Takashi Washizuka

Function, subcellular localization and assembly of a novel mutation of KCNJ2 in Andersen's syndrome.

Andersens syndrome (AS) (which is characterized by periodic paralysis, cardiac arrhythmias and dysmorphic features), a hereditary disease, and missense mutations of KCNJ2 (which encodes an inward rectifying potassium channel) have been reported recently. We performed clinical and molecular analyses of a patient with AS, and found a novel mutation (G215D) of KCNJ2. Twelve-lead electrocardiography revealed a long QT interval and frequent premature ventricular contractions, and polymorphic ventricular tachycardia was induced by programmed electrical stimulation. Use of a conventional whole-cell patch-clamp system with COS7 cells demonstrated that the G215D mutant was non-functional, and that co-expression of wild type (WT)- and mutant-KCNJ2 shows a dominant negative effect on both inward and outward currents. We performed confocal laser scanning microscopy to assess the cellular trafficking of WT- and mutant-KCNJ2 subunits tagged with yellow fluorescent protein (YFP) and cyan fluorescent protein (CFP), respectively. Tagging with the YFP did not affect the channel function of WT-KCNJ2 and both proteins showed similar plasma membrane fluorescence patterns. Furthermore, the result of fluorescence resonance energy transfer (FRET) studies at the plasma membrane region suggested that both YFP-tagged WT- and CFP-tagged mutant-KCNJ2 combine to construct a hetero-multimer of the potassium channel. In conclusion, the G215D mutant of KCNJ2 is distributed normally in the plasma membrane, but exhibits a dominant-negative effect and reduces the Kir2.1 current, presumably due to hetero-multimer construction.

Myocardium Extending from the Left Atrium onto the Pulmonary Veins: A Comparison Between Subjects with and Without Atrial Fibrillation

TAGAWA, M., et al.: Myocardium Extending from the Left Atrium onto the Pulmonary Veins: A Comparison Between Subjects with and Without Atrial Fibrillation. Rapid discharges from the myocardium extending from the left atrium onto the pulmonary vein (PV) have been shown to initiate AF, and AF may be eradicated by the catheter ablation within the PV. However, if there is any difference in the distribution patterns of the myocardial sleeve onto the PV between the subjects with and without AF is to be determined. Twenty‐one autopsied hearts were examined. Eleven patients previously had AF before death and another 10 patients had normal sinus rhythm as confirmed from the medical records including ECGs before death. After exposing the heart, the distance to the peripheral end of the myocardium was measured from the PV‐atrial junction in each PV. Then, the PVs were sectioned and stained and the distal end of myocardium and the distribution pattern were studied. The anteroposterior diameter of the left atrium was also measured. In 74 of 84 PVs, the myocardium extended beyond the PV‐atrial junction. The myocardium was localized surrounding the vascular smooth muscle layer forming a myocardial sleeve. The peripheral end of the myocardial sleeve was irregular and the maximal and minimal distances were measured in each PV. The myocardium extended most distally in the superior PVs compared to the inferior ones and the maximal distance to the peripheral end was similar between the AF and non‐AF subjects (8.4 ± 2.8 vs 8.7 ± 4.4 mm for the left superior and 6.5 ± 3.5 vs 5.1 ± 3.9 mm for the right superior PV, respectively). A significant difference was found in the maximal distance in the inferior PVs: 7.3 ± 4.6 vs 3.3 ± 2.8 mm for the left (P < 0.05) and 5.7 ± 2.4 vs 1.7 ± 1.9 mm for the right inferior PV (P < 0.001) in the subjects with and without AF, respectively. The diameter of left atrium was slightly dilated in AF patients but insignificantly (4.1 ± 0.1 vs 3.6 ± 0.1 cm, P > 0.07). The myocytes on the PV were less uniform and surrounded by more fibrosis in patients with AF compared to those without AF. In conclusion, the myocardium extended beyond the atrium‐vein junction onto the PVs. The distribution patterns of the myocardium was almost similar between subjects with and without AF, but the histology suggested variable myocytes in size and fibrosis in patients with AF.

Thyroid hormone regulates mRNA expression and currents of ion channels in rat atrium.

Atrial fibrillation is one of the common arrhythmias associated with hyperthyroidism. This study examined the effects of thyroid hormone (T3) on mRNA expression and currents of major ionic channels determining the action potential duration (APD) in the rat atrium using the RNase protection assay and the whole-cell patch-clamp technique, respectively. T3 increased the Kv1.5 mRNA expression and decreased the L-type calcium channel mRNA expression, while the Kv4.2 mRNA expression did not change. APD was shorter in hyperthyroid than in euthyroid myocytes. The ultrarapid delayed rectifier potassium currents were remarkably increased in hyperthyroid than in euthyroid myocytes, whereas the transient outward potassium currents were unchanged. L-type calcium currents were decreased in hyperthyroid than in euthyroid myocytes. T3 shifted the current-voltage relationship for calcium currents negatively. In conclusion, T3 increased the outward currents and decreased the inward currents. The resultant changes of ionic currents shortened APD, providing a substrate for atrial fibrillation.

