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AimsAtrial fibrillation (AF), the most prevalent clinical arrhythmia, is associated with atrial remodelling manifesting as acute and chronic alterations in expression, function, and regulation of atrial electrophysiological and Ca2+-handling processes. These AF-induced modifications crosstalk and propagate across spatial scales creating a complex pathophysiological network, which renders AF resistant to existing pharmacotherapies that predominantly target transmembrane ion channels. Developing innovative therapeutic strategies requires a systems approach to disentangle quantitatively the pro-arrhythmic contributions of individual AF-induced alterations.Methods and resultsHere, we built a novel computational framework for simulating electrophysiology and Ca2+-handling in human atrial cardiomyocytes and tissues, and their regulation by key upstream signalling pathways [i.e. protein kinase A (PKA), and Ca2+/calmodulin-dependent protein kinase II (CaMKII)] involved in AF-pathogenesis. Populations of atrial cardiomyocyte models were constructed to determine the influence of subcellular ionic processes, signalling components, and regulatory networks on atrial arrhythmogenesis. Our results reveal a novel synergistic crosstalk between PKA and CaMKII that promotes atrial cardiomyocyte electrical instability and arrhythmogenic triggered activity. Simulations of heterogeneous tissue demonstrate that this cellular triggered activity is further amplified by CaMKII- and PKA-dependent alterations of tissue properties, further exacerbating atrial arrhythmogenesis.ConclusionsOur analysis reveals potential mechanisms by which the stress-associated adaptive changes turn into maladaptive pro-arrhythmic triggers at the cellular and tissue levels and identifies potential anti-AF targets. Collectively, our integrative approach is powerful and instrumental to assemble and reconcile existing knowledge into a systems network for identifying novel anti-AF targets and innovative approaches moving beyond the traditional ion channel-based strategy.

Aims Recent studies suggest that bioactive mediators called resolvins promote an active resolution of inflammation. Inflammatory signalling is involved in the development of the substrate for atrial fibrillation (AF). The aim of this study is to evaluate the effects of resolvin-D1 on atrial arrhythmogenic remodelling resulting from left ventricular (LV) dysfunction induced by myocardial infarction (MI) in rats. Methods and results MI was produced by left anterior descending coronary artery ligation. Intervention groups received daily intraperitoneal resolvin-D1, beginning before MI surgery (early-RvD1) or Day 7 post-MI (late-RvD1) and continued until Day 21 post-MI. AF vulnerability was evaluated by performing an electrophysiological study. Atrial conduction was analysed by using optical mapping. Fibrosis was quantified by Masson's trichrome staining and gene expression by quantitative polymerase chain reaction and RNA sequencing. Investigators were blinded to group identity. Early-RvD1 significantly reduced MI size (17 ± 6%, vs. 39 ± 6% in vehicle-MI) and preserved LV ejection fraction; these were unaffected by late-RvD1. Transoesophageal pacing induced atrial tachyarrhythmia in 2/18 (11%) sham-operated rats, vs. 18/18 (100%) MI-only rats, in 5/18 (28%, P < 0.001 vs. MI) early-RvD1 MI rats, and in 7/12 (58%, P < 0.01) late-RvD1 MI rats. Atrial conduction velocity significantly decreased post-MI, an effect suppressed by RvD1 treatment. Both early-RvD1 and late-RvD1 limited MI-induced atrial fibrosis and prevented MI-induced increases in the atrial expression of inflammation-related and fibrosis-related biomarkers and pathways. Conclusions RvD1 suppressed MI-related atrial arrhythmogenic remodelling. Early-RvD1 had MI sparing and atrial remodelling suppressant effects, whereas late-RvD1 attenuated atrial remodelling and AF promotion without ventricular protection, revealing atrial-protective actions unrelated to ventricular function changes. These results point to inflammation resolution–promoting compounds as novel cardio-protective interventions with a particular interest in attenuating AF substrate development. [ABSTRACT FROM AUTHOR]

