Role Of Gap Junction In Ventricular Arrhythmias Of Long QT Syndrome

Long QT syndrome (LQTS) is either an inherited or acquired disorder of delayed ventricular repolarization characterized by an excessively prolonged QT interval on the ECG. It often manifests clinically as recurrent syncope or sudden cardiac death as the consequence of polymorphic ventricular tachycardia known as torsades de pointes (TdP). LQTS is characterized by abnormally prolonged ventricular repolarization that predispose the affected individuals to arrhythmia, typically polymorphic ventricular tachycardia in the form of torsade de pointes (TdP) and an increased risk of sudden cardiac death.In addition to prolonged repolarization time, an increasing number of studies have indicated that amplified transmural dispersion of repolarization (TDR) is essential for the development of TdP in both congenital and acquired LQTS. The increase in TDR facilitates transmural propagation of EAD, produces trigger beat (i.e., R-on-T extrasystole) capable of initiating TdP, and also contributes to the maintenance of TdP by serving as a functional reentrant substrate. In vitro studies have revealed that TDR is a result of significant heterogeneity in the expression of ion channels among different cell types across the ventricular wall. Important electrical and functional heterogeneity exists between the myocytes from different regions of the heart.TDR is mainly attributed by significant heterogeneities of ion channel expression among different cell types, of which adjacent myocytes are coupled via gap junction. Gap junctions are composed of intercellular channels that allow the transfer of electrical current and small molecules between adjacent cells. The constituent proteins of gap junction channels, connexins, play a critical role in impulse propagation and electrical synchronization between myocytes. In an intact heart, such intrinsic electrophysiological heterogeneity diminishes due to the existence of gap junctions that permits movement of small molecules along the electrochemical gradient and thus helps in electrical synchronization of adjacent myocytes. In the intact heart where cells are well-coupled, transmural repolarization heterogeneity is significantly smaller than the intrinsic difference in action potential duration (APD) among these cells. Recent studies have found that expression and distribution of connexin43 (Cx43), the principle constitutional protein of gap junction in the ventricle, influence transmural electrical heterogeneities in both normal and failing hearts. Changes in phosphorylation of connexin can result in gap junction uncoupling by its effects on the gating of gap junction channels and connexin turnover dynamics. A gap junction uncoupler, carbenoxolone, enhances dispersion of repolarization, whereas high-degree coupling between myocardial layers diminishes or eradicates transmural dispersion in repolarization time and masks M cells. Agents such as antiarrhythmic peptide 10 (AAP10) and antiarrhythmic peptide HP-5, which facilitate the function of gap junctions, reduce dispersion of action potential duration (APD) and may have antiarrhythmic action. Also, augmentation of gap junction coupling can prevent abnormal amplification of TDR.Increased TDR and uncoupling of the gap junctions are observed in many heart diseases such as ventricular hypertrophy and heart failure. All of these findings indicate that changes in intercellular coupling of gap junctions can modify the intrinsic difference in repolarization properties of cells spanning the ventricular wall. Therefore, we hypothesized that increasing gap junction coupling may be capable of reducing transmural heterogeneities of repolarization and hence, preventing ventricular arrhythmias under conditions of LQTS. To test this hypothesis, we investigated the effects of AAP10, a gap junction enhancer, on TDR and induction of TdP in a rabbit LQT3 and canine LQT2 model. Objective Increased transmural dispersion of repolarization (TDR) contributes importantly to the development of torsades de pointes (TdP) in long QT syndrome (LQTS). Intercellular electrical coupling via gap junctions plays an important role in maintaining TDR in both normal and diseased hearts. This study examined the effects of antiarrhythmic peptide AAP10, a gap junction enhancer, on TDR and induction of TdP in a rabbit LQT3 model.Methods An arterially perfused rabbit left ventricular preparation and sea anemone toxinⅡ(ATX-Ⅱ,20 nM) were used to establish a LQT3 model. Transmural ECG as well as action potentials from both endocardium and epicardium were simultaneously recorded. Changes in nonphosphorylated connexin43 (Cx43) were measured by immunoblotting.The ventricular wedge preparations were allowed to equilibrate in the tissue bath for 1 hour prior to electrical recordings. Unless noted otherwise, all drugs used in this study were dissolved in Tyrode's solution and infused into the wedge preparation via the cannulated artery. ATX-II (20 nM, Calbiochem Corp., La Jolla, CA, USA) was used to augment the late INa and create a model of LQT3. AAP10 (Chinese Peptide Co., Hangzhou, Zhejiang, China), a gap junction enhancer, was used to enhance gap junction coupling. After baseline data acquisition, preparations were divided into four groups:Control group (n=10):wedge preparations were kept perfusion with Tyrode's solution; LQT3 group (n=10):wedge preparations were perfused with ATX-II (20 nM) to mimic