Canine Left Ventricular Wedge Preparation Research Articles

Effect of Flecainide and Ibutilide Alone and in Combination to Terminate and Prevent Recurrence of Atrial Fibrillation.

There is a need for improved approaches to rhythm control therapy of atrial fibrillation (AF). The effectiveness of flecainide (1.5 µmol/L) and ibutilide (20 nmol/L), alone and in combination, to cardiovert and prevent AF recurrence was studied in canine-isolated coronary-perfused right atrioventricular preparations. We also examined the safety of the combination of flecainide (1.5 µmol/L) and ibutilide (50 nmol/L) using canine left ventricular wedge preparations. Sustained AF (>1 hour) was inducible in 100%, 60%, 20%, and 0% of atria in the presence of acetylcholine alone, acetylcholine+ibutilide, acetylcholine+flecainide, and acetylcholine+ibutilide+flecainide, respectively. When used alone, flecainide and ibutilide cardioverted sustained AF in 40% and 20% of atria, respectively, but in 100% of atria when used in combination. Ibutilide prolonged atrial and ventricular effective refractory period by 15% and 8%, respectively, at a cycle length of 500 ms (P<0.05 for both). Flecainide increased the effective refractory period in atria by 27% (P<0.01) but by only 2% in the ventricles. The combination of the 2 drugs lengthened the effective refractory period by 42% in atria (P<0.01) but by only 7% (P<0.05) in the ventricles. In left ventricular wedges, ibutilide prolonged QT and Tpeak-Tend intervals by 25 and 55%, respectively (P<0.05 for both; cycle length, 2000 ms). The addition of flecainide (1.5 µmol/L) partially reversed these effects (P<0.05 for both parameters versus ibutilide alone). Torsades de Pointes score was relatively high with ibutilide alone and low with the drug combination. In our experimental model, a combination of flecainide and ibutilide significantly improves cardioversion and prevents the recurrence of AF compared with monotherapies with little to no risk for the development of long-QT-mediated ventricular proarrhythmia.

PO-01-148 COMBINATION OF FLECAINIDE AND IBUTILIDE FOR EFFECTIVE AND SAFE MANAGEMENT OF ATRIAL FIBRILLATION

There is a need for improved approaches to rhythm control therapy of atrial fibrillation (AF). To investigate a novel pharmacological approach for rhythm control management of AF. We studied the effectiveness of clinically relevant concentrations of flecainide (1.5 μM) and ibutilide (20 nM), alone and in combination, to prevent the induction of AF and to cardiovert persistent AF, defined here as AF lasting > 1 hr, using canine isolated coronary-perfused right atrial preparations. In addition, we examined the safety of the flecainide and ibutilide (50 nM) combination using canine left ventricular (LV) wedge preparations. Persistent AF could be induced in 100% of preparations following exposure to acetylcholine (0.5 μM; n=5). Persistent AF remained inducible in 60% of atria in the presence of ibutilide alone and in 20% of preparations in the presence of flecainide alone, but in 0% of preparations in the presence of the combination of ibutilide and flecainide (n=5 per group). When used alone, flecainide and ibutilide cardioverted persistent AF in 40% and 20% of atria, respectively, but in 100% of atria when used in combination (n=5 per group). Ibutilide prolonged both atrial and ventricular effective refractory period (ERP; by 15 and 8%, respectively; p<0.05 for both; Cycle length [CL] = 500 ms; n=5 for each). Flecainide significantly prolonged ERP in atria (by 27%; p<0.001) but not in ventricles (2% increase) due to atrial predominant induction of post-repolarization refractoriness. The combination of the two drugs prolonged ERP by 42% in atria (p<0.001 vs. control and 0<0.05 vs. flecainide alone) but by only 7% in ventricles (p<0.05; n=5, CL=500 ms). In separate experiments involving LV wedge preparations, risk factors for Torsade de Pointes were accentuated by ibutilide (50 nM) and slowed pacing (CL= 2000 ms; n=5). Under these conditions, ibutilide induced significant prolongation of QT and Tpeak /Tend intervals (by 25 and 55%, respectively; p<0.01 for both; n=5). Further addition of flecainide (1.5 μM) partially reversed these effects leading to an increase of these parameters by only 15 and 19%, respectively; p<0.05, combination vs. ibutilide; n=5). Torsade de Pointes was never observed with the drug combination. Our experimental data indicate that a combination of flecainide and ibutilide significantly improves pharmacological cardioversion of AF as well as prevention of AF with very little or no risk of long QT-mediated ventricular proarrhythmia.

