Ranolazine normalizes action potential repolarization of hypertrophied ventricular cardiomyocytes

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ABSTRACTS / Journal of Molecular and Cellular Cardiology 42 (2007) S1–S23

coronary infusion two times for 5 min, 5 min prior to the prolonged ischaemia. In some dogs samples were taken for Western blot analysis to determine 3-nitrotyrosine (3-NT) formation. Compared to the controls both PC and PN significantly reduced the numbers of ventricular premature beats (VPBs; 289 ± 84 vs. 93 ± 36 and 84 ± 34) and episodes of ventricular tachycardia (VT; 9.6 ± 3.4 vs. 1.9 ± 1.1 and 1.9 ± 1.7) as well as the incidence of ventricular fibrillation (VF; 50% vs. 0% and 13%) during prolonged occlusion. Furthermore both PC and PN increased survival (0% vs. 40% and 50%) and attenuated 3-nitrotyrosine formation following reperfusion. We conclude that PN provides protection, similar to PC, and this protection can be attributed to a reduced PN formation during the prolonged ischaemia. Acknowledgment The work is supported by OTKA (TO 37520). Keywords: Arrhythmias; Preconditioning; Free radicals doi:10.1016/j.yjmcc.2007.03.027

The effect of flecainide on conduction is not diminished in human atrial fibrillation Torsten Christ, Erich Wettwer, Ursula Ravens. Department of Pharmacology and Toxicology, Dresden University of Technology, Dresden, Germany Success rate of pharmacological cardioversion of atrial fibrillation (AF) with Class I antiarrhythmic drugs declines with duration of AF within few months. The reason for this increasing resistance of AF against sodium channel blocker treatment is unclear. Therefore, we have studied whether chronic AF influences the effect of the Class IC agent flecainide on action potentials parameters measured with conventional microelectrode technique in human right atrial trabeculae obtained from patients who were either in sinus rhythm (SR) or in chronic AF. In AF APD90 was smaller compared to SR (AF, 207 ± 9 ms, n = 6 vs. SR, 245 ± 13 ms, n = 6; p < 0.05). Resting membrane potential was significantly more negative in AF than in SR (AF, − 74 ± 1 mV vs. SR − 68 ± 2 mV, n = 6, p < 0.05). Maximum upstroke velocity (dV/dtmax) was slightly higher in AF than in SR (AF, 244 ± 14 V/s vs. SR 197 ± 15 V/s, n = 6; p < 0.05) whereas conduction time was not different. Flecainide decreased dV/dtmax to complete block with no difference in EC50 between SR and AF (SR, −log EC50 (M) 4.7 ± 0.2 vs. AF, 4.9 ± 0.3). Flecainide (10 μM) decreased action potential amplitude in SR from 93 ± 2 mV to 84 ± 3 mV and in AF from 105 ± 2 mV to 92 ± 4 mV and plateau potential in SR from − 15 ± 2 mV to − 20 ± 3 mV and in AF from 4 ± 6 mV to − 5 ± 5 mV. We conclude that the conduction delaying effects of flecainide are well preserved also in chronic AF. Our data therefore suggest that the Nachannel blocking activity of flecainide in chronic AF persists and that the lack of the drug's efficacy to cardiovert long-

standing AF is due to additional mechanisms most probably induced by ongoing electrical and structural remodeling. Keywords: Atrial fibrillation; Na+ channels; Antiarhythmic drugs doi:10.1016/j.yjmcc.2007.03.028

