In silico assessment of genetic variation in KCNA5 reveals multiple mechanisms of human atrial arrhythmogenesis

A recent experimental study investigating patients with lone atrial fibrillation identified six novel mutations in the KCNA5 gene. The mutants exhibited both gain- and loss-of-function of the atrial specific ultra-rapid delayed rectifier K+ current, IKur. The aim of this study is to elucidate and quantify the functional impact of these KCNA5 mutations on atrial electrical activity. A multi-scale model of the human atria was updated to incorporate detailed experimental data on IKur from both wild-type and mutants. The effects of the mutations on human atrial action potential and rate dependence were investigated at the cellular level. In tissue, we assessed the effects of the mutations on the vulnerability to unidirectional conduction patterns and dynamics of re-entrant excitation waves. Gain-of-function mutations shortened the action potential duration in single cells, and stabilised and accelerated re-entrant excitation in tissue. Loss-of-function mutations had heterogeneous effects on action potential duration and promoted early-after-depolarisations following beta-adrenergic stimulation. In the tissue model, loss-of-function mutations facilitated breakdown of excitation waves at more physiological excitation rates than the wild-type, and the generation of early-after-depolarisations promoted unidirectional patterns of excitation. Gain- and loss-of-function IKur mutations produced multiple mechanisms of atrial arrhythmogenesis, with significant differences between the two groups of mutations. This study provides new insights into understanding the mechanisms by which mutant IKur contributes to atrial arrhythmias. In addition, as IKur is an atrial-specific channel and a number of IKur-selective blockers have been developed as anti-AF agents, this study also helps to understand some contradictory results on both pro- and anti-arrhythmic effects of blocking IKur

In-silico investigations of the functional impact of KCNA5 mutations on atrial mechanical dynamics

A recent study has identified six novel genetic variations (D322H, E48G, A305T, D469E, Y155C, P488S) in KCNA5 (encoding Kv1.5 which carries the atrial-specific ultra-rapid delayed rectifier current, I ) in patients with early onset of lone atrial fibrillation. These mutations are distinctive, resulting in either gain-of-function (D322H, E48G, A305T) or loss-of-function (D469E, Y155C, P488S) of I channels. Though affecting potassium channels, they may modulate the cellular active force and therefore atrial mechanical functions, which remains to be elucidated. The present study aimed to assess the inotropic effects of the identified six KCNA5 mutations on the human atria. Multiscale electromechanical models of the human atria were used to investigate the impact of the six KCNA5 mutations on atrial contractile functions. It was shown that the gain-of-function mutations reduced active contractile force primarily through decreasing the calcium transient (CaT) via a reduction in the L-type calcium current (I ) as a secondary effect of modulated action potential, whereas the loss-of-function mutations mediated positive inotropic effects by increased CaT via enhancing the reverse mode of the Na /Ca exchanger. The 3D atrial electromechanical coupled model predicted different functional impacts of the KCN5A mutation variants on atrial mechanical contraction by either reducing or increasing atrial output, which is associated with the gain-of-function mutations or loss-of-function mutations in KCNA5, respectively. This study adds insights to the functional impact of KCNA5 mutations in modulating atrial contractile functions. Kur Kur CaL + 2

Disease Modeling and Disease Gene Discovery in Cardiomyopathies: A Molecular Study of Induced Pluripotent Stem Cell Generated Cardiomyocytes

The in vitro modeling of cardiac development and cardiomyopathies in human induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CMs) provides opportunities to aid the discovery of genetic, molecular, and developmental changes that are causal to, or influence, cardiomyopathies and related diseases. To better understand the functional and disease modeling potential of iPSC-differentiated CMs and to provide a proof of principle for large, epidemiological-scale disease gene discovery approaches into cardiomyopathies, well-characterized CMs, generated from validated iPSCs of 12 individuals who belong to four sibships, and one of whom reported a major adverse cardiac event (MACE), were analyzed by genome-wide mRNA sequencing. The generated CMs expressed CM-specific genes and were highly concordant in their total expressed transcriptome across the 12 samples (correlation coefficient at 95% CI =0.92 ± 0.02). The functional annotation and enrichment analysis of the 2116 genes that were significantly upregulated in CMs suggest that generated CMs have a transcriptomic and functional profile of immature atrial-like CMs; however, the CMs-upregulated transcriptome also showed high overlap and significant enrichment in primary cardiomyocyte (p-value = 4.36 × 10−9), primary heart tissue (p-value = 1.37 × 10−41) and cardiomyopathy (p-value = 1.13 × 10−21) associated gene sets. Modeling the effect of MACE in the generated CMs-upregulated transcriptome identified gene expression phenotypes consistent with the predisposition of the MACE-affected sibship to arrhythmia, prothrombotic, and atherosclerosis risk