Transmural conduction time in an early repolarization syndrome model.

In the pathological aspect of J wave syndrome, delayed depolarization is defined as the difference in local conduction velocity of the ventricular myocardium. If polymorphic ventricular tachycardia is induced without local conduction velocity heterogeneity, this contradicts the delayed depolarization theory. In the present study, the transmural conduction time at was evaluated at several transmural locations in a canine early repolarization model. The transmural pseudo-electrocardiogram and endocardial/epicardial action potentials were recorded from coronary-perfused canine left ventricular wedge preparations (n=18). The Ito agonist NS5806 (9-10 µM), Ca2+ channel blocker verapamil (2 µM) and acetylcholine (ACh) (2 µM) were used to pharmacologically mimic early repolarization syndrome genotypes. The transmural conduction times were measured at five fixed epicardial unipolar electrodes before and after the perfusion of provocative agents. The transmural conduction time was defined as the time from endocardial stimulation to the maximal negative deflection (dV/dt) of the endocardial electrogram at the unipolar electrode. Polymorphic ventricular tachycardia developed in 14/18 preparations. In the transmembrane action potentials, there was no definite delayed phase 0 upstroke in any induced polymorphic ventricular tachycardia preparations. In all preparations, the transmural conduction time increased significantly after perfusing the Ito agonist NS5806, verapamil and Ach; however, the increase was only 2.6±0.4 msec, and dispersion of the transmural conduction time did not exhibit significant heterogeneity (7.16±0.93 vs. 7.76±1.21 msec; P=0.240). In the early repolarization model, polymorphic ventricular tachycardia was induced without any regional conduction velocity heterogeneity. This finding suggests that local depolarization heterogeneity would not be a major contributor to the generation of ventricular arrhythmia in the early repolarization syndrome wedge preparation model.

Experimental verification of the value of the Tpeak -Tend interval in ventricular arrhythmia inducibility in an early repolarization syndrome model.

In patients with early repolarization patterns on ECG, many researchers have studied to find predictors of fatal arrhythmia. However, there are no satisfying clinical predictors. We evaluated the value of the Tpeak -Tend interval on pseudo-ECG in canine myocardial wedge preparation models of early repolarization syndrome. Transmural pseudo-ECG and endocardial/epicardial action potentials were recorded from coronary-perfused canine left ventricular wedge preparations (n = 34). The Ito agonist NS5806 (8-10 µM), the calcium channel blocker verapamil (3 µM) and acetylcholine (2-3 µM) were used to mimic the disease model. A ventricular arrhythmia induction test was performed. QTpeak , QTend , Tpeak -Tend , and Tpeak -Tend /QTend were measured at 15 to 20 minutes after the provocative agent infusion. Polymorphic ventricular tachycardias (pVT) developed in 23 of the 34 preparations (67%). The maximal values of Tpeak -Tend and Tpeak -Tend /QTend were recorded just before pVT induction. At baseline, without the provocative agents, Tpeak -Tend and Tpeak -Tend /QTend were not different between pVT-induced and pVT-noninduced preparations. The Tpeak -Tend of the pVT-induced preparations was longer than that of non-induced preparations (58 ± 26.8 msec vs 33 ± 6.8 msec, P < .001). The Tpeak -Tend /QTend of pVT- induced preparations was larger than that of noninduced preparations (0.220 ± 0.1017 vs 0.128 ± 0.0312, P < .001). The transmural and epicardial dispersion of repolarization of pVT-induced preparations were larger than those of pVT-noninduced preparations. The transmural dispersion of repolarization showed a positive correlation with Tpeak -Tend . Tpeak -Tend predicted malignant ventricular arrhythmias in early repolarization syndrome models. Tpeak -Tend reflects the repolarization heterogeneity of ventricular myocardium.

Epicardial Substrate as a Target for Radiofrequency Ablation in an Experimental Model of Early Repolarization Syndrome.