Comparison between the effects of Etomoxir and Ranolazine on cardiac arrhythmias Moslem Najafi, Tahereh Eteraf Oskouei. School of Pharmacy and Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran Comparison between the effects of Etomoxir (ETM) and Ranolazine (RAZ) on ischemia/reperfusion-induced arrhythmias was aimed. Isolated rat hearts were subjected to 30 min regional ischemia and 30 min reperfusion. The hearts were perfused with drug-free (control) or enriched Krebs solution with ETM (1 μM) or RAZ (20 μM) during ischemia/ reperfusion. The total number of ischemic ventricular ectopic beats (VEBs) in the control group was 667 ± 116 while perfusion of RAZ and ETM reduced it to 33 ± 16 (p < 0.001) and 501 ± 165, respectively. The number of ischemic ventricular tachycardia (VT) was also lowered by RAZ and ETM from 280 ± 50 (control) to 0 (p < 0.001) and 146 ± 50, respectively. Similarly, the number of reperfusion VEBs and VT were lowered by RAZ from 349 ± 73 and 154 ± 29 (control) to 86 ± 38 (p < 0.01) and 0 (p < 0.001), respectively. RAZ also reduced the incidence of total ventricular fibrillation (VF) and the time spent for reversible VF from 63% and 218 ± 69 s (control) to 0% (p < 0.05) and 0 sec (p < 0.01), respectively. At the same time, ETM showed significant anti-arrhythmic effects but in the lower extent in comparison with RAZ. Except a significant difference in the reduction of VEBs by RAZ compared to ETM, the effect was not significant between the agents. Regarding the inhibitory role of ETM and RAZ on fatty acid metabolism, their beneficial anti-arrhythmic effects are probably related to indirect increase in glucose oxidation. Keywords: Etomoxir; Ranolazine; Arrhythmias doi:10.1016/j.yjmcc.2007.03.029

Ranolazine normalizes action potential repolarization of hypertrophied ventricular cardiomyocytes Silvia Suffredini, Simona Brogioni, Elisabetta Cerbai, Luiz Belardinelli1, Alessandro Mugelli. CIMMBA, Unifi, Italy. 1CV Therapeutics, CA, USA Ranolazine (RAN) is a piperazine derivative recently approved as an add-on therapy for the treatment of patients with stable angina. Recent studies suggest that RAN reduces calcium overload in the ischemic myocyte by inhibiting the late sodium current. Thus, we studied the electrophysiological effect

ABSTRACTS / Journal of Molecular and Cellular Cardiology 42 (2007) S1–S23

of RAN on ventricular myocytes (VCMs) isolated from hypertrophied hearts of 15-month-old hypertensive rats (SHR), a model exhibiting marked electrophysiological alterations. Action potentials (APs) were recorded from SHR and normotensive WKY using the perforated-patch technique. APs were elicited at 0.2, 0.5 and 1 Hz and AP duration (APD) measured at − 50 mV (APD50) and 90% of repolarization (APD90). The effect of 3 and 10 μM RAN and 1 μM tetrodotoxin (TTX) were determined. In SHR VCMs, APD was significantly increased with respect to WKY (p < 0.05, n = 9). RAN caused a concentration-dependent decrease of APD in SHR VCMs. At 0.2 Hz, APD50 was 170 ± 21 ms in the absence and 133 ± 17 ms and 106 ± 18 ms in the presence of 3 μM and 10 μM RAN, respectively (p < 0.05, n = 13). Also APD90 was shortened by 3 and 10 μM RAN (p < 0.005, n = 13). The effect was similar at higher driving rates (0.5 or 1 Hz). In WKY VCMs, the effect of RAN was less pronounced and significantly smaller than in SHR. As a result, after exposure to 10 μM RAN, APD became similar in SHR and WKY VCMs (0.2 Hz; n = 9). The effect of RAN on SHR VCMs was mimicked by 1 μM TTX. Thus, increased Na entry through non-inactivating Na channels may contribute to delayed repolarization in hypertrophied myocytes, an alteration which is attenuated by RAN. Keywords: Hypertrophy; Ion channels; Cardiomyocytes doi:10.1016/j.yjmcc.2007.03.030