Aims Atrial fibrillation (AF), the most prevalent clinical arrhythmia, is associated with atrial remodelling manifesting as acute and chronic alterations in expression, function, and regulation of atrial electrophysiological and Ca2+-handling processes. These AF-induced modifications crosstalk and propagate across spatial scales creating a complex pathophysiological network, which renders AF resistant to existing pharmacotherapies that predominantly target transmembrane ion channels. Developing innovative therapeutic strategies requires a systems approach to disentangle quantitatively the pro-arrhythmic contributions of individual AF-induced alterations. Methods and results Here, we built a novel computational framework for simulating electrophysiology and Ca2+-handling in human atrial cardiomyocytes and tissues, and their regulation by key upstream signalling pathways [i.e. protein kinase A (PKA), and Ca2+/calmodulin-dependent protein kinase II (CaMKII)] involved in AF-pathogenesis. Populations of atrial cardiomyocyte models were constructed to determine the influence of subcellular ionic processes, signalling components, and regulatory networks on atrial arrhythmogenesis. Our results reveal a novel synergistic crosstalk between PKA and CaMKII that promotes atrial cardiomyocyte electrical instability and arrhythmogenic triggered activity. Simulations of heterogeneous tissue demonstrate that this cellular triggered activity is further amplified by CaMKII- and PKA-dependent alterations of tissue properties, further exacerbating atrial arrhythmogenesis. Conclusions Our analysis reveals potential mechanisms by which the stress-associated adaptive changes turn into maladaptive pro-arrhythmic triggers at the cellular and tissue levels and identifies potential anti-AF targets. Collectively, our integrative approach is powerful and instrumental to assemble and reconcile existing knowledge into a systems network for identifying novel anti-AF targets and innovative approaches moving beyond the traditional ion channel-based strategy. [ABSTRACT FROM AUTHOR]

Cardiac electrophysiology is regulated by continuous trafficking and internalization of ion channels occurring over minutes to hours. Kv11.1 (also known as hERG) underlies the rapidly activating delayed‐rectifier K+ current (IKr), which plays a major role in cardiac ventricular repolarization. Experimental characterization of the distinct temporal effects of genetic and acquired modulators on channel trafficking and gating is challenging. Computer models are instrumental in elucidating these effects, but no currently available model incorporates ion‐channel trafficking. Here, we present a novel computational model that reproduces the experimentally observed production, forward trafficking, internalization, recycling and degradation of Kv11.1 channels, as well as their modulation by temperature, pentamidine, dofetilide and extracellular K+. The acute effects of these modulators on channel gating were also incorporated and integrated with the trafficking model in the O'Hara–Rudy human ventricular cardiomyocyte model. Supraphysiological dofetilide concentrations substantially increased Kv11.1 membrane levels while also producing a significant channel block. However, clinically relevant concentrations did not affect trafficking. Similarly, severe hypokalaemia reduced Kv11.1 membrane levels based on long‐term culture data, but had limited effect based on short‐term data. By contrast, clinically relevant elevations in temperature acutely increased IKr due to faster kinetics, while after 24 h, IKr was decreased due to reduced Kv11.1 membrane levels. The opposite was true for lower temperatures. Taken together, our model reveals a complex temporal regulation of cardiac electrophysiology by temperature, hypokalaemia, and dofetilide through competing effects on channel gating and trafficking, and provides a framework for future studies assessing the role of impaired trafficking in cardiac arrhythmias. Key points: Kv11.1 channels underlying the rapidly activating delayed‐rectifier K+ current are important for ventricular repolarization and are continuously shuttled from the cytoplasm to the plasma membrane and back over minutes to hours.Kv11.1 gating and trafficking are modulated by temperature, drugs and extracellular K+ concentration but experimental characterization of their combined effects is challenging. Computer models may facilitate these analyses, but no currently available model incorporates ion‐channel trafficking.We introduce a new two‐state ion‐channel trafficking model able to reproduce a wide range of experimental data, along with the effects of modulators of Kv11.1 channel functioning and trafficking.The model reveals complex dynamic regulation of ventricular repolarization by temperature, extracellular K+ concentration and dofetilide through opposing acute (millisecond) effects on Kv11.1 gating and long‐term (hours) modulation of Kv11.1 trafficking.This in silico trafficking framework provides a tool to investigate the roles of acute and long‐term processes on arrhythmia promotion and maintenance. [ABSTRACT FROM AUTHOR]

