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Cardiovascular Research 2007 75(3):510-518; doi:10.1016/j.cardiores.2007.04.028
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Copyright © 2007, European Society of Cardiology

Mechanism of U wave and polymorphic ventricular tachycardia in a canine tissue model of Andersen–Tawil syndrome

Hiroshi Morita, Douglas P. Zipes, Shiho T. Morita and Jiashin Wu*

Krannert Institute of Cardiology, Indiana University School of Medicine, 1800 N. Capitol Ave., Indianapolis, IN 46202, USA

* Corresponding author. Tel.: +1 317 962 0075; fax: +1 317 962 0069. jiaswu{at}iupui.edu

Received 14 February 2007; revised 2 April 2007; accepted 23 April 2007


   Abstract
Top
 Abstract
1. Introduction
 2. Methods
 3. Results
 4. Discussion
 References
 
Objective Andersen–Tawil syndrome (ATS) is a channelopathy affecting inward rectifier potassium IK1 with QT prolongation, large U waves, and frequent ventricular tachycardia (VT). Although ATS is clinically defined and genetically identified, its electrophysiological mechanism is still unclear, and thus, was the subject of the current study.

Methods and results We replicated the major electrophysiological features of ATS with cesium chloride (CsCl, at IK1 blockade concentration of 5–10 mmol/l) in 23 isolated canine left ventricular tissues perfused arterially with Tyrode's solution having normal or low potassium concentrations, [K+]o. We mapped action potentials (APs) on the cut-exposed transmural surface of the wedges in control, after CsCl, and CsCl with 0.15 µmol/l isoproterenol (CsCl+ISP). CsCl delayed late phase 3 repolarization and prolonged the duration of the AP, more so during low [K+]o perfusion. Rapid pacing induced delayed afterdepolarizations (DADs) in all low [K+]o and in 71% of normal [K+]o preparations after CsCl treatment. Addition of ISP induced DADs in all preparations. DADs originated in mid-to-endocardium, and initiated VT after CsCl+ISP. Migration of DAD–VT foci resulted in multifocal VT. Alternating DADs at 2 foci resulted in bidirectional VT. There were more foci and longer durations of VT at low [K+]o than at normal [K+]o. Delayed late phase 3 repolarization of APs and DADs generated U waves. Verapamil abolished all DADs and VT.

Conclusions CsCl blockade of IK1 produced a ventricular wedge model of ATS. Suppressing IK1 generated U waves by delaying late repolarization of APs and creating DADs, and promoted polymorphic VT by triggering DADs at multiple shifting sites.

KEYWORDS Arrhythmia (mechanisms); ECG; Membrane potential; Mapping; Repolarization


   1. Introduction
Top
 Abstract
 1. Introduction
2. Methods
 3. Results
 4. Discussion
 References
 
Andersen–Tawil syndrome is a genetic disorder associated with potassium sensitive periodic paralysis, frequent ventricular arrhythmias and dysmorphic features [1,2]. Mutations of the gene KCNJ2, which encodes Kir2.1 [3–5], have been identified in Type 1 Andersen–Tawil syndrome (ATS1). Kir2.1 forms the inward rectifier potassium channel (current: IK1), which plays major roles in the final repolarization of the action potential (AP) and in maintaining the resting potential of cardiac and skeletal muscle membrane [6–8]. Cardiac involvement in ATS1 includes repolarization abnormalities consisting of mild prolongation of the QT interval (less than the other types of long QT syndrome), TU complex formed by prominent U waves in the electrocardiogram (ECG), and frequent ventricular arrhythmias [1,2,4,9]. ATS1 is also described as type 7 long QT syndrome (LQT7) due to mild QT prolongation and low occurrence of torsades de pointes [4]. It has been demonstrated that ATS1-associated ventricular arrhythmia was initiated by frequent bigeminal premature ventricular complexes (PVCs) that were increased by hypokalemia [2,4], and characterized by QRS alternans in association with bidirectional ventricular tachycardia (VT) [9]. Exercise is one of the most important triggers of syncope and polymorphic VT in ATS1 [9]. Although over 20 mutations in KCNJ2 have been reported in ATS1, the mechanism of its arrhythmogenesis and origin of its characteristic TU complex are still unknown. We investigated the mechanisms of arrhythmogenesis and formation of U wave in in vitro models of ATS1 created with drugs in isolated perfused canine left ventricular preparations.