In silico Assessment of Pharmacotherapy for Human Atrial Patho-Electrophysiology Associated With hERG-Linked Short QT Syndrome

Short QT syndrome variant 1 (SQT1) arises due to gain-of-function mutations to the human Ether-à-go-go-Related Gene (hERG), which encodes the α subunit of channels carrying rapid delayed rectifier potassium current, IKr. In addition to QT interval shortening and ventricular arrhythmias, SQT1 is associated with increased risk of atrial fibrillation (AF), which is often the only clinical presentation. However, the underlying basis of AF and its pharmacological treatment remain incompletely understood in the context of SQT1. In this study, computational modeling was used to investigate mechanisms of human atrial arrhythmogenesis consequent to a SQT1 mutation, as well as pharmacotherapeutic effects of selected class I drugs–disopyramide, quinidine, and propafenone. A Markov chain formulation describing wild type (WT) and N588K-hERG mutant IKr was incorporated into a contemporary human atrial action potential (AP) model, which was integrated into one-dimensional (1D) tissue strands, idealized 2D sheets, and a 3D heterogeneous, anatomical human atria model. Multi-channel pharmacological effects of disopyramide, quinidine, and propafenone, including binding kinetics for IKr/hERG and sodium current, INa, were considered. Heterozygous and homozygous formulations of the N588K-hERG mutation shortened the AP duration (APD) by 53 and 86 ms, respectively, which abbreviated the effective refractory period (ERP) and excitation wavelength in tissue, increasing the lifespan and dominant frequency (DF) of scroll waves in the 3D anatomical human atria. At the concentrations tested in this study, quinidine most effectively prolonged the APD and ERP in the setting of SQT1, followed by disopyramide and propafenone. In 2D simulations, disopyramide and quinidine promoted re-entry termination by increasing the re-entry wavelength, whereas propafenone induced secondary waves which destabilized the re-entrant circuit. In 3D simulations, the DF of re-entry was reduced in a dose-dependent manner for disopyramide and quinidine, and propafenone to a lesser extent. All of the anti-arrhythmic agents promoted pharmacological conversion, most frequently terminating re-entry in the order quinidine > propafenone = disopyramide. Our findings provide further insight into mechanisms of SQT1-related AF and a rational basis for the pursuit of combined IKr and INa block based pharmacological strategies in the treatment of SQT1-linked AF

In silico Assessment of Pharmacotherapy for Human Atrial Patho-Electrophysiology Associated With hERG-Linked Short QT Syndrome

Short QT syndrome variant 1 (SQT1) arises due to gain-of-function mutations to the human Ether-à-go-go-Related Gene (hERG), which encodes the α subunit of channels carrying rapid delayed rectifier potassium current, IKr. In addition to QT interval shortening and ventricular arrhythmias, SQT1 is associated with increased risk of atrial fibrillation (AF), which is often the only clinical presentation. However, the underlying basis of AF and its pharmacological treatment remain incompletely understood in the context of SQT1. In this study, computational modeling was used to investigate mechanisms of human atrial arrhythmogenesis consequent to a SQT1 mutation, as well as pharmacotherapeutic effects of selected class I drugs–disopyramide, quinidine, and propafenone. A Markov chain formulation describing wild type (WT) and N588K-hERG mutant IKr was incorporated into a contemporary human atrial action potential (AP) model, which was integrated into one-dimensional (1D) tissue strands, idealized 2D sheets, and a 3D heterogeneous, anatomical human atria model. Multi-channel pharmacological effects of disopyramide, quinidine, and propafenone, including binding kinetics for IKr/hERG and sodium current, INa, were considered. Heterozygous and homozygous formulations of the N588K-hERG mutation shortened the AP duration (APD) by 53 and 86 ms, respectively, which abbreviated the effective refractory period (ERP) and excitation wavelength in tissue, increasing the lifespan and dominant frequency (DF) of scroll waves in the 3D anatomical human atria. At the concentrations tested in this study, quinidine most effectively prolonged the APD and ERP in the setting of SQT1, followed by disopyramide and propafenone. In 2D simulations, disopyramide and quinidine promoted re-entry termination by increasing the re-entry wavelength, whereas propafenone induced secondary waves which destabilized the re-entrant circuit. In 3D simulations, the DF of re-entry was reduced in a dose-dependent manner for disopyramide and quinidine, and propafenone to a lesser extent. All of the anti-arrhythmic agents promoted pharmacological conversion, most frequently terminating re-entry in the order quinidine > propafenone = disopyramide. Our findings provide further insight into mechanisms of SQT1-related AF and a rational basis for the pursuit of combined IKr and INa block based pharmacological strategies in the treatment of SQT1-linked AF.</p