Early repolarization syndrome (ERS) is an inherited cardiac arrhythmia syndrome associated with sudden cardiac death. Approaches to therapy are currently very limited. This study probes the mechanisms underlying the electrocardiographic and arrhythmic manifestation of experimental models of ERS and of the ameliorative effect of radiofrequency ablation. Action potentials, bipolar electrograms, and transmural pseudo-ECGs were simultaneously recorded from coronary-perfused canine left ventricular wedge preparations (n=11). The Ito agonist NS5806 (7-10 μmol/L), calcium channel blocker verapamil (3 μmol/L), and acetylcholine (1-3 μmol/L) were used to pharmacologically mimic the effects of genetic defects associated with ERS. The provocative agents induced prominent J waves in the ECG secondary to accentuation of the action potential notch in epicardium but not endocardium. Bipolar recordings displayed low-voltage fractionated potentials in epicardium because of temporal and spatial variability in appearance of the action potential dome. Concealed phase 2 reentry developed when action potential dome was lost at some epicardial sites but not others, appearing in the bipolar electrogram as discrete high-frequency spikes. Successful propagation of the phase 2 reentrant beat precipitated ventricular tachycardia/ventricular fibrillation. Radiofrequency ablation of the epicardium destroyed the cells displaying abnormal repolarization and thus suppressed the J waves and the development of ventricular tachycardia/ventricular fibrillation in 6/6 preparations. Our findings suggest that low-voltage fractionated electrical activity and high-frequency late potentials recorded from the epicardial surface of the left ventricle can identify regions of abnormal repolarization responsible for ventricular tachycardia/ventricular fibrillation in ERS and that radiofrequency ablation of these regions in left ventricular epicardium can suppress ventricular tachycardia/ventricular fibrillation by destroying regions of ER.

Cellular and ionic mechanisms underlying the effects of cilostazol, milrinone, and isoproterenol to suppress arrhythmogenesis in an experimental model of early repolarization syndrome

Early repolarization syndrome (ERS) is associated with polymorphic ventricular tachycardia (PVT) and ventricular fibrillation, leading to sudden cardiac death. The present study tests the hypothesis that the transient outward potassium current (Ito)-blocking effect of phosphodiesterase-3 (PDE-3) inhibitors plays a role in reversing repolarization heterogeneities responsible for arrhythmogenesis in experimental models of ERS. Transmembrane action potentials (APs) were simultaneously recorded from epicardial and endocardial regions of coronary-perfused canine left ventricular (LV) wedge preparations, together with a transmural pseudo-electrocardiogram. The Ito agonist NS5806 (7-15 μM) and L-type calcium current (ICa) blocker verapamil (2-3 μM) were used to induce an early repolarization pattern and PVT. After stable induction of arrhythmogenesis, the PDE-3 inhibitors cilostazol and milrinone or isoproterenol were added to the coronary perfusate. All were effective in restoring the AP dome in the LV epicardium, thus abolishing the repolarization defects responsible for phase 2 reentry and PVT. Arrhythmic activity was suppressed in 7 of 8 preparations by cilostazol (10 μM), 6 of 7 by milrinone (2.5 μM), and 7 of 8 by isoproterenol (0.1-1 μM). Using voltage clamp techniques applied to LV epicardial myocytes, both cilostazol (10 μM) and milrinone (2.5 μM) were found to reduce Ito by 44.4% and 40.4%, respectively, in addition to their known effects to augment ICa. Our findings suggest that PDE-3 inhibitors exert an ameliorative effect in the setting of ERS by producing an inward shift in the balance of current during the early phases of the epicardial AP via inhibition of Ito as well as augmentation of ICa, thus reversing the repolarization defects underlying the development of phase 2 reentry and ventricular tachycardia/ventricular fibrillation.