   2. Methods
Top
 Abstract
 1. Introduction
 2. Methods
3. Results
 4. Discussion
 References
 
2.1 Isolated and arterially perfused left ventricular tissue preparations
The investigation conforms to the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH Publication No. 85-23, revised 1996). We prepared tissues with procedures similar to those used previously [10–12]. In brief, we harvested hearts from 23 anesthetized (pentobarbital sodium at ~30 mg/kg body weight) adult male mongrel dogs (25–30 kg) and immediately perfused the hearts retrogradely through the aorta with a cardioplegic solution (Tyrode's solution, see below, containing 15 mmol/l KCl, 4 °C), which washed out the blood and pentobarbital sodium and protected the hearts during subsequent tissue isolation. We then isolated a transmural wedge from the left ventricular free wall of each cardioplegic perfused heart. Each wedge (20–30 mm long by 4–7 mm wide on the epicardium, and 12–20 mm transmural) contained a section of coronary artery (diameter: ~1 mm) along its length. Two plastic cannulas, one for perfusion, the other for arterial pressure monitoring, were inserted into the two openings of the artery. Major arterial leaks in the wedges were ligated with silk sutures. Under-perfused tissue was trimmed from the wedges. The isolated tissues were mounted in a warmed chamber with the cut-exposed transmural observation surface up, perfused with 37 °C Tyrode's solution (in mmol/l, NaCl 128.0, NaHCO3 22.0, NaH2PO4 0.65, MgCl2 0.50, dextrose 11.1, CaCl2 2.0, and gassed with 95% O2–5% CO2) having either normal (KCl 4.69, n=7) or low (KCl 2.50, n=16) potassium concentration, [K+]o, at an arterial pressure of 40–50 mm Hg, and immersed in the perfusion efflux. The normal and low [K+]o groups were used to evaluate the effects of hypokalemia on arrhythmogenicity in the ATS1 models.

2.2 Drug-induced tissue model of Andersen–Tawil syndrome
We created ventricular tissue models of ATS1 by adding to the perfusate cesium chloride (CsCl, 5 and 10 mmol/l, Sigma Chemical, St. Louis, MO), which suppresses mainly IK1 in the above range of concentrations [6,7,13]. To investigate the effects of sympathetic activation, we added isoproterenol (ISP; 0.05, 0.15 µmol/l, Sigma Chemical, St. Louis, MO) to the perfusate of 11 low [K+]o and 7 normal [K+]o CsCl-treated preparations and added both norepinephrine (NE, 0.15 µmol/l, Research Biochemical Incorporated, Natick, MA) and ISP (0.15 µmol/l) to the perfusate of another 5 low [K+]o CsCl-treated preparations. To evaluate the contribution of the Ca2+ current to the VT in ATS1, we added verapamil (3.0 µmol/l, Sigma Chemical, St. Louis, MO), a calcium channel blocker, to the CsCl-treated preparations having VT.

2.3 Electrophysiological recording
Two silver electrodes were placed in the bath, 10 mm away from the epicardium (anode) and endocardium (cathode), to register the transmural ECG. The tissue preparations were stained with di-4-ANEPPS (Biotium, Inc., Hayward, CA, ~4 mmol/l in perfusate), a membrane potential sensitive fluorescent dye having no known electrophysiological effects in canines and used widely in optical mapping studies. An optical mapping system [11] with a 256-element (16x16) photodiode camera (C4675, Hamamatsu, Japan) collected the fluorescence signals from a 19.5x19.5 mm2 observation area (1.1x1.1 mm2 per element) on the tissue surface and converted it into 256 channels of electrical signals (APs). We paced (2 ms, 2x diastolic current threshold, with a bipolar electrode) the endocardium of preparations at cycle lengths (CLs) of 1000 ms during recovery. Sufficient time was allowed for the preparations to recover from the isolation process until full stabilization [with consistent arterial resistance, contractions, AP durations (APDs), velocity of conduction, and ECG for >10 min]. We evaluated the healthiness of the preparations as we have done before [10–12], noting that well-perfused preparations had a vivid reddish color, low-noise optical signals with normal APD, and strong contractions upon stimulation. Healthy wedges were immobilized with cytochalasin D (Fermentek Ltd. Jerusalem, Israel, 20–30 µmol/l), which depolymerizes cytoskeletal actin filaments and reduces the Ca2+ sensitivity of myofilaments without affecting canine ventricular APs, as we verified previously [11]. Tissue immobilization eliminated motion artifacts in optically recorded APs and facilitated the identification of early and delayed afterdepolarizations (EAD and DAD). We verified that these procedures produced stable preparations having physiologically normal transmural dispersion of APD and conduction velocity [10,12].