Arrhythmia mechanisms in human induced pluripotent stem cell-derived cardiomyocytes

Despite major efforts by clinicians and researchers, cardiac arrhythmia remains a leading cause of morbidity and mortality in the world. Experimental work has relied on combining high-throughput strategies with standard molecular and electrophysiological studies, which are, to a great extent, based on the use of animal models. As this poses major challenges for translation, the progress in the development of novel antiarrhythmic agents and clinical care has been mostly disappointing. Recently, the advent of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) has opened new avenues for both basic cardiac research and drug discovery: now there is an unlimited source of CMs of human origin, both from healthy individuals and patients with cardiac diseases. Understanding arrhythmic mechanisms is one the main use-cases of hiPSC-CMs, in addition to pharmacological cardiotoxicity and efficacy testing, in vitro disease modeling, developing patient-specific models and personalized drugs, and regenerative medicine. Here, we review the advances that the hiPSC-based modeling systems have brought so far regarding the understanding of both arrhythmogenic triggers and substrates, while also briefly speculating about the possibilities in the future.publishedVersionPeer reviewe

Murine Electrophysiological Models of Cardiac Arrhythmogenesis

Cardiac arrhythmias can follow disruption of the normal cellular electrophysiological processes underlying excitable activity and their tissue propagation as coherent wavefronts from the primary sinoatrial node pacemaker, through the atria, conducting structures and ventricular myocardium. These physiological events are driven by interacting, voltage-dependent, processes of activation, inactivation, and recovery in the ion channels present in cardiomyocyte membranes. Generation and conduction of these events are further modulated by intracellular Ca$^{2+}$ homeostasis, and metabolic and structural change. This review describes experimental studies on murine models for known clinical arrhythmic conditions in which these mechanisms were modified by genetic, physiological, or pharmacological manipulation. These exemplars yielded molecular, physiological, and structural phenotypes often directly translatable to their corresponding clinical conditions, which could be investigated at the molecular, cellular, tissue, organ, and whole animal levels. Arrhythmogenesis could be explored during normal pacing activity, regular stimulation, following imposed extra-stimuli, or during progressively incremented steady pacing frequencies. Arrhythmic substrate was identified with temporal and spatial functional heterogeneities predisposing to reentrant excitation phenomena. These could arise from abnormalities in cardiac pacing function, tissue electrical connectivity, and cellular excitation and recovery. Triggering events during or following recovery from action potential excitation could thereby lead to sustained arrhythmia. These surface membrane processes were modified by alterations in cellular Ca$^{2+}$ homeostasis and energetics, as well as cellular and tissue structural change. Study of murine systems thus offers major insights into both our understanding of normal cardiac activity and its propagation, and their relationship to mechanisms generating clinical arrhythmias.Medical Research Council, Wellcome Trust, McVeigh Benefaction, British Heart Foundation, Helen Kirkland Trust, Sudden Arrhythmic Death Syndrome - SADS U

Applying computational approaches to the understanding of the consequences and opportunities of ion channel properties in atrial fibrillation