Cellular Mechanism Underlying Hypothermia-Induced Ventricular Tachycardia/Ventricular Fibrillation in the Setting of Early Repolarization and the Protective Effect of Quinidine, Cilostazol, and Milrinone

Hypothermia has been reported to induce ventricular tachycardia and fibrillation (VT/VF) in patients with early repolarization (ER) pattern. This study examines the cellular mechanisms underlying VT/VF associated with hypothermia in an experimental model of ER syndrome and examines the effectiveness of quinidine, cilostazol, and milrinone to prevent hypothermia-induced arrhythmias. Transmembrane action potentials were simultaneously recorded from 2 epicardial and 1 endocardial site of coronary-perfused canine left ventricular wedge preparations, together with a pseudo-ECG. A combination of NS5806 (3-10 μmol/L) and verapamil (1 μmol/L) was used to pharmacologically model the genetic mutations responsible for ER syndrome. Acetylcholine (3 μmol/L) was used to simulate increased parasympathetic tone, which is known to promote ER. In controls, lowering the temperature of the coronary perfusate to induce mild hypothermia (32°C-34°C) resulted in increased J-wave area on the ECG and accentuated epicardial action potential notch but no arrhythmic activity. In the setting of ER, hypothermia caused further accentuation of the epicardial action potential notch, leading to loss of the action potential dome at some sites but not others, thus creating the substrate for development of phase 2 reentry and VT/VF. Addition of the transient outward current antagonist quinidine (5 μmol/L) or the phosphodiesterase III inhibitors cilostazol (10 μmol/L) or milrinone (5 μmol/L) diminished the ER manifestations and prevented the hypothermia-induced phase 2 reentry and VT/VF. Hypothermia leads to VT/VF in the setting of ER by exaggerating repolarization abnormalities, leading to development of phase 2 reentry. Quinidine, cilostazol, and milrinone suppress the hypothermia-induced VT/VF by reversing the repolarization abnormalities.

Mechanisms underlying the development of the electrocardiographic and arrhythmic manifestations of early repolarization syndrome

Early repolarization pattern in the ECG has been associated with increased risk for ventricular tachycardia/fibrillation (VT/VF), particularly when manifest in inferior leads. This study examines the mechanisms underlying VT/VF in early repolarization syndrome (ERS). Transmembrane action potentials (APs) were simultaneously recorded from 2 epicardial sites and 1 endocardial site of coronary-perfused canine left-ventricular (LV) wedge preparations, together with a pseudo-ECG. Transient outward current (Ito) was recorded from epicardial myocytes isolated from the inferior and lateral LV of the same heart. J wave area (pseudo-ECG), epicardial AP notch magnitude and index were larger in inferior vs. lateral wall preparations at baseline and after exposure to provocative agents (NS5806+verapamil+acetylcholine (ACh)). Ito density was greater in myocytes from inferior vs. lateral wall (18.4±2.3pA/pF vs. 11.6±2.0pA/pF; p<0.05). A combination of NS5806 (7μM) and verapamil (3μM) or pinacidil (4μM), used to pharmacologically model the genetic defects responsible for ERS, resulted in prominent J-point and ST-segment elevation. ACh (3μM), simulating increased vagal tone, precipitated phase-2-reentry-induced polymorphic VT/VF. Using identical protocols, inducibility of arrhythmias was 3-fold higher in inferior vs. lateral wedges. Quinidine (10μM) or isoproterenol (1μM) restored homogeneity and suppressed VT/VF. Our data support the hypothesis that 1) ERS is caused by a preferential accentuation of the AP notch in the LV epicardium; 2) this repolarization defect is accentuated by elevated vagal tone; 3) higher intrinsic levels of Ito account for the greater sensitivity of the inferior LV wall to development of VT/VF; and 4) quinidine and isoproterenol exert ameliorative effects by reversing the repolarization abnormality.

Abstract 214: Therapeutic Hypothermia Promotes Arrhythmogenic Calcium-Mediated Delayed Afterdepolarizations During Sympathetic Stimulation