We recorded 256 channels of APs, transmural ECG, and arterial pressure sequentially after >10 pacing stimuli at the CLs of 4000, 2000, 1000, and 500 ms using a custom data acquisition system at 1000 samples · channel–1 s–1. The sequences of baseline data recording were performed after the tissue stabilization and verification (as the baseline control data) and repeated after 20 min of stabilization following each dose of CsCl, ISP, NE, and verapamil.

2.4 Data processing and analysis
We measured AP duration (APD) from the interval between the maximum rate of depolarization (time of depolarization) and peak of the second-order derivative of the AP (time of repolarization) as we have done previously [10–12], as well as at 50% repolarization (APD50). We statistically analyzed APDs at the recording sites along the epicardial, midmyocardial, and endocardial layers in the preparations. We measured the QT interval in the transmural ECG.

Statistical analysis was performed with Student's t test for paired data or ANOVA coupled with Scheffe's test, as appropriate. Data were expressed as mean±SD values. Significance was defined as p<0.05.


   3. Results
Top
 Abstract
 1. Introduction
 2. Methods
 3. Results
4. Discussion
 References
 
3.1 ECG and action potentials in the tissue model of Andersen–Tawil syndrome
CsCl delayed late phase 3 repolarization of the AP, and thus prolonged the QT interval and the APD (Fig. 1), with dose and CL dependency (Fig. 2), consistent with the effects of IK1 blockade. These effects were greater in the low [K+]o preparations than in the normal [K+]o preparations (Fig. 3). APDs were longer in the endocardium than in the epicardium with gradual transmural transition without M-cell-like midmyocardial preferential prolongation, both before (baseline) and after CsCl administration. Therefore, M cells were not involved in this tissue model of ATS1.


Figure 1
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  		Fig. 1 Transmural ECG and action potential (AP) distribution in normal (A) and low (B) [K+]o preparations. CsCl (10 mmol/l) delayed the late phase 3 repolarization of AP, mildly prolonged the QT interval, and produced U waves in ECG (left). The large arrows show the end of AP in control (gray arrows) and in CsCl (black arrows). Action potential duration (APD) increased gradually from the epicardium to endocardium without M-cell-like midmyocardial maximum (right, transmural distribution of APD in ms).

 

Figure 2
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  		Fig. 2 The effects of CsCl on QT interval and transmural APD in 7 normal (A) and 16 low (B) [K+]o preparations. CsCl prolonged QT interval and APD dose-dependently. These effects were enhanced by long cycle length (CL) and low [K+]o perfusion.

 

Figure 3
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  		Fig. 3 Effects of ISP and verapamil on endocardial APD in the CsCl induced model of ATS1. Drugs were added sequentially (Control, CsCl 5 mmol/l, CsCl 10 mmol/l, ISP 0.05 µmol/l, ISP 0.15 µmol/l, verapamil 3 µmol/l). CsCl prolonged APD dose-dependently. Additional ISP did not change APDs but verapamil shortened APDs. Compared to normal [K+]o, low [K+]o perfusion promoted the drug-induced prolongation of APD. There were 11 low [K+]o and 7 normal [K+]o preparations. CL: 4000 ms.

 
There was no U wave in the ECG during the baseline recording in all preparations. However, U waves were observed following the T waves, forming a TU complex, after the administration of CsCl in 75% (12/16) of the low [K+]o preparations and in 29% (2/7) of the normal [K+]o preparations (p<0.05). These U waves were associated with a significant delay in the late phase 3 repolarization of the AP (Fig. 1).

