Inducible Atrial Arrhythmias Research Articles

Abstract Mo065: An Anti-arrhythmic Action and Novel Molecular Mechanisms of Alda-1 in Holiday Heart Syndrome

Introduction: Holiday Heart Syndrome (HHS) is caused by excessive binge alcohol intake. Atrial fibrillation (AF) is the most common arrhythmia in HHS patients. With a worldwide rising trend in heavy alcohol consumption despite significant prevention efforts, effective drugs against alcohol-evoked AF are in urgent need. Methods: We assessed the effect of Alda-1, a cardiac protective agent, on mitigating binge alcohol-evoked AF and alcohol-activated stress kinase JNK2 in 50mM alcohol-exposed (24hr) H9c2 differentiated myocytes and in a well-characterized HHS mouse model (2g/kg ethanol i.p., 4 injections, every other day). Alda-1’s effect on alcohol-evoked RyR2 channel-mediated Ca 2+ waves/sparks & tachycardia/fibrillation (AT/AF) incidence were measured in intact mouse atria. RyR2 channel open probability was assessed in human RyR2 single-channel recordings. Results: We found that Alda-1 significantly reduced atrial arrhythmia inducibility in HHS mouse atria, as evidenced by a decreased incidence of alcohol-evoked AT/AF (0 vs. 0.21 incidences/pacing attempts/animal; n=6, 8). Alda-1 is an agonist of aldehyde dehydrogenase 2 (ALDH2; a key enzyme in alcohol detoxification), which inhibits cellular apoptosis. We found that ALDH2 was increased by 42%±6 in HHS mice compared to controls, and which remained elevated upon Alda-1 treatment (n=6,7,7; p<0.05). HHS hearts showed unchanged apoptotic signaling pathways, suggesting alternative mechanisms in AF genesis. Our lab recently revealed a critical role of activated cardiac stress kinase JNK2 in AF risk. Intriguingly, Alda-1 suppressed alcohol-activated JNK2 by 70%±13 (immunoprecipitated JNK2-specific activity; n=9,9, p<0.001) (but not JNK1, the other cardiac JNK isoform) in cells, while JNK2 inhibition alleviated alcohol-evoked RyR2 channel dysfunction in human RyR2 channels as well as Ca 2+ -triggered atrial arrhythmic activities and AT/AF (0 out of 8; p<0.05) in intact mouse hearts. Conclusion: Alda-1 effectively mitigates binge alcohol-evoked atrial arrhythmogenesis by suppressing JNK2 activity independently of an ALDH2-mediated anti-apoptosis effect. Our findings highlight the potential of Alda-1 as a therapeutic agent for the increasing incidence of alcohol-evoked AF.

Atrial-specific reduction of Kir2.1 channel pore diameter in addition to loss of inward-going rectification underlies inducible atrial fibrillation in a mouse model of short QT syndrome type 3

Abstract Introduction Short QT Syndrome Type 3 (SQTS3) is an extremely rare arrhythmogenic disease caused by gain-of-function mutations in the KCNJ2 gene coding the inward rectifier potassium channel Kir2.1. We investigated arrhythmogenic mechanisms associated with a de-novo mutation (E299V) in Kir2.1 in an 11-year-old boy presenting an extremely abbreviated QT interval, paroxysmal atrial fibrillation, and mild left ventricular dysfunction. Amino acid E299 in the Kir2.1 sequence is necessary for polyamine binding induced inward rectification. Purpose Test the hypothesis that Kir2.1E299V induces reduced conductance and lack of rectification that causes electrical defects in atrial cardiomyocytes, predisposing patients to atrial arrhythmias. Methods We used intravenous adeno-associated virus-mediated gene transfer to generate mice expressing wild-type (WT) and the E299V mutant protein. We used ECG, intracardiac stimulation, patch-clamp, molecular biology and computational modelling to characterize the models and study arrhythmia mechanisms in the atria and ventricles. Results We confirmed WT or mutant Kir2.1 gene expression specifically in the mouse heart. On ECG, the corrected QT (QTc) interval of Kir2.1E299V mice was significantly shorter than Kir2.1WT mice (p<0.0001). The PR interval in Kir2.1E299V was also significantly shorter than WT mice (p<0.0001). On intracardiac stimulation, the largest proportion of arrhythmic events occurred in the atria, as 7 out of 9 Kir2.1E299V mice presented >1 second atrial flutter/fibrillation, while only 2 out of 10 Kir2.1WT mice showed this type of arrhythmia (p=0.023). On patch clamping, both atrial and ventricular cardiomyocytes expressing Kir2.1E299V had extremely abbreviated action potential durations (APD90) at all frequencies studied (p<0.0001). The current/voltage relation of ventricular Kir2.1E299V cardiomyocytes revealed an absence of inward-going rectification and increased IK1 at voltages positive to −80 mV compared to Kir2.1WT cardiomyocytes (p<0.0001). In contrast, while in the atrial Kir2.1E299V cardiomyocytes the outward IK1 was increased at voltages positive to −80 mV with loss of rectification, IK1 was significantly reduced at voltages negative to −80 mV (p<0.0001), suggesting a loss of function leading to atrial arrhythmia inducibility. A higher proportion of Kir2.2 at atrial level and atomic in-silico 3D simulations suggested that the mutation impaired polyamine block of the Kir2.1E299V-Kir2.2 channel while reducing the pore diameter. Conclusions This first in-vivo mouse model of cardiac-specific SQTS3 recapitulates the electrophysiological phenotype of a patient with the Kir2.1E299V mutation. The mutation results in a Kir2.1 gain-of-function mediated by and absence of rectification. The predominant arrhythmias induced in these SQTS3 mice were supraventricular likely due to the combined lack of inward rectification and atrial-specific reduced pore diameter of the Kir2.1E299V-Kir2.2 channel. Funding Acknowledgement Type of funding sources: Private company. Main funding source(s): La Caixa FoundationLa Maratό TV3 Foundation