Cystic fibrosis transmembrane conductance regulator mediates sulphonylurea block of the inwardly rectifying K+ channel Kir6.1

1 Recombinant ATP‐sensitive K+ channels (KATP channels) were heterologously expressed in the NIH3T3 mouse cell line, and the electrophysiological properties were studied using patch‐clamp techniques. 2 The NIH3T3 cell lines transfected with the inwardly rectifying K+ channel Kir6.1 alone or with both Kir6.1 and cystic fibrosis transmembrane conductance regulator (CFTR) exhibited time‐independent K+ currents with weak inward rectification. In contrast, no measurable K+ conductance was observed in mock‐transfected cells or in cells transfected with CFTR alone. Regardless of co‐transfection with Kir6.1, the transfection with CFTR produced a Cl− conductance that was activated by cell dialysis with cAMP (1 mM). The conductance was reversibly suppressed by glibenclamide (30 μM). 3 Whole‐cell currents at +60 mV were blocked in a concentration‐dependent manner by Ba2+ ions with similar IC50 values: 89.3 ± 23.3 μM (Kir6.1 alone) and 67.3 ± 24.9 μM (Kir6.1‐CFTR). 4 The currents recorded from Kir6.1‐transfected cells were not affected by glibenclamide, whereas glibenclamide did inhibit the conductance expressed in cells co‐transfected with CFTR (IC50= 35.9 ± 6.6 μM). 5 In the cell‐attached mode with a 150 mM K+ pipette solution, both Kir6.1‐ and Kir6.1‐CFTR‐transfected cells displayed a class of K+ channels showing weak inward rectification and a slope conductance of 50.7 ± 1.0 and 52.4 ± 4.9 pS, respectively. 6 In the inside‐out mode, the single‐channel currents recorded from both types of cells were not inhibited by intracellular ATP (1 mM). However, glibenclamide was found to block the single‐channel activities in the co‐transfected cells.

Endothelin-1 Inhibits the Slow Component of Cardiac Delayed Rectifier K+ Currents via a Pertussis Toxin–Sensitive Mechanism

Endothelin-1 (ET-1) is a 21-amino acid peptide hormone released from myocardial and endothelial cells, whose receptors (both ETA and ETB are expressed in the myocardium. We report here that ET-1 inhibits the cardiac delayed rectifier K+ current (IK) via a pertussis toxin (PTX)-sensitive mechanism. Ventricular myocytes enzymatically isolated from guinea pig hearts were voltage-clamped by the conventional whole-cell and nystatin-perforated patch technique (intrapipette and extrapipette K+ concentrations, 150 and 5.4 mmol/L, respectively) in the presence of nifedipine (2 mumol/L). Amplitudes of tail and steady state (2-second pulse) currents were measured as IK. ET-1 suppressed the basal IK by 20.9 +/- 2.3% in a concentration-dependent manner, with an IC50 of 1.1 +/- 0.3 nmol/L (n = 19), although it did not suppress the basal IK using the nystatin method. E-4031 (5 mumol/L), a blocker of the rapid component of IK (IKr), did not prevent the inhibitory action of ET-1. ET-1 reduced by 63.4 +/- 6.5% the slow component of IK (IKs) that had been enhanced to approximately 2-fold by isoproterenol (ISO, 20 nmol/L). The action was concentration dependent, with an IC50 of 0.7 +/- 0.4 nmol/L (n = 22), and was also observed using the nystatin method. The effect of ET-1 appeared to be mediated by an ETA receptor, because it was prevented by FR139317, an ETA-selective antagonist (1 mumol/L, n = 4), and sarafotoxin s6c, an ETB-selective agonist (100 nmol/L, n = 4), could not inhibit the ISO-enhanced IK. ET-1 antagonized IKs enhanced by histamine (250 nmol/L, n = 7) and forskolin (500 nmol/L, n = 7) but did not inhibit IKs enhanced by the internal application of cAMP (100 mumol/L, n = 6). Preincubation of myocytes with PTX (5 micrograms/mL for > 60 minutes at 36 degrees C) completely abolished the inhibitory action of ET-1 on the ISO-enhanced IKs (n = 4). Thus, nanomolar ET-1 inhibits IKs via the ETA receptor/PTX-sensitive G protein/PKA pathway.