3.2 Delayed afterdepolarizations and ventricular tachycardia in the tissue model of Andersen–Tawil syndrome
The preparations were stable, free of non-paced spontaneous activity. Rapid pacing (CL≥1000 ms) induced slow phase 4 diastolic depolarization in the form of DADs following full repolarization of the preceding AP after the CsCl treatment (Fig. 4A). Prominent DADs initiated new activations. Repetitive DAD-initiated activations generated VT (Fig. 4B). Concordant relationships between the pacing CLs and first post pacing intervals of triggered beat were observed in 76% of preparations (Fig. 4E). Progressive prolongation of the activation intervals led to spontaneous termination of VTs, which was followed by additional subthreshold DADs arising from the same focal sites causing the VT (Fig. 4B). Burst pacing during VT abbreviated the CL of the VT (overdrive acceleration), but did not entrain the VT (Fig. 4C), and could lead to the termination of VT (19%) within several beats after the cessation of rapid pacing (Fig. 4D).


Figure 4
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  		Fig. 4 Delayed afterdepolarization (DAD), U wave, and VT in the CsCl-treated preparations. (A): Burst pacing induced a DAD (arrow). St = stimulation. (B): Repetitive triggered activity at the peak of DADs (indicated with triangles) produced VT. V = VT. An isolated DAD and corresponding U wave appeared after termination of VT. (C): Rapid pacing shortened the CL of ongoing VT (overdrive acceleration), but did not entrain the VT. (D): VT was terminated in several beats after cessation of burst pacing (delayed termination). (E): Shorter pacing CL induced shorter coupling interval of DADs and VT. (F): Effects of verapamil on VT induction. a) Rapid pacing (CL=500 ms) induced DADs (arrow heads) and VT. b) Verapamil (3 µmol/l) suppressed the occurrence of DADs and VT. Panel A: CsCl 10 mmol/l. Panels B–D, and F: CsCl 10 mmol/l, ISP 0.05 µmol/l.

 
In the normal [K+]o preparations, DADs were not induced during both control and low CsCl (5 mmol/l) conditions, but were induced in 71% of the preparations treated with high dose CsCl (10 mmol/l). VT did not occur in any of these preparations. Sympathetic activation with ISP (0.05, 0.15 µmol/l) induced DADs in 86% of the normal [K+]o preparations treated with CsCl. VT was induced in 14% of the preparations at 0.05 µmol/l ISP and in 71% of the preparations at 0.15 µmol/l ISP.

In the low [K+]o preparations, DADs were not present during control recordings, but appeared after low dose CsCl (5 mmol/l) treatment in 25% of these preparations. VT was not present during both control and low dose CsCl conditions in all preparations. High dose CsCl (10 mmol/l) induced DADs in 100% and VT in 38% of the low [K+]o preparations. Subsequent addition of ISP (0.05, 0.15 µmol/l) induced both DADs and VT in all low [K+]o preparations. Therefore, lower [K+]o promoted arrhythmogenicity in the tissue model of ATS1, consistent with clinical observations of increased arrhythmogenicity by hypokalemia in Andersen syndrome [2,4].

The addition of ISP increased both the duration and rate of VT, with greater effects in the low [K+]o than in normal [K+]o preparations. Multiple foci caused polymorphic VT, especially in the low [K+]o preparations with ISP. The foci of DADs and VT distributed transmurally with an endocardial preference (Fig. 5).


Figure 5
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  		Fig. 5 Characteristics of induced VT. Drugs were added sequentially (CsCl 10 mmol/l, ISP 0.05 µmol/l, and ISP 0.15 µmol/l). (A) Compared to the normal [K+]o preparations, low [K+]o preparations had longer duration of induced VTs, which were further prolonged by ISP. (B) There were multiple VT foci in the low [K+]o preparations. Adding ISP increased number of VT foci in the CsCl-treated tissue. (C) ISP shortened the cycle length (CL) of VT in the low [K+]o preparations. (D and E) Transmural distributions of VT foci in the normal (D) and low [K+]o preparations (E). Focus of VT occurred most frequently in the endocardium and enhanced by ISP. There were 11 low [K+]o and 7 normal [K+]o preparations.