Pharmacological nNOS inhibition modified small-conductance Ca2+-activated K+ channel without altering Ca2+ dynamics.

Atrial fibrillation (AF) is associated with electrical remodeling processes that promote a substrate for the maintenance of AF. Although the small-conductance Ca2+-activated K+ (SK) channel is a key factor in atrial electrical remodeling, the mechanism of its activation remains unclear. Regional nitric oxide (NO) production by neuronal nitric oxide synthase (nNOS) is involved in atrial electrical remodeling. In this study, atrial tachyarrhythmia (ATA) induction and optical mapping were performed on perfused rat hearts. nNOS is pharmacologically inhibited by S-methylthiocitrulline (SMTC). The influence of the SK channel was examined using a specific channel inhibitor, apamin (APA). Parameters such as action potential duration (APD), conduction velocity, and calcium transient (CaT) were evaluated using voltage and calcium optical mapping. The dominant frequency was examined in the analysis of AF dynamics. SMTC (100 nM) increased the inducibility of ATA and apamin (100 nM) mitigated nNOS inhibition-induced arrhythmogenicity. SMTC caused abbreviations and enhanced the spatial dispersion of APD, which was reversed by apamin. By contrast, conduction velocity and other parameters associated with CaT were not affected by SMTC or apamin administration. Apamin reduced the frequency of SMTC-induced ATA. In summary, nNOS inhibition abbreviates APD by modifying the SK channels. A specific SK channel blocker, apamin, mitigated APD abbreviation without alteration of CaT, implying an underlying mechanism of posttranslational modification of SK channels.NEW & NOTEWORTHY We demonstrated that pharmacological nNOS inhibition increased the atrial arrhythmia inducibility and a specific small-conductance Ca2+-activated K+ channel blocker, apamin, reversed the enhanced atrial arrhythmia inducibility. Apamin mitigated APD abbreviation without alteration of Ca2+ transient, implying an underlying mechanism of posttranslational modification of SK channels.

Profibrillatory Structural and Functional Properties of the Atrial-Pulmonary Junction in the Absence of Remodeling.

Background: Sole pulmonary vein (PV) isolation by ablation therapy prevents atrial fibrillation (AF) in patients with short episodes of AF and without comorbidities. Since incomplete PV isolation can be curative, we tested the hypothesis that the PV in the absence of remodeling and comorbidities contains structural and functional properties that are proarrhythmic for AF initiation by reentry.Methods: We performed percutaneous transvenous in vivo endocardial electrophysiological studies and quantitative histological analysis of PV from healthy sheep.Results: The proximal PV contained more myocytes than the distal PV and a higher percentage of collagen and fat tissue relative to myocytes than the left atrium. Local fractionated electrograms occurred in both the distal and proximal PVs, but a large local activation (>0.75 mV) was more often present in the proximal PV than in the distal PV (86 vs. 50% of electrograms, respectively, p = 0.017). Atrial arrhythmias (run of premature atrial complexes) occurred more often following the premature stimulation in the proximal PV than in the distal PV (p = 0.004). The diastolic stimulation threshold was higher in the proximal PV than in the distal PV (0.7 [0.3] vs. 0.4 [0.2] mA, (median [interquartile range]), p = 0.004). The refractory period was shorter in the proximal PV than in the distal PV (170 [50] vs. 248 [52] ms, p < 0.001). A linear relation existed between the gradient in refractoriness (distal-proximal) and atrial arrhythmia inducibility in the proximal PV.Conclusion: The structural and functional properties of the native atrial-PV junction differ from those of the distal PV. Atrial arrhythmias in the absence of arrhythmia-induced remodeling are caused by reentry in the atrial-PV junction. Ablative treatment of early paroxysmal AF, rather than complete isolation of focal arrhythmia, may be limited to inhibition of reentry.