Introduction: Most patients who undergo therapeutic hypothermia (TH) suffer cardiac arrest due to ischemia-induced ventricular fibrillation (VF). Ischemia and hypothermia both cause cellular calcium (Ca) overload, a substrate for Ca-mediated triggered arrhythmias. Previously, we showed that under non-ischemic conditions, TH suppresses Ca-mediated arrhythmias. However, the mechanistic relationship between hypothermia-induced Ca dysregulation and arrhythmias during ischemia, the most common clinical scenario, is unknown. We hypothesized that TH and ischemia would synergistically promote spontaneous Ca release and subsequent arrhythmogenic delayed afterdepolarizations (DADs). We examined this relationship during sympathetic stimulation, as occurs when epinephrine is given during resuscitation from VF. Methods and Results: Dual optical mapping of Ca transients and action potentials was performed in canine left ventricular wedge preparations during 10 minutes of no flow ischemia during normothermia (NT, 36°C, n=5) and TH (32°C, n=7) with and without beta-sympathetic stimulation (isoproterenol, 0.2uM, each n=5). Multicellular spontaneous Ca release events (mSCRs) and resultant DADs were induced by rapid pacing. During ischemia, TH increased mSCR amplitude (44±3%) vs. NT (26±4%, p&lt;0.04). In contrast, the slope (amplitude/rise time) of mSCRs, an important determinant of triggered activity (TA), and DAD amplitudes were similar in TH and NT (24±6 and 21%±2% respectively, both p=NS). Importantly, when sympathetic stimulation was present during ischemia, TH increased the amplitude mSCRs compared to NT (61±5 vs. 31±7%, respectively p&lt;.01) and the slope of mSCRs was 2-fold greater (p&lt;.01), resulting in greater amplitude of DADs (50±9 vs. 28±7%, p=.08) and TA was observed. Conclusions: During ischemia, TH is not arrhythmogenic unless sympathetic stimulation is introduced, where it promotes spontaneous Ca release and DADs. Moreover, these data suggest that when TH is initiated during ischemia, as in intra-arrest cooling, sympathetic stimulation during cardiac arrest may be proarrhythmic by increasing susceptibility to Ca-mediated triggered arrhythmias.

Enhanced Dispersion of Repolarization Explains Increased Arrhythmogenesis in Severe Versus Therapeutic Hypothermia

Hypothermia is proarrhythmic, and, as the use of therapeutic hypothermia (TH) increases, it is critically important to understand the electrophysiological effects of hypothermia on cardiac myocytes and arrhythmia substrates. We tested the hypothesis that hypothermia-enhanced transmural dispersion of repolarization (DOR) is a mechanism of arrhythmogenesis in hypothermia. In addition, we investigated whether the degree of hypothermia, the rate of temperature change, and cooling versus rewarming would alter hypothermia-induced arrhythmia substrates. Optical action potentials were recorded from cells spanning the transmural wall of canine left ventricular wedge preparations at baseline (36°C), during cooling and during rewarming. Electrophysiological parameters were examined while varying the depth of hypothermia. On cooling to 26°C, DOR increased from 26±4 ms to 93±18 ms (P=0.021); conduction velocity decreased from 35±5 cm/s to 22±5 cm/s (P=0.010). On rewarming to 36°C, DOR remained prolonged, whereas conduction velocity returned to baseline. Conduction block and reentry was observed in all severe hypothermia preparations. Ventricular fibrillation/ventricular tachycardia was seen more during rewarming (4/5) versus cooling (2/6). In TH (n=7), cooling to 32°C mildly increased DOR (31±6 to 50±9, P=0.012), with return to baseline on rewarming and was associated with decreased arrhythmia susceptibility. Increased rate of cooling did not further enhance DOR or arrhythmogenesis. Hypothermia amplifies DOR and is a mechanism for arrhythmogenesis. DOR is directly dependent on the depth of cooling and rewarming. This provides insight into the clinical observation of a low incidence of arrhythmias in TH and has implications for protocols for the clinical application of TH.

Is there a significant transmural gradient in repolarization time in the intact heart?

Is there a significant transmural gradient in repolarization time in the intact heart?

Pharmacological Enhancement of Cardiac Gap Junction Coupling Prevents Arrhythmias in Canine LQT2 Model

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. 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. 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. These data indicate that gap junction enhancer AAP10 can prevent augmentation of TDR and suppress TdP by preventing dephosphorylation of Cx43 in a LQT2 model.