Frequency of presumed reentry with an excitable gap in sustained ventricular tachycardia unassociated with coronary artery disease

In sustained ventricular tachycardia (VT) unrelated to coronary artery disease, the incidence of reentry with an excitable gap was examined, and rapid pacing was performed to entrain VT in 48 episodes in 42 consecutive patients. Coronary artery disease was excluded by coronary arteriography. The underlying heart diseases were postoperative congenital heart diseases (n = 5), dilated (n = 7) or hypertrophic (n = 4) cardiomyopathy, arrhythmogenic right ventricular dysplasia (n = 6) and miscellaneous heart diseases (n = 5), as well as no demonstrable heart disease (n = 15) in which 8 patients had verapamil-responsive VT. Except for 1 patient with hypertrophic cardiomyopathy, 48 morphologically distinct monomorphic sustained VTs were induced. Twenty-five VTs showed right bundle branch block morphology and 23 left bundle branch block morphology, and VT was entrained in 84 and 96%, respectively. The overall incidence of the entrainment was 89.6% (43 of 48 monomorphic VTs), and the frequency of the ability to entrain VT ranged between 33.3 and 100% in the subgroups. The lowest frequency was found in hypertrophic cardiomyopathy. In conclusion, most inducible monomorphic sustained VT unassociated with coronary artery disease was presumed to be reentry with an excitable gap.

Successful Radiofrequency Catheter Ablation for Macroreentrant Ventricular Tachycardias in a Patient with Tetralogy of Fallot After Corrective Surgery

CHINUSHI, M., et al.: Successful Radiofrequency Catheter Ablation for Macroreentrant Ventricular Tachycardias in a Patient with Tetralogy of Fallot After Corrective Surgery. Radiofrequency (RF) catheter ablation was applied to two macroreentrant ventricular tachycardias (VTs) documented after corrective operation for tetralogy of Fallot. The activation wavefront of VT with a right bundle branch block pattern was found to revolve in a clockwise manner around a presumed myotomy scar in the right ventricle, and VT with a left bundle branch block pattern revolved around the same anatomical obstacle in a counterclockwise manner. In both VTs, the biggest conduction delay was confirmed at the right ventricular outflow tract. RF applications to the slow conduction area terminated each VT within a few seconds but were insufficient to cure the VTs. RF lesions were then applied to the, slow conduction area in a line to intersect the macroreentrant circuit, and both VTs became noninducible.

Intravenous Administration of Class I Antiarrhythmic Drugs Induced T Wave Alternans in a Patient with Brugada Syndrome

T Wave Alternans in Brugada Syndrome. A 71‐year‐old man who experienced aborted sudden death was referred to our hospital. Coronary artery disease and cerebral accident were ruled out by conventional tests. The 12‐lead ECG obtained at rest showed a right bundle branch block pattern and ST segment elevation in leads V1 to V3. Double ventricular extrastimuli at coupling intervals > 180 msec induced ventricular fibrillation (VF) twice during electrophysiologic study. Intravenous administration of procainamide accentuated ST segment elevation in leads V1 to V3, and visible T wave alternans was induced in leads V2 and V3 at a dose of 450 mg. Initiation of T wave alternans was not associated with changes of the cardiac cycle or development of premature beats. When procainamide infusion was discontinued, T wave alternans disappeared before the elevated ST segment returned to the control level. Pilsicainide also accentuated ST segment elevation and induced similar T wave alternans in leads V2 and V3. Class I antiarrhythmic drug‐related T wave alternans has been reported rarely in Brugada syndrome, but it may represent enhanced arrhythmogenicity of VF. We need to monitor closely and study the clinical implications of T wave alternans in Brugada syndrome.

Electrophysiological Findings in Idiopathic Recurrent Ventricular Fibrillation: Special Reference to Mode of Induction, Drug Testing, and Long‐Term Outcomes

Electrophysiological studies can be useful in the presence of idiopathic ventricular fibrillation (VF) and may be used when selecting antiarrhythmic drugs. However, the yield, the mode, and the long‐term reproducibility of the induction of VF have not yet been fully elucidated. Eight patients with idiopathic VF underwent electrophysiological study. The mean age (± SD) was 45 ± 17 years. Six were males and two were females. Diagnosis was done by exclusion. VF was induced in 6 (75%) of 8 patients using double extra stimuli at coupling intervals of 233 ± 39 and 191 ± 20 ms for the first and second extra stimuli, respectively. Of note, VF was induced by stimulation exclusively at the origin of the premature ventricular beat, which was the first complex of VF in two patients. In another patient, VF was initiated by two premature stimuli and also by a pause produced by rapid pacing. The inducibility of VF was reproduced 9–18 months after the first induction in all of the four patients studied. When the ability of antiarrhythmic drugs to suppress VF inducibility was confirmed, no recurrence was observed during the follow‐up period of 40–160 months, but a recurrence of VF was observed in one of two nonresponders. In one patient, amiodarone administration failed in preventing VF induction 9 months after initiation of therapy, and reassessment of long‐term drug‐efficacy might be indicated in some patients. In conclusion, idiopathic VF was highly inducible (75%) with double extra stimuli. In this study, it was induced from a specific site (2/8) or by a pause (1/8). Induction of VF seemed to be reproduced 9–18 months after the first study. The outcome was considered favorable when the inducibility of VF was suppressed by antiarrhythmic drugs.

Relationship between dominant prolongation of the filtered QRS duration in the right precordial leads and clinical characteristics in Brugada syndrome.

Background: Electrical abnormalities in the RVOT may be involved in Brugada syndrome.