Atrial Stretch‐Dependent Tumor Necrosis Factor (TNF)‐Mediated Adverse Atrial Remodeling and Atrial Arrhythmia Inducibility in a Mouse Model of Aortic Regurgitation

Atrial fibrillation (AF) is the most common sustained supraventricular arrhythmia worldwide, with its incidence linked to cardiovascular (CV) disease and, paradoxically, endurance exercise. Most conditions linked to AF are associated with elevated atrial pressures and stretch, which are powerful stimuli for atrial hypertrophy, fibrosis, and inflammation. We previously established adverse atrial changes and arrhythmogenesis required the pro-inflammatory and mechanosensitive cytokine tumor necrosis factor (TNF) in intense swim exercised mice. Thus, we hypothesize that stretch-mediated TNF-dependent signaling may provide a unifying mechanism linking AF in exercise and disease. We developed a clinically-relevant mouse model of aortic regurgitation (AR), which is characterized by chronic diastolic volume overload and elevated left ventricular end-diastolic (LVEDPs) and atrial pressures, to study atrial stretch-dependent TNF-mediated AF pathogenesis. AR was induced by retrograde puncture of the aortic valve in 8-week-old CD1 wild-type and whole-body TNF knockout (TNF-/-) mice and cardiac and electrophysiological changes were assessed using echocardiography, invasive hemodynamics, histology, in vivo intracardiacs, and ex vivo optical mapping in isolated atria. Four weeks after regurgitation, AR resulted in volume overload-mediated progressive LV dilatation, functional impairment, hypertrophy, and elevated LVEDPs in the absence of ventricular arrhythmia inducibility in both groups. In wild-type mice, AR resulted in adverse atrial remodeling, characterized by atrial hypertrophy and fibrosis, decreased conduction velocity, reduced atrial effective refractory period and action potential duration, and increased in vivo and ex vivo AF susceptibility. By contrast, TNF-/- prevented AR-induced adverse atrial remodeling and arrhythmia inducibility, independent of ventricular changes. Our results establish that adverse atrial remodeling and AF vulnerability with AR requires TNF, providing a mechanistic link between elevated atrial pressures, adverse atrial remodeling, and AF susceptibility with CV disease and exercise.

High-Dose Dobutamine for Inducibility of Atrial Arrhythmias During Atrial Fibrillation Ablation.

This study sought to compare the effect of high-dose dobutamine (DBT) with that of high-dose isoproterenol (IPN) in eliciting triggers during atrial fibrillation (AF) ablation. High-dose IPN is commonly used to elicit triggers during AF ablation. However, it is not available worldwide and, in the United States, its cost per dose has significantly increased. DBT is a similarly nonselective β-agonist and, as such, is a potential alternative. This was a prospective, randomized 2×2 crossover study of patients undergoing AF ablation. Patients were assigned to receive IPN (20 to 30μg/min for 10min) followed by DBT (40 to 50μg/kg/min for 10min) or vice versa in a 1:1 fashion. The type, number, and location of triggers as well as heart rate, blood pressure, and side effects were noted. Fifty patients were included in the study. Both drugs caused a significant increase in heart rate, with a consistently lower peak for DBT. Blood pressure significantly increased with DBT, while there was a significant reduction with IPN, despite phenylephrine support. Atrial arrhythmias induced during DBT were comparable to that induced during IPN. In patients with IPN-inducible outflow tract premature ventricular contractions, a similar effect was noted with DBT. No major complications occurred during either drug challenge. High-dose DBT is safe and comparable to high-dose IPN in respect of eliciting AF triggers, withtheadvantage to maintain systemic pressure without the need of additional vasopressor support. This studysupports the use of high-dose DBT in electrophysiology laboratories in which IPN is not readily available andfor those patients in whom hypotension is a concern.

Acute Left Ventricular Unloading Reduces Atrial Stretch and Inhibits Atrial Arrhythmias

BackgroundLeft atrium (LA) physiology is influenced by changes in left ventricular (LV) performance and load. ObjectivesThe purpose of this study was to define the effect of acute changes in LV loading conditions on LA physiology in subacute myocardial infarction (MI). MethodsMI was percutaneously induced in 19 Yorkshire pigs. One to 2 weeks after MI, 14 pigs underwent acute LV unloading using a percutaneous LV assist device, Impella. The remaining 5 pigs underwent acute LV loading by percutaneous induction of aortic regurgitation. A pressure-volume catheter was inserted into the LA using a percutaneous transseptal approach, and LA pressure-volume loops were continuously monitored. Atrial arrhythmia inducibility was examined by burst-pacing of the right atrium. Nicotinamide adenine dinucleotide phosphate oxidase (NOX) levels and ryanodine receptor phosphorylation were examined in LA tissues to study the potential effect of stretch-dependent oxidative stress. ResultsMI resulted in reduced LV ejection fraction and increased LV end-diastolic pressure with concomitant increase in LA pressure and volumes. Acute LV unloading resulted in a reduction of LV end-diastolic pressure, which led to proportional decreases in mean LA pressure and maximum LA volume. LA pressure-volume loops exhibited a pump flow-dependent, left-downward shift. This was associated with reduced LA passive stiffness, suggesting the alleviation of the LA stretch that was present after MI. Prior to acute unloading of the LV, 71% of the pigs were arrhythmia-inducible; LV unloading reduced this to 29% (p = 0.02). Time to spontaneous termination of atrial arrhythmias was decreased from median 55 s (range 5 to 300 s) to 3 s (range 0 to 59 s). In contrast, acute LV loading with aortic regurgitation increased LA pressure without a significant effect on arrhythmogenicity. Molecular analysis of LA tissue revealed that NOX2 expression was increased after MI, whereas acute LV unloading reduced NOX2 levels and diminished ryanodine receptor phosphorylation. ConclusionsAcute LV unloading relieves LA stretch and reduces atrial arrhythmogenicity in subacute MI.