Cardiac arrhythmias are disorders of the electrical system of the heart and an often clinically-challenging group of disorders. Atrial fibrillation (AF) is the most common cardiac arrhythmia in the general population; it is associated with significant morbidity and mortality. Available antiarrhythmic drugs (AADs) for the treatment of AF are older molecules with sub- optimal efficacy and safety profiles. Recent advances in basic electrophysiology and the development of sophisticated mathematical modeling approaches could help in expanding our understanding of the basic mechanisms of AF and assist in the development of novel AF- selective AADs. The purpose of this thesis was to utilize computational approaches to the understanding of the consequences and opportunities of ion channel properties, with a special emphasis on AF. The cardiac action potential is the basic functional unit of the electrical system of the heart and is the manifestation of coordinated current fluxes through specialized proteins known as ion channels. Antiarrhythmic drugs act through modulation of ion channel properties. We hypothesized that mathematical modeling could be used to study and optimize the pharmacodynamic properties of AADs for the treatment of AF. We demonstrated that the pharmacodynamic properties (binding/unbinding characteristics) of a state-dependent Na+- channel blocker modulate the drug’s anti-/proarrhythmic actions with inactivated-state blockers being optimally AF-selective. The optimized drug’s selectivity for AF was the result of its rate- selectivity (stronger effects at fast vs slow cardiomyocyte activation rates) with relatively mild atrial-selective (stronger effects in atrial vs ventricular cardiomyocytes) actions. We found that the optimally AF-selective Na+-channel blocker had sub-optimal anti-AF efficacy, but that slightly less selective drugs had favorable AF-termination rates. We then sought to explore potential current-block combinations with synergistic AF- selective properties. Using mathematical modeling and laboratory experiments, we demonstrated that the combination of optimized state-dependent Na+-channel block and K+- channel block had synergistic effects, significantly augmenting AF termination rates for any level of AF-selectivity vs pure Na+-channel block. The mechanisms of these synergistic effects were found to be mediated by the functional interaction between the action potential prolonging- v effects of K+-channel block, the Na+-channel blocker’s voltage-dependent binding/unbinding properties and the Na+ channel’s inactivation characteristics, highlighting the non-linear nature of the cardiac action potential’s dynamics. Traditional K+ currents targeted by AADs have significant ventricular proarrhythmic liabilities. Using recent experimental observations, we updated the mathematic formulation for the inactivation dynamics of the ultra-rapid delayed-rectifier K+ current (IKur), an atrial-specific current. Using this model, we showed that, contrary to what had been proposed in the published literature, IKur rate-dependent properties are mediated by its activation properties with minimal contribution from inactivation, under physiological conditions. We also demonstrated that the contribution of IKur to action potential repolarization is preserved, or even increased, in the setting of electrical remodeling-induced IKur downregulation. Finally, we described the mechanisms of the forward rate-dependent of IKur block, mediated by functional non-linear interactions with the rapid delayed inward-rectifier K+ current (IKr), the only K+ current with such properties. Until recently, fibroblasts were considered to be electrically inactive. More recently, experimental work demonstrated the presence of functional ionic current on the fibroblast and possible cardiomyocyte-fibroblast coupling. Here, we described a novel kind of heart failure- induced electrical remodeling involving the fibroblasts ion channels. This was characterized by downregulation of the fibroblast voltage-dependent K+ current (IKv,fb) and upregulation of the fibroblast inward-rectifier K+ current (IKir,fb). We then implemented our experimental findings into a mathematical model of cardiomyocyte-fibroblast coupling and found fibroblast electrical remodeling to have significant effects on the cardiomyocyte’s electrophysiological properties. In a 2-dimension model of simulated AF, downregulation of IKv,fb had an antiarrhythmic effect whereas IKir,fb upregulation was found to be proarrhythmic. The studies presented here utilized mathematical modeling to study non-linear systems in cardiac electrophysiology to tackle questions that would have been difficult to approach with traditional laboratory-based experimentation. They also showcased how theoretical results can help orient and receive confirmation with subsequent experimental work or, conversely, novel experimental findings results be implemented into a mathematical model to investigate potential consequences. Mathematical modeling is a promising tool to help in studying the complex and vi non-linear effects of pharmacological modulation of ion channel properties and assist in the development of optimized antiarrhythmics for the treatment of AF, a major unmet need in clinical medicine. As models increase in sophistication to better represent the cardiomyocyte’s electrophysiology, they will almost certainly play an ever-growing role in expanding our understanding of the mechanisms of complex arrhythmias.Les arythmies cardiaques représentent une famille de pathologies du système électrique cardiaque. La fibrillation auriculaire (FA), est l’arythmie cardiaque la plus fréquente dans la population générale