 
The addition of NE did not change the occurrence of DADs or VT in a separate group of 5 low [K+]o tissues treated with CsCl. Monomorphic VTs occurred in 1 of these tissues with the same duration of 15 s both before and after NE treatment. Subsequent addition of ISP after NE produced polymorphic VT in all 5 tissues. Verapamil shortened APD, eliminated the DADs and VT completely, and prevented their induction by burst pacing, in both low and normal [K+]o preparations with ISP (Fig. 4F).

3.3 Effects of DADs on the QRS complex of ventricular tachycardia and on the U wave in the tissue model of Andersen–Tawil syndrome
Induced ventricular arrhythmias took the form of frequent premature activations (PVAs) and monomorphic and polymorphic VT. Frequent PVAs occurred as bigeminy arising from a single DAD focus (Fig. 6A). Monomorphic VT had a single DAD focus with stable QRS configuration (Fig. 6B). Migration of multiple foci (Fig. 7) and competitive activation from multiple foci (Fig. 8A) produced polymorphic (multifocal) VT in all low [K+]o preparations treated with ISP, but in only 57% of normal [K+]o preparations having ISP. Alternating discharges from 2 DAD foci with coordinated timing resulted in bidirectional VT (Fig. 8B), which occurred in 64% of low [K+]o preparations having ISP, but not in the normal [K+]o preparations (p<0.05).


Figure 6
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  		Fig. 6 Examples of premature ventricular activations (PVAs) (A) and monomorphic VT (B) in low [K+]o preparations. ECGs and APs at the focal sites (a) are shown in A-1 and B-1. St = stimulation. The numbers in the ECG and AP traces show the coupling intervals of PVAs (A-1) or cycle lengths (CLs) of VT (B-1) in ms. A-2 and B-2 show the transmural distribution maps of activation isochrone (in ms, left) and relative amplitude of DAD (in % of the phase 0 amplitude of local AP, right) of a PVA beat and a VT beat, respectively. A: PVA (bigeminy) initiated at the peak of DAD (arrows in the AP trace) at an endocardial focus (a) having the largest DAD and conducted to the rest of the tissue (A-2). B: Monomorphic VT, formed by repetitive activations arising from the DADs (arrows in B-1), conducted from at a single midmyocardial focus to the rest of tissue (B-2). [K+]o: 2.5 mmol/l, CsCl: 10 mmol/l, ISP: 0.05 µmol/l.

 

Figure 7
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  		Fig. 7 An example of polymorphic VT with multiple migrating foci. There were three QRS morphologies (QRS-1 to -3) in the transmural ECG (A). APs at the foci (a, b, and c) of QRS-1 to -3 and corresponding ECG are shown in B (with the arrows indicating the earliest activations of each beat and the numbers showing the CLs in ms). The activation isochronal maps (in ms) of the three QRS complexes and the maps of DAD amplitude (in % of the phase 0 amplitude of local AP) are shown in C. The migration of foci within the midmyocardium (a, QRS-1), sub-epicardium (b, QRS-2), and sub-endocardium (c, QRS-3) produced polymorphic VT. [K+]o: 2.5 mmol/l, CsCl: 10 mmol/l.

 

Figure 8
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  		Fig. 8 Examples of polymorphic (A) and bidirectional (B) VTs. Traces of ECG and AP (at the foci) are shown in A-1 and in B-1. The numbers in the ECG and AP traces indicate the CL (in ms). The arrows in the AP traces indicate the earliest activations of each beat. Maps of activation isochrone (in ms) and of DAD amplitude (in % of the phase 0 of local AP) are shown in A-2 and B-2. A: Competition of two DAD foci caused polymorphic VT with beat-by-beat QRS alterations (QRS-1 and -2). QRS-1 was initiated at the left focus. QRS-2 had two simultaneous foci and was taller and narrower than QRS-1. The foci were at the regions having the largest DADs. [K+]o: 2.5 mmol/l, CsCl: 10 mmol/l, ISP: 0.05 µmol/l. B: Alternating DADs from two endocardial foci (a and b) generated QRS alternans and triggered bidirectional VT. [K+]o: 2.5 mmol/l, CsCl: 10 mmol/l.

 
The manifestation of DADs caused a U wave-like slow upstroke potential in the transmural ECG. The QRS complexes during VT timed with the peaks of prominent U waves. This U wave-like potential also timed with the occurrence of isolated DADs after the termination of VT in all tissues that developed VT (Fig. 4B).