Ventricular transmural repolarization sequence: its relationship with ventricular relaxation and role in ventricular diastolic function

We investigated the role of ventricular repolarization sequence in ventricular diastolic function. Arterially perfused canine left ventricular wedge preparation with simultaneous recording of action potentials and isometric contractile force was used to establish the relationship between ventricular repolarization and relaxation sequences. An isolated rabbit working heart model was used to investigate role of ventricular repolarization sequence in ventricular diastolic function. Under controlled conditions, similar to transmural dispersion of repolarization (TDR), there existed a time difference between initiation of epicardial and endocardial relaxation (TR(Epi-Endo), 47.4 +/- 13.9 ms) with epicardium relaxing earlier. There was a strong correlation between TDR and TR(Epi-Endo) (r(2) = 0.99, n = 5) and the interventions that changed transmural repolarization sequence led to parallel changes in transmural relaxation sequence. In isolated rabbit working hearts, reversal of the transmural repolarization sequence that manifested as negative T wave was associated with a significant increase in isovolumic relaxation time (from 49.2 +/- 19.1 to 76.4 +/- 12.1 ms, n = 5, P = 0.001). There is a strong correlation between transmural repolarization and relaxation sequences. A positive T wave that denotes transmural repolarization sequence from epicardium to endocardium is essential for normal diastolic function of ventricle and the reversal of such sequence is associated with ventricular diastolic dysfunction.

Cellular basis for arrhythmogenesis in an experimental model of the SQT1 form of the short QT syndrome

Short QT syndrome (SQTS) is a primary electrical disease of the heart associated with a high risk of sudden cardiac death. A gain-of-function in I(Kr), due to a mutation in KCNH2, underlies SQT1. This study sought to examine the cellular basis for arrhythmogenesis in an experimental model of SQT1 created using PD-118057, a novel I(Kr) agonist. Transmembrane action potentials were simultaneously recorded from epicardial, M, and endocardial regions of arterially perfused canine left ventricular (LV) wedge preparations, together with a pseudo-electrocardiogram. PD-118057 (10 micromol/l) abbreviated the QT interval from 267 +/- 4 to 232 +/- 4 ms and increased transmural dispersion of repolarization (TDR) from 33.7 +/- 2.0 to 49.1 +/- 3.1 ms (P <.001). T-wave amplitude increased from 18.0% +/- 1.4% to 23.1% +/- 1.7% of R-wave amplitude (P =.027). Reversing the direction of activation of the LV wall (epicardial pacing) resulted in an increase in QT interval from 269 +/- 5 to 282 +/- 5 ms and an increase in TDR from 34.1 +/- 2.0 to 57.6 +/- 3.3 ms (P <.001) under baseline conditions. PD-118057 abbreviated the QT interval from 282 +/- 5 to 258 +/- 5 ms and produced a proportional decrease in effective refractory period (ERP). TDR increased from 57.6 +/- 3.3 to 77.6 +/- 4.3 ms (P <.001). Polymorphic ventricular tachycardia (pVT) was induced in 10 of 20 preparations with a single S(2) applied to epicardium. Quinidine (10 micromol/l) increased the ERP and QT interval, did not significantly alter TDR, and prevented induction of pVT in 5 of 5 preparations. Our results suggest that a combination of ERP abbreviation and TDR amplification underlie the development of pVT in SQT1 and that quinidine prevents pVT principally by prolonging ERP.

Ryanodine receptor dysfunction and triggered activity in the heart

Arrhythmogenesis has been increasingly linked to cardiac ryanodine receptor (RyR) dysfunction. However, the mechanistic relationship between abnormal RyR function and arrhythmogenesis in the heart is not clear. We hypothesize that, under abnormal RyR conditions, triggered activity will be caused by spontaneous calcium release (SCR) events that depend on transmural heterogeneities of calcium handling. We performed high-resolution optical mapping of intracellular calcium and transmembrane potential in the canine left ventricular wedge preparation (n = 28). Rapid pacing was used to initiate triggered activity under normal and abnormal RyR conditions induced by FKBP12.6 dissociation and beta-adrenergic stimulation (20-150 microM rapamycin, 0.2 microM isoproterenol). Under abnormal RyR conditions, almost all preparations experienced SCRs and triggered activity, in contrast to control, rapamycin, or isoproterenol conditions alone. Furthermore, under abnormal RyR conditions, complex arrhythmias (monomorphic and polymorphic tachycardia) were commonly observed. After washout of rapamycin and isoproterenol, no triggered activity was observed. Surprisingly, triggered activity and SCRs occurred preferentially near the epicardium but not the endocardium (P < 0.01). Interestingly, the occurrence of triggered activity and SCR events could not be explained by cytoplasmic calcium levels, but rather by fast calcium reuptake kinetics. These data suggest that, under abnormal RyR conditions, triggered activity is caused by multiple SCR events that depend on the faster calcium reuptake kinetics near the epicardium. Furthermore, multiple regions of SCR may be a mechanism for multifocal arrhythmias associated with RyR dysfunction.