Lack of prognostic value of atrial arrhythmia inducibility and change in inducibility status after catheter ablation of atrial fibrillation

Previous studies have suggested a role of atrial arrhythmia inducibility as an endpoint of catheter ablation of atrial fibrillation (AF). The prognostic value of noninducibility after ablation and of a change in inducibility status has not been investigated in large studies. The purpose of this study was to evaluate the prognostic role of noninducibility and of a change in inducibility status after ablation of AF. We studied 305 consecutive patients with AF (66% paroxysmal) undergoing antral pulmonary vein (PV) isolation plus non-PV triggers ablation. All patients underwent a standardized induction protocol before and after ablation from the coronary sinus and right atrium: 15-beat burst pacing at 250 ms and decrementing to 180 ms (up to 20 μg/min isoproterenol). Inducibility was defined as any sustained AF or organized atrial tachycardia (AT) lasting >2 minutes. A total of 197 patients (65%) had inducible AF/AT at baseline compared to 118 (39%) after ablation. One hundred seven patients (57%) changed their inducibility status from inducible preablation to noninducible postablation. After 19 ± 7 months of follow-up, 212 patients (70%) remained free from any recurrent AF/AT. Noninducibility of AF/AT postablation (log-rank P = .236) or change in inducibility status (log-rank P = .429) was not associated with reduced risk of recurrent AF/AT. Results were consistent across the paroxysmal and nonparoxysmal subgroups. Noninducibility of atrial arrhythmia or change in inducibility status after PV isolation and non-PV trigger ablation is not associated with long-term freedom from recurrent arrhythmia and should not be used as an ablation endpoint or to support the appropriateness of additional ablation lesion sets.

Middle-term results of hybrid atrial fibrillation ablation using AtriCure system

IntroductionLong-term results of catheter ablation (CA) for persistent and long-standing persistent atrial fibrillation (AF) are disappointing. The hybrid approach is currently one of options for overcoming the limitations of CA. AimTo evaluate the safety and medium-term efficacy of the hybrid approach in patients with persistent and long-standing persistent AF. MethodsAll patients underwent epicardial thoracoscopic radiofrequency (RF) pulmonary vein (PV) isolation using the AtriCure clamp followed by left atrial (LA) linear lesions (using a linear pen), Marshall ligament disruption, and LA appendage exclusion using an Atriclip. All patients underwent an electrophysiological study (EPS) and RF CA 2–3 months after the initial surgery to eliminate recurrent conductions from/to the PVs or across the linear lesions, and to eliminate all spontaneous and inducible atrial arrhythmias. 7-Day ECG Holter monitoring was performed every 3 months during the first year and every 6 months afterwards to evaluate possible recurrent arrhythmias. ResultsSeventy patients (49 male, median 63.5 years) took part in the study. EPS was performed 87 days (median) after the thoracoscopic surgery. Seventy-six percent of patients were in normal sinus rhythm (SR) at the start of the EPS, 7% had typical atrial flutter, 11% had atrial tachycardia, and 6% were in AF. After completion of the hybrid approach, all PVs were isolated, while a complete conduction block across the linear lines was achieved in 88.6% of patients. Twelve months after the procedure, 77.1% of patients had a stable SR without any anti-arrhythmic medication or re-ablation. If we included those on anti-arrhythmic drugs and re-ablation procedures, SR was achieved in 96.5% of patients during follow-up (936±432 days). ConclusionThe sequential hybrid approach is probably the most effective and relatively safe invasive treatment for persistent and long-term persistent AF with very low medium- to long-term recurrences. Introduction of the hybrid approach to clinical practice requires extensive cooperation between cardiologic and cardiothoracic teams.