   4. Discussion
Top
 Abstract
 1. Introduction
 2. Methods
 3. Results
 4. Discussion
References
 
4.1 New observations in the tissue model of Andersen–Tawil syndrome
This study demonstrated a potential origin of the U wave and mechanisms of arrhythmogenesis in a model of ATS1 created by CsCl-perfusion in isolated wedges of the canine left ventricular free wall. This tissue model reproduced major cardiac electrophysiological characteristics of Andersen–Tawil syndrome: mild prolongation of QT interval, U wave in ECG, and frequent premature activations, ventricular arrhythmia and bidirectional tachycardia that could be exaggerated by hypokalemia, by isoproterenol, and by increased heart rate.

We found that repetitive DAD-activations caused VT. Migration of multiple DAD foci caused polymorphic VT and paired alternating DAD foci at similar cycle lengths resulted in bidirectional VT. The occurrence of DADs and VT was enhanced by burst pacing, low [K+]o in the perfusate, and ISP infusion, and was prevented by calcium channel blockade with verapamil. These results suggest the critical roles of [K+]o, heart rate, resting membrane potential, autonomic activity, and Ca2+ current in the arrhythmogenesis of ATS1.

4.2 U wave in Andersen–Tawil syndrome
The origin of the U wave in the ECG has not been fully understood, even after more than a century of investigation [14,15]. Although the U wave occurs during repolarization of the Purkinje fiber AP, the small mass makes Purkinje fibers an unlikely source. A U wave or U wave-like notch on the T wave has also been demonstrated coincident with delayed midmyocardial repolarization of M cells and with EADs in isolated ventricular tissue models of the long QT syndrome [16]. In this model of ATS1, we observed that U waves were associated with delayed phase 3 repolarization of the AP and with the occurrence of DADs. The dual sources of U waves could be responsible for the polymorphism of the U wave (e.g., the biphasic or enlarged U waves) observed clinically [9].

4.3 Mechanism of ventricular arrhythmia in Andersen–Tawil syndrome
ATS1 is a disease of the IK1 channel, which plays a major role in the late phase 3 repolarization (<–40 mV) [17] and maintains the stability of the resting potential [6–8]. We used CsCl to suppress IK1, thus, delaying late phase 3 repolarization (Fig. 1). We also evaluated the effects of hypokalamia (Figs. 1–3, 5–8GoGoGoGoGoGo), which causes additional reduction of the IK1 conductance with a square root relationship [7]. It is well recognized that delayed repolarization increases calcium influx, leading to cytosolic calcium overload and DADs [18]. The instability of the resting potential and calcium overload facilitated DAD formation (Fig. 4). Rapid pacing enhanced the calcium overload [18], thus promoting the occurrence of DADs and VT (Fig. 4A and E). Both hyperpolarization induced by hypokalemia and ISP enhance the pacemaker current, If [19]. ISP also increases the Ca2+ current, ICa [20]. Therefore, hypokalemia and ISP contributed to the formation of DADs by reducing IK1 and by enhancing If and ICa. The elimination of DADs and VT by calcium blockade with verapamil (Fig. 4F) suggested that Ca2+ overload was a major trigger of arrhythmia in this model of ATS1. Thus, cytosolic calcium overload and If could be the direct triggers, reduced IK1 provided the substrate, and hypokalemia and ISP caused exaggeration of the arrhythmias in ATS1.

We observed that the longest APDs, frequent DADs, and foci of triggered activity (VT) occurred preferentially in the endocardium of this tissue model of ATS1. This can be caused by the endocardial presence of the longest APDs. Purkinje fibers, which distribute mostly in the endocardium in the ventricular free wall [21], can also be a possible origin of DAD, especially following elevation of intracellular calcium concentration [18].

4.4 Correlations with clinical and in vivo observations
The above mechanism correlates well with clinical and in vivo observations of enhanced arrhythmogenesis by hypokalemia [2,4]. ISP infusion enhanced arrhythmogenicity while burst pacing triggered DADs, initiated (Fig. 4B) and terminated VT (Fig. 4D), consistent with clinical observations that exercise can worsen ventricular arrhythmia and induce syncope [9], as well as reduce arrhythmias [22]. A DAD-like potential has been observed in a patient with ATS1 [23], in support of the DAD mediated arrhythmogenesis in our tissue model. About one forth of Andersen–Tawil syndrome patients have polymorphic VT, and among them, over 60% had bidirectional VT [9,18,24], also found in this tissue model of ATS1 (Fig. 8B).

