 % in the NI and I group, resectively) relauve to vehicle (66 % and 50 % in the NI and I group, respectively). Although infarct sizes were significantly greater in the I groups, there was no difference between vehicle and dofetilide animals within these groups. In conclusion, dofetilide increased ERP and APD values, but did not affect dispersion of refractoriness. Thus, changes in dispersion of refractori ness may be used as a marker for inducibility in untreated animals, but it did not predict the antiarrhythmic effects observed with dofetilide.

Effects of cromakalim or glibenclamide on arrhythmias and dispersion of refractoriness in chronically infarcted anesthetized dogs

The proarrhythmic effects of the ATP-sensitive potassium channel modulators cromakalim (n = 10; 0.01 to 0.3 mg/kg i.v.), glibenclamide (n = 10; 0.3 to 10 mg/kg i.v.) or volume equivalents of vehicle (n = 10) were evaluated in post-infarcted anaesthetised dogs. Dogs were anaesthetised, subjected to an anterior-apical myocardial infarction, and allowed to recover. At 7.4 ± 0.7 days post infarction, animals were anaesthetised again, electrophysiologic measurements (effective refractory periods, QT-intervals and ventricular fibrillation thresholds) were taken, and animals were tested for arrhythmias using a programmed electrical stimulation protocol. Only animals that did not have programmed electrical stimulation-inducible arrhythmias were used. Ventricular fibrillation thresholds were determined twice, once before the first dose then after the last dose of drug. At the end of the experiment, animals were subjected to ligation of the left circumflex coronary artery and survival was measured over the next two hours. Cromakalim significantly increased heart rate and decreased blood pressure. Although cromakalim significantly reduced effective refractory periods, it neither increased electrical dispersion, as determined by the standard deviation or coefficient of variance of the effective refractory period, nor did it enhance inducibility (0 out of 10 in both vehicle and cromakalim treated animals), change ventricular fibrillation thresholds, or reduce sudden death survival relative to vehicle. Glibenclamide did not increase electrical dispersion, but slightly increased the incidence of programmed electrical stimulation-induced arrhythmias (3 out of 10), and lowered ventricular fibrillation thresholds values. However, these changes were not statistically significant. Glibenclamide did not significantly affect survival relative to vehicle. Infarct sizes of the left ventricle were not statistically different among groups. In conclusion, cromakalim and glibenclamide did not affect dispersion of refractoriness. Glibenclamide did demonstrate a propensity towards proarrhythmic activity. However, the doses needed to observe proarrhythmic activity with glibenclamide were significantly higher than those needed for clinical treatment of hyperglycemia.