The Electrophysiologic Effects of Acute Mitral Regurgitation in a Canine Model

Atrial fibrillation (AF) occurs in 30% of patients with mitral regurgitation referred for surgical intervention. However, the underlying mechanisms in this population are poorly understood. This study examined the effects of acute left atrial volume overload on atrial electrophysiology and the inducibility of AF. Ten canines underwent insertion of an atrioventricular shunt between the left ventricle and left atrium. Shunt and aortic flows were calculated, and the shunt was titrated to a shunt fraction to 40% to 50% of cardiac output. An epicardial plaque with 250 bipolar electrodes was used to determine activation and refractory periods. Biatrial pressures and volumes, conduction times, and atrial fibrillation inducibility were recorded. Data were collected at baseline and 20 minutes after shunt opening and closure. Mean shunt flow was 1.3 ± 0.5 L/min with a shunt fraction of 43% ± 6% simulating moderate to severe mitral regurgitation. Compared with baseline, left atrial volumes and maximum pressures increased by 27% and 29%, respectively, after shunt opening. Biatrial effective refractory periods did not change significantly after shunt opening or closure. Conduction times increased by 9% with shunt opening and returned to baseline after closure. AF duration or inducibility did not change with shunt opening. This canine model of mitral regurgitation demonstrated that acute left atrial volume overload did not increase the inducibility of atrial arrhythmias in contrast with experimental and clinical findings of chronic left atrial volume overload. This suggests that the substrates for AF in patients with mitral regurgitation are a result of chronic remodeling.

Abstract 16912: Molecular Mechanisms Underlying the Beneficial Effects of Inhibition of Soluble Epoxide Hydrolase in the Prevention of Atrial Fibrillation

Introduction: Atrial fibrillation (AF) represents one of the most common arrhythmias seen clinically and is associated with a significant increase in morbidity and mortality, yet, current treatment paradigms have proven largely inadequate. Inflammation has been shown to play critical roles in the pathophysiology of AF. Soluble epoxide hydrolase (sEH) catalyzes the hydrolysis of anti-inflammatory epoxyeicosatrienoic acids to corresponding pro-inflammatory diols. We have previously shown beneficial effects of sEH inhibitors (sEHIs) in cardiac failure and in the reduction of atrial arrhythmia inducibility. The objective of this study is to elucidate the molecular mechanisms underlying the prevention of atrial arrhythmia inducibility by sEHIs. Methods: In vivo pressure-overload model was used to replicate chronic hypertension. Flow cytometry, electrophysiologic studies and optical mapping were used to determine the mechanistic roles of sEHIs on atrial fibrosis and the reversal of electrical remodeling in atrial myocytes. Results: We demonstrate positive effects of sEHI in AF prevention by reducing 1) inflammation, 2) atrial structural remodeling by suppression of atrial fibrosis and hypertrophy (Fig A-D), and 3) electrical remodeling in atrial myocytes (Fig E-G). Mechanistically, inhibition of sEH enzyme results in a more than three-fold decrease in cytokines and chemokines (n=5, p&lt;0.05), inhibition of NF-κB activation, reduction of oxidative and endoplasmic reticulum stress, and a more than five-fold decline in the phosphorylation of key signaling molecules including ERK1/2 and Smad2/3 in atrial myocytes and atrial fibroblasts (n=4, p&lt;0.05). Conclusions: This study provides insights into the underlying mechanisms leading to AF by inflammation and represents a paradigm shift from conventional antiarrhythmic drugs, which block downstream molecules to a novel upstream therapeutic target by counteracting the inflammatory processes in AF.

Atrial SERCA2a Overexpression Has No Affect on Cardiac Alternans but Promotes Arrhythmogenic SR Ca2+ Triggers.

BackgroundAtrial fibrillation (AF) is the most common arrhythmia in humans, yet; treatment has remained sub-optimal due to poor understanding of the underlying mechanisms. Cardiac alternans precede AF episodes, suggesting an important arrhythmia substrate. Recently, we demonstrated ventricular SERCA2a overexpression suppresses cardiac alternans and arrhythmias. Therefore, we hypothesized that atrial SERCA2a overexpression will decrease cardiac alternans and arrhythmias.MethodsAdult rat isolated atrial myocytes where divided into three treatment groups 1) Control, 2) SERCA2a overexpression (Ad.SERCA2a) and 3) SERCA2a inhibition (Thapsigargin, 1μm). Intracellular Ca2+ was measured using Indo-1AM and Ca2+ alternans (Ca-ALT) was induced with a standard ramp pacing protocol.ResultsAs predicted, SR Ca2+ reuptake was enhanced with SERCA2a overexpression (p< 0.05) and reduced with SERCA2a inhibition (p<0.05). Surprisingly, there was no difference in susceptibility to Ca-ALT with either SERCA2a overexpression or inhibition when compared to controls (p = 0.73). In contrast, SERCA2a overexpression resulted in increased premature SR Ca2+ (SCR) release compared to control myocytes (28% and 0%, p < 0.05) and concomitant increase in SR Ca2+ load (p<0.05). Based on these observations we tested in-vivo atrial arrhythmia inducibility in control and Ad.SERCA2a animals using an esophageal atrial burst pacing protocol. There were no inducible atrial arrhythmias in Ad.GFP (n = 4) animals though 20% of Ad.SERCA2a (n = 5) animals had inducible atrial arrhythmias (p = 0.20).ConclusionsOur findings suggest that unlike the ventricle, SERCA2a is not a key regulator of cardiac alternans in the atrium. Importantly, SERCA2a overexpression in atrial myocytes can increase SCR, which may be arrhythmogenic.