4.5 CsCl and arrhythmia models
Five to 10 mmol/l CsCl exerts its major effect by reducing IK1, although it can have non-specific K+ blocking activity at higher concentrations [7,13,28,29]. Experimental studies and computer simulations showed that IK1 dysfunction reduced diastolic membrane conductivity, elevated and made unstable the resting membrane potential, and delayed late phase 3 repolarization of the AP, which subsequently contributed to cytosolic Ca2+ overload, EADs and DADs, automaticity, and finally, PVC and VT [4,26,27]. In contrast, reduction of the delayed rectifier currents delays phase 2 and early phase 3 repolarization [16]. These theoretically anticipated consequences of IK1 dysfunction match well with the electrophysiological characteristics of the CsCl-treated preparation. The late phase 3 repolarization delay and frequent DADs along with minimal effects on the phase 2 and early phase 3 repolarization after CsCl treatment in the current study are consistent with the effects of 5 to 10 mmol/l CsCl mostly on IK1 blockade [4] without major effects on the delayed rectifier currents, thus, validating the CsCl-induced ATS1 model.

Previous in vivo canine studies have demonstrated that CsCl can prolong the QT and TU complex, causing polymorphic VT, torsades de pointes, and ventricular fibrillation, triggered by EADs and DADs, and augmented by bradycardia, sympathetic nerve stimulation, and calcium overload [30–38]. CsCl can also modulate autonomic nervous function and alter blood pressure in vivo [36]. We did not find EADs in the current study, possibly due to the absence of neural activity in the isolated preparations or CsCl dose.

4.6 Other models
Our CsCl-induced ATS1 model differed from another model of ATS1 created with barium chloride (BaCl2) [25]. Although both models had mild QT prolongation in the transmural ECG, the BaCl2-induced model had repolarization delay in the early phase 3 of the AP, without expressing prominent U waves, polymorphic and bidirectional VT. In contrast, the CsCl-created ATS1 model had repolarization delay in the late phase 3 of AP, U waves in the ECG, and frequent polymorphic and bidirectional VT. These electrophysiological differences could be caused by the different effects of CsCl and BaCl2 on the kinetics of IK1 and other ion currents.

4.7 Clinical Implications
The results of this study support the therapeutic strategies to manage VT in patients having ATS1 by reducing cytosolic Ca2+ overload, preventing and limiting excessive heart rate, controlling autonomic activity, elevating plasma potassium concentration, increasing the conductivity of membrane at rest, and reducing the ventricular If.

4.8 Limitation
Differences exist between IK1 reduction caused by genetic mutations or pathological down-regulation in patients and in the laboratory by CsCl blockade. In addition, the isolated tissue model of ATS1 contains only a small wedge of ventricular free wall. Canine cardiac tissue can also have different electrophysiological properties from human.


   Acknowledgements
 
This research was supported by award 0455517Z from the American Heart Association Midwest Affiliation and by the Herman C. Krannert Fund, Indianapolis.


   References
Top
 Abstract
 1. Introduction
 2. Methods
 3. Results
 4. Discussion
 References
 

  1. Andersen E.D., Krasilnikoff P.A., Overvad H. Intermittent muscular weakness, extrasystoles, and multiple developmental anomalies. A new syndrome? Acta Paediatr Scand (1971) 60:559–564.[ISI][Medline]
  2. Tawil R., Ptacek L.J., Pavlakis S.G., DeVivo D.C., Penn A.S., Ozdemir C., et al. Andersen's syndrome: potassium-sensitive periodic paralysis, ventricular ectopy, and dysmorphic features. Ann Neurol (1994) 35:326–330.[CrossRef][ISI][Medline]
  3. Plaster N.M., Tawil R., Tristani-Firouzi M., Canun S., Bendahhou S., Tsunoda A., et al. Mutations in Kir2.1 cause the developmental and episodic electrical phenotypes of Andersen's syndrome. Cell (2001) 105:511–519.[CrossRef][ISI][Medline]
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