a LQT3 situation; AAP-100nM group (n=10) and AAP-500nM group (n=10):15 minutes prior to ATX-II administration, wedge preparations were pretreated with 100 nM and 500 nM AAP10, respectively.The development of spontaneous and programmed electrical stimulation (PES)-induced TdP was assessed at baseline conditions, pretreatment of AAP10, and in the presence of ATX-II. PES-induced arrhythmias were evaluated by use of single extrastimulus (S2) applied to the endocardial surface of the wedge. The QT interval was defined as the time from the onset of the QRS to the point at which the final downslope of the T wave crossed the isoelectric line. APD was measured at 90% repolarization (APD90). TDR was defined as the difference between the endocardial and the epicardial epolarization times across the left ventricular wall.Results Compared with the control group, the QT interval, TDR, early afterdepolariztion (EAD), R-on-T extrasystole, and TdP increased sharply with augmented nonphosphorylated Cx43 in the LQT3 group (P<0.001 for both). EAD and R-on-T extrasystole were induced by ATX-II (20 nM) in 10 of 10 preparations (P<0.001 vs. control) at a BCL of 2000 ms, among which five (P=0.033 vs. control) spontaneously degenerated into TdP in the LQT3 group. A single extrastimulus reproducibly induced TdP in nine of 10 preparations in the LQT3 group (P<0.001 vs. control). Interestingly, compared with the LQT3 group,500 nM AAP10 reduced QT interval, TDR (P<0.001 for both), and prevented EAD, R-on-T extrasystole, and TdP (P=0.003, P=0.001, P=0.02) with a parallel decrease in nonphosphorylated Cx43 in the presence of ATX-Ⅱ(P<0.001).Conclusions Gap junction enhancer AAP10 is capable of abbreviating the QT interval, reducing TDR, and suppressing TdP in a rabbit LQT3 model probably via its effect by preventing dephosphorylation of Cx43. These data suggest that increasing intercellular coupling may reduce TDR and, therefore, prevent TdP in LQTS. Objective Gap junctions contribute to the transmural heterogeneity of repolarization in the normal heart and under conditions of prolonged QT interval in the diseased heart. This study examined whether enhancing of gap junction coupling can reduce transmural dispersion of repolarization (TDR) and prevent torsade de pointes (TdP) in a canine LQT2 model.Methods Canine left ventricular wedge preparations were perfused with delayed rectifier potassium current (IKr) blocker d-sotalol to mimic LQT2 and the antiarrhythmic peptide 10 (AAP10) was used as a gap junction coupling enhancer. Western blot and immunofluorescence were used for examining the expression and distribution of Cx43.The ventricular wedge preparations were allowed to equilibrate in the tissue bath for 1 h prior to electrical recordings. Unless noted otherwise, all drugs used in this study were dissolved in Tyrode's solution and delivered to the wedge preparation via the cannulated artery. d-Sotalol (Sigma, St. Louis, MO, USA), a blocker of the rapidly activating delayed rectifier potassium current (IKr), was used to create the LQT2 model. Antiarrhythmic peptide 10 (AAP10), a peptide that enhances gap junction coupling, was obtained from Chinese Peptide (Hangzhou, Zhejiang, China).Three groups of experiments were carried out. Control group (n=10):wedge preparations were kept perfusion with Tyrode's solution; LQT2 group (n=10):wedge preparations were perfused with d-sotalol (100μM) to mimic a LQT2 situation;AAP10 group (n=10):wedge preparations were pretreated with 500 nM AAP10 for 15 min, then perfused with both d-sotalol (100μM) and AAP10 (500 nM). Development of spontaneous and programmed electrical stimulation (PES)-induced TdP was assessed at baseline conditions, after pretreatment of AAP10, and in the presence of d-sotalol. The QT interval was defined as the time from the onset of the QRS to the point atwhich the final downslope of the T wave crossed the isoelectric line. APD was measured at 90% repolarization (APD90). TDR was defined as the difference between the longest and shortest repolarization times of intracellular action potentials recorded across the ventricular wall.Results As compared with the control group, the LQT2 group had significantly augmented TDR and higher incidence of TdP associated with increased nonphosphorylated connexin 43 (Cx43). AAP10 prevented augmentation of TDR and induction of TdP while rescuing Cx43 phosphorylation. There was no significant change in the quantity and spatial distribution of Cx43.Pretreatment with AAP10 significantly attenuated effects of d-sotalol on APD90 in the M cells (P<0.001), but not in the endocardial or epicardial cells. As a net effect, the QT interval and TDR were significantly lower in the AAP 10 group as compared with the LQT2 group (P<0.001). AAP10 pretreatment completely suppressed spontaneous TdP (from 4/10 to 0/10; P=0.087) and significantly reduced PES-induced TdP episodes (7/10 in d-sotalol group versus 1/10 in AAP10+d-sotalol group; P=0.02). Pretreatment with AAP 10 did not alter QT interval, APD90, TDR, or QRS duration (P=NS). No arrhythmia was observed in the samples receiving AAP 10 alone.Conclusions The present study showed that gap junction enhancer AAP 10 reduces TDR and incidence of TdP by preventing dephosphorylation of Cx43 in a canine ventricular model of LQT2. These results imply that dephosphorylation of Cx43 may be involved in the genesis of transmural heterogeneity of repolarization, and ultimately ventricular arrhythmias in LQT2.