Electrophysiological properties and mechanism of aging for the susceptibility of left atrium to arrhythmogenesis in rabbits

To explore the electrophysiological properties and mechanism of aging for the susceptibility of left atrium (LA) to arrhythmogenesis in rabbits. A total of 50 male New Zealand rabbits were divided into young (6 months, n = 25) and aged (26 months, n = 25) groups. According to comparisons before and after isoproterenol (ISO) dosing, young rabbits were further divided into young control and medication groups while aged ones into aged control and medication groups. Electrophysiological study was performed on perfused isolated left atrium and conventional microelectrodes were used for recording transmenbrane action potentials (tAP), resting membrane potential (RMP), amplitude of APs (APA), maximum upstroke velocity (Vmax), APs duration at 20%, at 50% or at 90% repolarization (APD20, APD50, APD90), atrial effective refractory periods (AERP) and atrial arrhythmia inducibility before and after ISO dosing. L-type calcium current (ICa-L) was measured via whole-cell patch clamp technique. And fluorescent intensity of intracellular Ca²⁺ was detected with Flup-3/AM loading by laser scanning confocal microscope in enzymatically dissociated single LA myocytes before and after dosing. Compared with major parameters of tAP in young control group, APDs and AERP significantly increased and APA, Vmax, RMP and AERP/APD90 obviously decreased in aged control group (all P < 0.05). Otherwise, the conduction of tAP was rapid and delayed afterdepolarizations (DADs) were induced in both young and aged medication groups after ISO dosing. However, as compared with major parameters of young medication group, APDs dramatically declined while APA, Vmax, RMP and DAD amplitude significantly increased. And triggered activity was induced in aged medication group (all P < 0.05). With voltage clamp model, ICa-L significantly declined in aged control group versus young control group. The peak current density of ICa-L dramatically decreased ((6.3 ± 0.8) pA/pF in aged control group vs (9.5 ± 1.3) pA/pF in young control group, P < 0.01) under 10 mV clamp voltage. And current-voltage (I-V) curve of ICa-L was greatly upgraded in aged control group on the top of all I-V curves without changing direction. The current density of ICa-L increased much more in aged medication group than that in young medication group ((15.9 ± 3.1) vs (11.3 ± 2.6) pA/pF, P < 0.01). And I-V curve of ICa-L shifted the bottom of all I-V curves in aged medication group. Fluorescent intensities of intracellular free Ca²⁺ became completely attenuated ((437.9 ± 21.7) µmol/L in aged control group vs (778.3 ± 19.5) µmol/L in young control group, P < 0.01). Under the effect of ISO administration, fluorescent intensities of intracellular free Ca²⁺ significantly increased in both young and aged medication groups. But [Ca²⁺]i were altered more in aged medication group than that in young medication group ((1 635.6 ± 51.6) vs (1 008.7 ± 34.5) µmol/L, P < 0.05). The electrophysiological characteristics are significantly altered with electrical remodeling before and after ISO dosing in aging LA in rabbits. And it may contribute to the vulnerability and pathophysiological basis for reentry and atrial arrhythmogenesis. The electrophysiological mechanism is probably attributed to the alterations of ICa-L and [Ca²⁺]i before and after ISO perfusion in aging LA myocytes.

205 Atrial Arrhythmia Susceptibility in Arrhythmogenic Right Ventricular Cardiomyopathy

BackgroundArrhythmogenic right ventricular cardiomyopathy (ARVC) is an inherited disorder that causes sudden death and right ventricular heart failure in young athletes. Loss-of-function mutations in desmosomal proteins including plakoglobin deficiency are...

Critical Roles of SK3 Calcium-Activated Potassium Channels in the Repolarization of Atrial Myocytes

Small conductance Ca2+-activated K+ channels (SK channels) have been first identified in central nervous system, where they aid in integrating changes in intracellular Ca2+ with the intrinsic excitability of neurons and affect the synaptic transmission and plasticity. We have demonstrated that three members of SK channel family (SK1, SK2 and SK3) are present in human and mouse cardiac myocytes and contribute significantly to the repolarization process in both mouse and human atria. Moreover, the three members can heteromultimerize to form functional channels. In this study, we directly tested the contribution of SK3 channels to the overall repolarization of atrial action potentials. We used a mouse model with site-specific insertion of a tetracycline-based genetic switch in the 5' untranslated region of the KCNN3 (SK3 channel) gene so that SK3 expression could be decreased by dietary doxycycline administration without interfering with the normal profile of SK3 expression. Whole-cell patch-clamp recording showed a significant shortening of the action potential duration mainly at 90% repolarization (APD90) in atrial myocytes from the homozygous SK3T/T animals. Conversely, treatment with dietary doxycycline results in a significant prolongation of APD90 in atrial myocytes from SK3T/T animals. We further demonstrated that the shortening of action potential durations in SK3 over-expression mice predisposes the animals to inducible atrial arrhythmias. In conclusion, SK3 contributes toward atrial action potential repolarization, suggesting the pivotal role of the SK channels in atrial myocytes.