β-Subunits kchip2, kcne2 and dpp6 modulate effects of tedisamil on transient outward current ITO Susanne Radicke1, Diego Cotella2, Ursula Ravens1, Erich Wettwer1. 1Department of Pharmacology, Dresden, Germany. 2 Department of Medical Sciences, Novara, Italy The pharmacological sensitivity of ion-channel blockers is not only determined by the interaction with the ion-conducting α-subunit but also by accessory β-subunits. The aim of this study was to investigate whether β-subunits modify the effects of the antiarrhythmic agent tedisamil, a blocker of the transient outward current Ito encoded by Kv4.3 in human myocardium. Here we have examined the influence of β-subunits on the effects of tedisamil (30 μM) on Ito measured in CHO cells coexpressing the α-subunit Kv4.3 and the β-subunits KCNE2 and DPP6 (dipeptidyl-peptidase protein 6) with standard voltage-clamp technique. Tedisamil concentrations of half maximum inhibition (IC50) were either calculated from drug effects on charge carried by Ito (AUC) or on peak Ito. The IC50 values for Kv4.3+KChIP2 currents (AUC: 12 μM; peak: 16 μM) did not differ. In contrast, IC50 values for Kv4.3+KChIP2+KCNE2/DPP6 currents were lower for peak (3/9 μM) than for AUC (15/35 μM). Time course of Ito activation and inactivation was accelerated with the additional β-subunits KCNE2 and DPP6 under control conditions. Tedisamil did not affect the activation but apparently accelerated time course of inactivation in all channel combinations.

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Our data suggest that β-subunits modulate the effects of tedisamil on Ito current. We speculate that the larger potency of tedisamil in reducing peak Ito of channel complexes with the βsubunits KCNE2 and DPP6 reflects acceleration of block development. Keywords: Antiarrhythmics; Ion channel; Potassium doi:10.1016/j.yjmcc.2007.03.031

Cardioprotective role of prostaglandin E2 in ventricular cardiomyocytes involving Stat3 M.A. Frias, C. Gerber-Wicht, U. Lang The transcription factor Stat3 as well as cyclooxygenase (COX-2) products appear to be involved in adaptive growth and survival of cardiac myocytes. We have previously shown that in neonatal rat ventricular cardiomyocytes, the major COX-2 product prostaglandin E2 (PGE2) activates Stat3 leading to an increase in cell size and protein synthesis. In this study we determined whether PGE2 counteracts the apoptotic influence of the antitumor drug doxorubicin (doxo) in ventricular cardiomyocytes cells and whether Stat3 is involved. Cardiomyocytes were incubated with doxo (0.5 μM) and apoptosis was determined by investigating caspase3 activation and apoptotic DNA fragmentation. We found that doxo promoted a strong increase in caspase3 activation which, in the presence of PGE2 (1 μM), was reduced by approximately 70% and 44% after 4 and 24 h. Similarly, PGE2 inhibited the doxo-induced DNA fragmentation by 58 ± 5% after 24 h. We further showed that the specific inhibitors of ERK1/2, U0126 and PD98059, strongly reduced the antiapoptotic effect of PGE2 on doxo-induced caspase3 activation and DNA fragmentation. By contrast, the p38 MAP kinase inhibitor, SB203580, did not affect the antiapoptotic influence of PGE2. To evaluate the role Stat3, we transfected cardiomyocytes with a small interfering RNA (siRNA) targeting rat Stat3. This treatment resulted in 50– 60% silencing of Stat3, 48 h after transfection. In these Stat3downregulated cells, the inhibitory effect of PGE2 on doxoinduced caspase3 activation and DNA fragmentation was significantly reduced by approximately 50%. Thus, our result show that PGE2 plays a cardioprotective role by eliciting antiapoptotic responses in ventricular cardiomyocytes which are mediated through Stat3 involving the ERK1/2 but not the p38 MAP kinase pathway. Keywords: Apoptosis; Signal transduction; Cardiomyocytes doi:10.1016/j.yjmcc.2007.03.032

Mechanism of atrial reverse remodeling by combination of multi-ion channel blocker and angiotensine receptor blocker Hidehira Fukaya, Shinichi Niwano, Hiroe Niwano, Daisuke Satoh, Tohru Izumi. Department of Cardioangiology, Kitasato University, Japan