et est associée à un fardeau de morbidité et mortalité cardiovasculaire important. Les médicaments antiarythmiques utilisées dans le traitement de la FA sont de vieilles molécules avec une efficacité sous-optimale et des effets secondaires importants. Les avancées récentes en électrophysiologie cardiaque fondamentale et le développement d’outils de modélisation mathématique ont le potentiel d’élargir notre compréhension des mécanismes pathophysiologiques en FA et contribuer au développement de nouveaux médicaments antiarythmiques optimisés pour le traitement de la FA. L’objectif global de cette thèse est d’utiliser les méthodes de modélisation mathématique pour étudier les conséquences et opportunités thérapeutiques de la modulation des canaux ioniques cardiaques, avec une emphase sur la FA. Le potentiel d’action cardiaque est l’unité fonctionnelle de base du système électrique cardiaque ; il est le résultat du flux coordonné de courants électriques à travers de protéines spécialisées, les canaux ioniques. Les molécules antiarythmiques agissent à travers la modulation des canaux ioniques cardiaques. Nous avons posé l’hypothèse que des modèles mathématiques pourraient être utilisés pour étudier et optimiser les propriétés pharmacodynamiques d’un médicament antiarythmique pour le traitement de la FA. Nous avons démontré que les propriétés pharmacodynamiques (propriétés de liage et déliage) d’un bloqueur des canaux Na+ état-dépendant modulent les effets anti- et pro-arythmiques de la molécule ; un bloqueur Na+ sélectif pour l’état inactivé du canal serait maximalement FA-sélectif. Cette sélectivité pour la FA est la conséquence de la sélectivité pour la fréquence (effet thérapeutique plus important à des fréquences d’activation du cardiomyocyte élevées vs basses) avec une contribution relativement faible de la sélectivité auriculaire (effet thérapeutique plus important sur les cardiomyocytes auriculaires vs ventriculaires). Par la suite, nous avons exploré des combinaisons de bloqueurs ioniques ayant des propriétés anti-FA synergiques. En utilisant des modèles mathématiques et des expériences en laboratoire, nous avons démontré que la combinaison d’un bloqueur des canaux Na+ et d’un iii bloqueur des canaux K+ a des effets synergiques, augmentant de façon importante l’efficacité anti-FA pour un même degré de sélectivité vs un bloqueur des canaux Na+ seul. Le mécanisme de synergie a été élucidé et consiste d’effets fonctionnels médiés par l’interaction du prolongement de la durée du potentiel d’action causé par le bloque des canaux K+, les propriétés voltage-dépendantes du liage et déliage du bloqueur des canaux Na+ ainsi que des propriétés d’inactivation des canaux Na+, démontrant la nature hautement non-linéaire des dynamiques du potentiel d’action cardiaque. Les courants K+ ciblés par les médicaments antiarythmiques ont des effets proarythmiques ventriculaires importants. En utilisant des données expérimentales récentes, nous avons proposé une formulation mise à jour des dynamiques d’inactivation du courant K+ IKur, un courant auriculo-sélectif. En utilisant ce modèle, nous avons démontré que, contrairement à ce qui avait été précédemment proposé, les propriétés fréquence-dépendantes du courant IKur dépendent de ses caractéristiques d’activation avec une contribution négligeable de ses propriétés d’inactivation, sous conditions physiologiques normales. Nous avons également démontré que la contribution de IKur à la repolarisation du potentiel d’action est maintenue, voir augmentée, dans le contexte de la diminution de IKur en situation de remodelage électrique induit par la FA. Finalement, nous avons décrit le mécanisme qui sous-tend les propriétés fréquence-dépendantes du bloque de IKur, l’unique courant K+ avec de telles caractéristiques. Jusqu’à très récemment, les fibroblastes cardiaques étaient considérés comme électriquement inactifs. Des travaux expérimentaux ont démontré la présence de canaux ioniques sur la surface de ces fibroblastes ainsi que la possibilité de couplage électrique entre cardiomyocytes et fibroblastes. Nous avons décrit un nouveau type de remodelage électrique en situation d’insuffisance cardiaque, le remodelage des courants ioniques des fibroblastes cardiaques. Ce remodelage est caractérisé par une diminution du courant K+ voltage-dépendant IKv,fb et une augmentation du courant K+ IKir,fb. Nous avons par la suite incorporé ces trouvailles expérimentales dans un modèle mathématique simulant l’interaction électrique entre cardiomyocytes et fibroblastes et montré que le remodelage électrique des fibroblastes peut avoir un impact important sur les propriétés électrophysiologiques des cardiomyocytes. Dans iv un modèle 2-dimensionel de FA, nous avons trouvé que la diminution de IKv,fb a un effet antiarythmique alors que l’augmentation de IKir,fb a des effets proarythmiques. Les études ici présentées utilisent les méthodes de modélisation mathématique pour l’étude de systèmes non-linéaires en électrophysiologie cardiaque et aborder des avenues de recherche difficilement accessibles aux méthodes de laboratoire traditionnelles. Elles démontrent également comment des résultats théoriques peuvent orienter et trouver confirmation dans des travaux expérimentaux subséquents ou, à l’inverse, des trouvailles expérimentales peuvent être implémentées dans les modèles mathématiques pour investiguer les conséquences de celles-ci. La modélisation mathématique est un outil prometteur pour l’étude des effets complexes et non-linéaires de la modulation pharmacologique des canaux ioniques et ainsi contribuer au développement de médicaments antiarythmiques optimisés pour le traitement de la FA, un besoin clinique majeur