Electrophysiological consequence of skeletal myoblast transplantation in normal and infarcted canine myocardium

The purpose of this study is to test our hypothesis that injection of skeletal myoblasts (SkMbs) into viable tissue may alter impulse conduction but that injections into nonviable tissue (scar) will have negligible impact. Myocardial infarction (MI) is a major public health problem. SkMb transplantation after MI has been shown to have some beneficial effect on hemodynamic function. Previous studies have indicated that SkMbs do not electrically couple with viable host myocardium in vivo. We used optical mapping to measure impulse propagation and arrhythmia inducibility in the canine left ventricular wedge preparation with and without MI. MI was created by temporary ligation of a branch of the left anterior descending coronary artery (LAD) (7.0 +/- 3.8 ng/mL troponin 24 hours after MI). Labeled SkMbs (10(8) in 4 mL of serum-free basal solution) were injected from the epicardium (20-40 0.1 mL injections) into normal myocardium (n = 8) or the central zone of the MI (n = 6). During endocardial pacing in the absence of MI, transmural conduction velocity was similar with (35.75 +/- 3.4 cm/s) and without (37.42 +/- 3.6 cm/s) SkMb transplantation. However, pacing from the epicardium resulted in conduction slowing in regions that were DiI-positive and associated with the expression of skeletal myosin (fast) but not connexin-43. In all preparations with MI (n = 13), abnormal impulse propagation was seen regardless of SkMb transplantation. Arrhythmias (at least one extra beat after standard programmed stimulation) occurred most frequently in preparations with MI independent of SkMb transplantation. In preparations without MI (n = 8), SkMb transplantation did not significantly increase arrhythmia inducibility. We conclude that SkMbs transplanted into normal myocardium can cause abnormal impulse propagation. These data suggest that the location of SkMb transplantation may influence arrhythmia vulnerability associated with MI.

The Mechanism of Pause‐Induced Torsade de Pointes in Long QT Syndrome

Torsade de pointes (TdP), is often preceded by a short-long cycle length sequence. However, the causal relationship between the pause associated with a short-long cycle length sequence and TdP is not completely understood. This study tests the hypothesis that a pause enhances both dispersion of repolarization and EAD formation; however, EADs that form where APD is longest will be less likely to initiate TdP. We used optical mapping to measure transmural action potentials from the canine left ventricular wedge preparation. D-sotalol and ATX-II were used to mimic LQT2 and LQT3, respectively. The pause significantly enhanced mean APD (from 356 +/- 20 to 381 +/- 25 msec in LQT2, P < 0.05; from 609 +/- 92 to 675 +/- 98 msec in LQT3, P < 0.05) and transmural dispersion (from 35 +/- 9 to 46 +/- 11 msec in LQT2, P < 0.05; from 121 +/- 85 to 171 +/- 98 msec in LQT3, P < 0.05) compared to steady state pacing. Under LQT3 condition EADs, EAD-induced triggered activity, and TdP were more likely to occur following a pause. Interestingly, the triggered beat following a pause always broke through at the region of maximum local repolarization gradient. These data suggest that a pause accentuates transmural repolarization gradients and facilitates the formation of EADs and EAD-induced triggered activity. In contrast to our hypothesis, the findings of this study support the concept that M-cells (where APD is longest) can play an important role in both the origination of EAD-induced triggered activity and unidirectional block associated with TdP.