Overexpression of KCNN3 results in sudden cardiac death

A recent genome-wide association study identified a susceptibility locus for atrial fibrillation at the KCNN3 gene. Since the KCNN3 gene encodes for a small conductance calcium-activated potassium channel, we hypothesized that overexpression of the SK3 channel increases susceptibility to cardiac arrhythmias. We characterized the cardiac electrophysiological phenotype of a mouse line with overexpression of the SK3 channel. We generated homozygote (SK3(T/T)) and heterozygote (SK3(+/T)) mice with overexpression of the channel and compared them with wild-type (WT) controls. We observed a high incidence of sudden death among SK3(T/T) mice (7 of 19 SK3(T/T) mice). Ambulatory monitoring demonstrated that sudden death was due to heart block and bradyarrhythmias. SK3(T/T) mice displayed normal body weight, temperature, and cardiac function on echocardiography; however, histological analysis demonstrated that these mice have abnormal atrioventricular node morphology. Optical mapping demonstrated that SK3(T/T) mice have slower ventricular conduction compared with WT controls (SK3(T/T) vs. WT; 0.45 ± 0.04 vs. 0.60 ± 0.09 mm/ms, P = 0.001). Programmed stimulation in 1-month-old SK3(T/T) mice demonstrated inducible atrial arrhythmias (50% of SK3(T/T) vs. 0% of WT mice) and also a shorter atrioventricular nodal refractory period (SK3(T/T) vs. WT; 43 ± 6 vs. 52 ± 9 ms, P = 0.02). Three-month-old SK3(T/T) mice on the other hand displayed a trend towards a more prolonged atrioventricular nodal refractory period (SK3(T/T) vs. WT; 61 ± 1 vs. 52 ± 6 ms, P = 0.06). Overexpression of the SK3 channel causes an increased risk of sudden death associated with bradyarrhythmias and heart block, possibly due to atrioventricular nodal dysfunction.

Atrial Arrhythmia in Ageing Spontaneously Hypertensive Rats: Unraveling the Substrate in Hypertension and Ageing

BackgroundBoth ageing and hypertension are known risk factors for atrial fibrillation (AF) although the pathophysiological contribution or interaction of the individual factors remains poorly understood. Here we aim to delineate the arrhythmogenic atrial substrate in mature spontaneously hypertensive rats (SHR).MethodsSHR were studied at 12 and 15 months of age (n = 8 per group) together with equal numbers of age-matched normotensive Wistar-Kyoto control rats (WKY). Electrophysiologic study was performed on superfused isolated right and left atrial preparations using a custom built high-density multiple-electrode array to determine effective refractory periods (ERP), atrial conduction and atrial arrhythmia inducibility. Tissue specimens were harvested for structural analysis.ResultsCompared to WKY controls, the SHR demonstrated: Higher systolic blood pressure (p<0.0001), bi-atrial enlargement (p<0.05), bi-ventricular hypertrophy (p<0.05), lower atrial ERP (p = 0.008), increased atrial conduction heterogeneity (p = 0.001) and increased atrial interstitial fibrosis (p = 0.006) & CD68-positive macrophages infiltration (p<0.0001). These changes resulted in higher atrial arrhythmia inducibility (p = 0.01) and longer induced AF episodes (p = 0.02) in 15-month old SHR. Ageing contributed to incremental bi-atrial hypertrophy (p<0.01) and atrial conduction heterogeneity (p<0.01) without affecting atrial ERP, fibrosis and arrhythmia inducibility. The limited effect of ageing on the atrial substrate may be secondary to the reduction in CD68-positive macrophages.ConclusionsSignificant atrial electrical and structural remodeling is evident in the ageing spontaneously hypertensive rat atria. Concomitant hypertension appears to play a greater pathophysiological role than ageing despite their compounding effect on the atrial substrate. Inflammation is pathophysiologically linked to the pro-fibrotic changes in the hypertensive atria.