Modeling Human Atrial Patho-Electrophysiology from Ion Channels to ECG - Substrates, Pharmacology, Vulnerability, and P-Waves

Half of the patients suffering from atrial fibrillation (AF) cannot be treated adequately, today. This thesis presents multi-scale computational methods to advance our understanding of patho-mechanisms, to improve the diagnosis of patients harboring an arrhythmogenic substrate, and to tailor therapy. The modeling pipeline ranges from ion channels on the subcellular level up to the ECG on the body surface. The tailored therapeutic approaches carry the potential to reduce the burden of AF

Synergistic anti-arrhythmic effects in human atria with combined use of sodium blockers and acacetin

Atrial fibrillation (AF) is the most common cardiac arrhythmia. Developing effective and safe anti-AF drugs remains an unmet challenge. Simultaneous block of both atrial-specific ultra-rapid delayed rectifier potassium (K⁺) current (I Kur ) and the Na⁺ current (I Na ) has been hypothesized to be anti-AF, without inducing significant QT prolongation and ventricular side effects. However, the antiarrhythmic advantage of simultaneously blocking these two channels vs. individual block in the setting of AF-induced electrical remodeling remains to be documented. Furthermore, many I Kur blockers such as acacetin and AVE0118, partially inhibit other K⁺ currents in the atria. Whether this multi-K⁺ -block produces greater anti-AF effects compared with selective I Kur -block has not been fully understood. The aim of this study was to use computer models to (i) assess the impact of multi-K⁺-block as exhibited by many I Kur blokers, and (ii) evaluate the antiarrhythmic effect of blocking I Kur and I Na , either alone or in combination, on atrial and ventricular electrical excitation and recovery in the setting of AF-induced electrical-remodeling. Contemporary mathematical models of human atrial and ventricular cells were modified to incorporate dose-dependent actions of acacetin (a multichannel blocker primarily inhibiting I Kur while less potently blocking Ito, I Kr , and I Ks ). Rate- and atrial-selective inhibition of I Na was also incorporated into the models. These single myocyte models were then incorporated into multicellular two-dimensional (2D) and three-dimensional (3D) anatomical models of the human atria. As expected, application of I Kur blocker produced pronounced action potential duration (APD) prolongation in atrial myocytes. Furthermore, combined multiple K⁺-channel block that mimicked the effects of acacetin exhibited synergistic APD prolongations. Synergistically anti-AF effects following inhibition of I Na and combined I Kur /K⁺-channels were also observed. The attainable maximal AF-selectivity of I Na inhibition was greatly augmented by blocking I Kur or multiple K⁺-currents in the atrial myocytes. This enhanced anti-arrhythmic effects of combined block of Na⁺- and K⁺-channels were also seen in 2D and 3D simulations; specially, there was an enhanced efficacy in terminating re-entrant excitation waves, exerting improved antiarrhythmic effects in the human atria as compared to a single-channel block. However, in the human ventricular myocytes and tissue, cellular repolarization and computed QT intervals were modestly affected in the presence of actions of acacetin and I Na blockers (either alone or in combination). In conclusion, this study demonstrates synergistic antiarrhythmic benefits of combined block of I Kur and I Na , as well as those of I Na and combined multi K⁺-current block of acacetin, without significant alterations of ventricular repolarization and QT intervals. This approach may be a valuable strategy for the treatment of AF