Biophysical basis for monophasic action potential

We welcome Dr. Franz's comments relative to our recent publication dealing with the monophasic action potential (MAP) [1]. We agree and fully appreciate the fact that the Franz contact electrode technique has been successfully used for over 20 years by Dr. Franz and other investigators. This success stems from the fact that the two electrodes of the contact catheter are relatively close to each other so that their combined field of view is fairly localized. However, we do not agree that this success can be used as an argument against our conclusion that it is not the tip electrode that records the MAP. Our study investigated the physical principles behind the MAP recording. We started with a simple definition of voltage as the difference of potential between two points or two electrodes. To determine which of these two potentials carries time-dependent information, we placed the two electrodes at sites with different cellular activity and observed which activity is reflected in the MAP signal. The observations demonstrated in a very straightforward manner that the tip or inactivating electrode is not the recording electrode. This conclusion provides support for theories introduced as early as 1934 [2]. Response to specific comments: The size of the canine left ventricular wedge preparation is typically between 2 × 1.5 × 0.9 cm and 3 × 2 × 1.5 cm. The schemes in Figs. 4 and 8 correctly depict the relative dimensions of the contact MAP catheter and the wedge preparation. Moreover, in the schematics shown in Figs. 5, 6 and 7, the catheter is larger than its actual dimension relative to the size of the wedge. The pioneering study by Dower et al. [3], cited by Dr. Franz, was aimed at development of an intracellular recording technique. The authors recognized that microelectrodes with …

Delayed Afterdepolarization‐Mediated Triggered Activity Associated with Slow Calcium Sequestration Near the Endocardium

Previously, we have shown that cells near the endocardium are more prone to elevated diastolic intracellular calcium levels than cells near the epicardium. The arrhythmogenic consequence of such regional differences in calcium handling is not clear. Using optical mapping techniques, calcium transients and action potentials were recorded simultaneously from ventricular sites across the transmural wall of the arterially perfused canine left ventricular wedge preparation during control conditions, and under conditions of increased calcium entry (I(K) blockade and beta-adrenergic stimulation). Under conditions of enhanced calcium entry, the decay of the calcium transient and diastolic calcium levels during rapid pacing were slower (38%, P < 0.01) and higher (215%, P < 0.02), respectively, near (within approximately 3 mm) the endocardium compared to the epicardium (n = 9). Immediately after termination of rapid pacing under conditions of increased calcium entry, ectopic activity and simultaneous delayed after depolarizations and spontaneous calcium release events were observed. Over all experiments, ectopic activity occurred more frequently closer to the endocardium compared to the epicardium. Under conditions of enhanced calcium entry, myocytes closer to the endocardium exhibit a higher level of diastolic calcium and greater ectopic activity compared to the epicardium. We show for the first time simultaneous delayed after depolarization and spontaneous calcium release events from myocytes in a normally coupled multicellular preparation. These data combined suggest that myocytes near the endocardium are more susceptible to calcium-mediated triggered activity.

Cisapride-induced transmural dispersion of repolarization and torsade de pointes in the canine left ventricular wedge preparation during epicardial stimulation.

Cisapride, a gastrointestinal prokinetic agent, was recently withdrawn from the market because of its propensity to induce torsade de pointes (TdP) arrhythmias. The present study examines the electrophysiological actions of cisapride in the isolated arterially perfused canine left ventricular wedge preparation. Transmembrane action potentials from epicardial and M regions and a pseudo-ECG were simultaneously recorded. Cisapride (0.1 to 5 micromol/L) was added to the coronary perfusate. Cisapride prolonged the QT interval and increased transmural dispersion of repolarization (TDR) at relatively low but not at high concentrations. TdP could be induced with programmed electrical stimulation only at a low concentration of drug (0.2 micromol/L), when TDR was maximally prolonged. Moreover, TdP could only be induced during epicardial (but not endocardial) activation of the wedge, which was found to augment TDR. At higher concentrations of cisapride, QT was further prolonged, TDR was diminished, and TdP could no longer be induced. Tpeak-Tend interval and Tpeak-Tend area provided reasonable electrocardiographic indices of TDR. Our data (1) demonstrate a biphasic concentration/response relationship for the effect of cisapride to induce long-QT syndrome and TdP, (2) show the value of the left ventricular wedge preparation in identifying drugs that pose an arrhythmic risk, (3) support the hypothesis that risk for development of TdP is related to the increase in TDR rather than to prolongation of the QT interval, and (4) indicate that epicardial activation of the left ventricle, as occurs during biventricular pacing, can facilitate the development of TdP under long-QT conditions.