Aldosterone promotes atrial fibrillation

Hyperaldosteronism is associated with an increased prevalence of atrial fibrillation (AF). However, it is unclear whether this is the consequence of altered haemodynamics or a direct aldosterone effect. It was the aim of the study to demonstrate load-independent effects of aldosterone on atrial structure and electrophysiology. Osmotic mini-pumps delivering 1.5 µg/h aldosterone were implanted subcutaneously in rats (Aldo). Rats without aldosterone treatment served as controls. After 8 weeks, surface electrocardiogram, the inducibility of AF, and atrial pressures were recorded in vivo. In isolated working hearts, left ventricular function was measured, and conduction in the right atrium (RA) and the left atrium (LA) was mapped epicardially. The atrial effective refractory period (AERP) was determined. Atrial tissue was analysed histologically. Neither systolic nor diastolic ventricular function nor atrial pressures were altered in Aldo rats. All Aldo (11/11) showed inducible atrial arrhythmias vs. two of nine controls (P = 0.03). In Aldo, the P-wave duration and the total RA activation time were longer. Prolongation of local conduction times occurred more often in Aldo, whereas the AERP did not differ between both groups. In Aldo, atrial fibroblasts and interstitial collagen were increased, active matrix metalloproteinase 13 was reduced, and atrial myocytes were hypertrophied. The connexin 43 content was unaltered. Aldosterone causes a substrate for atrial arrhythmias characterized by atrial fibrosis, myocyte hypertrophy, and conduction disturbances. The described model imputes atrial proarrhythmia directly to aldosterone, since ventricular haemodynamics appeared unaltered in this model. This mechanism may have therapeutical impact for primary and secondary prevention of AF.

Remodeling of atrial ATP-sensitive K+ channels in a model of salt-induced elevated blood pressure

Hypertension is associated with the development of atrial fibrillation; however, the electrophysiological consequences of this condition remain poorly understood. ATP-sensitive K(+) (K(ATP)) channels, which contribute to ventricular arrhythmias, are also expressed in the atria. We hypothesized that salt-induced elevated blood pressure (BP) leads to atrial K(ATP) channel activation and increased arrhythmia inducibility. Elevated BP was induced in mice with a high-salt diet (HS) for 4 wk. High-resolution optical mapping was used to measure atrial arrhythmia inducibility, effective refractory period (ERP), and action potential duration at 90% repolarization (APD(90)). Excised patch clamping was performed to quantify K(ATP) channel properties and density. K(ATP) channel protein expression was also evaluated. Atrial arrhythmia inducibility was 22% higher in HS hearts compared with control hearts. ERP and APD(90) were significantly shorter in the right atrial appendage and left atrial appendage of HS hearts compared with control hearts. Perfusion with 1 μM glibenclamide or 300 μM tolbutamide significantly decreased arrhythmia inducibility and prolonged APD(90) in HS hearts compared with untreated HS hearts. K(ATP) channel density was 156% higher in myocytes isolated from HS animals compared with control animals. Sulfonylurea receptor 1 protein expression was increased in the left atrial appendage and right atrial appendage of HS animals (415% and 372% of NS animals, respectively). In conclusion, K(ATP) channel activation provides a mechanistic link between salt-induced elevated BP and increased atrial arrhythmia inducibility. The findings of this study have important implications for the treatment and prevention of atrial arrhythmias in the setting of hypertensive heart disease and may lead to new therapeutic approaches.

Cardiac Rac1 overexpression in mice creates a substrate for atrial arrhythmias characterized by structural remodelling

The small GTPase Rac1 seems to play a role in the pathogenesis of atrial fibrillation (AF). The aim of the present study was to characterize the effects of Rac1 overexpression on atrial electrophysiology. In mice with cardiac overexpression of constitutively active Rac1 (RacET), statin-treated RacET, and wild-type controls (age 6 months), conduction in the right and left atrium (RA and LA) was mapped epicardially. The atrial effective refractory period (AERP) was determined and inducibility of atrial arrhythmias was tested. Action potentials were recorded in isolated cells. Left ventricular function was measured by pressure-volume analysis. Five of 11 RacET hearts showed spontaneous or inducible atrial tachyarrhythmias vs. 0 of 9 controls (P < 0.05). In RacET, the P-wave duration was significantly longer (26.8 +/- 2.1 vs. 16.7 +/- 1.1 ms, P = 0.001) as was total atrial activation time (RA: 13.6 +/- 4.4 vs. 3.2 +/- 0.5 ms; LA: 7.1 +/- 1.2 vs. 2.2 +/- 0.3 ms, P < 0.01). Prolonged local conduction times occurred more often in RacET (RA: 24.4 +/- 3.8 vs. 2.7 +/- 2.1%; LA: 19.1 +/- 6.3 vs. 1.2 +/- 0.7%, P < 0.01). The AERP and action potential duration did not differ significantly between both groups. RacET demonstrated significant atrial fibrosis but only moderate systolic heart failure. RacET and statin-treated RacET were not significantly different regarding atrial electrophysiology. The substrate for atrial arrhythmias in mice with Rac1 overexpression is characterized by conduction disturbances and atrial fibrosis. Electrical remodelling (i.e. a shortening of AERP) does not play a role. Statin treatment cannot prevent the structural and electrophysiological effects of pronounced Rac1 overexpression in this model.