Cardiac arrhythmias are a major cause of death and disability. A large number of experimental cell and animal models have been developed to study arrhythmogenic diseases. These models have provided important insights into the underlying arrhythmia mechanisms and translational options for their therapeutic management. This position paper from the ESC Working Group on Cardiac Cellular Electrophysiology provides an overview of (i) currently available in vitro, ex vivo, and in vivo electrophysiological research methodologies, (ii) the most commonly used experimental (cellular and animal) models for cardiac arrhythmias including relevant species differences, (iii) the use of human cardiac tissue, induced pluripotent stem cell (hiPSC)-derived and in silico models to study cardiac arrhythmias, and (iv) the availability, relevance, limitations, and opportunities of these cellular and animal models to recapitulate specific acquired and inherited arrhythmogenic diseases, including atrial fibrillation, heart failure, cardiomyopathy, myocarditis, sinus node, and conduction disorders and channelopathies. By promoting a better understanding of these models and their limitations, this position paper aims to improve the quality of basic research in cardiac electrophysiology, with the ultimate goal to facilitate the clinical translation and application of basic electrophysiological research findings on arrhythmia mechanisms and therapies. [ABSTRACT FROM AUTHOR]

Aims: Atrial fibrillation (AF) prevalence increases with advanced stages of left ventricular (LV) dysfunction. Remote proarrhythmic effects of ventricular dysfunction on atrial electrophysiology remain incompletely understood. We hypothesized that repolarizing K2P3.1 K+channels, previously implicated in AF pathophysiology, may contribute to shaping the atrial action potential (AP), forming a specific electrical substrate with LV dysfunction that might represent a target for personalized antiarrhythmic therapy. Methods and results: A total of 175 patients exhibiting different stages of LV dysfunction were included. Ion channel expression was quantified by real-time polymerase chain reaction and Western blot. Membrane currents and APs were recorded from atrial cardiomyocytes using the patch-clamp technique. Severely reduced LV function was associated with decreased atrial K2P3.1 expression in sinus rhythm patients. In contrast, chronic (c)AF resulted in increased K2P3.1 levels, but paroxysmal (p)AF was not linked to significant K2P3.1 remodelling. LV dysfunction-related suppression of K2P3.1 currents prolonged atrial AP duration (APD) compared with patients with preserved LV function. In individuals with concomitant LV dysfunction and cAF, APD was determined by LV dysfunction-associated prolongation and by cAF-dependent shortening, respectively, consistent with changes in K2P3.1 abundance. K2P3.1 inhibition attenuated APD shortening in cAF patients irrespective of LV function, whereas in pAF subjects with severely reduced LV function, K2P3.1 blockade resulted in disproportionately high APD prolongation. Conclusion: LV dysfunction is associated with reduction of atrial K2P3.1 channel expression, while cAF leads to increased K2P3.1 abundance. Differential remodelling of K2P3.1 and APD provides a basis for patient-tailored antiarrhythmic strategies. [ABSTRACT FROM AUTHOR]

Atrial fibrillation (AF) is an extremely prevalent arrhythmia that presents a wide range of therapeutic challenges. AF usually begins in a self-terminating paroxysmal form (pAF). With time, the AF pattern often evolves to become persistent (nonterminating within 7 days). Important differences exist between pAF and persistent AF in terms of clinical features, in particular the responsiveness to antiarrhythmic drugs and ablation therapy. AF mechanisms have been extensively reviewed, but few or no Reviews focus specifically on the pathophysiology of pAF. Accordingly, in this Review, we examine the available data on the electrophysiological basis for pAF occurrence and maintenance, as well as the molecular mechanisms forming the underlying substrate. We first consider the mechanistic insights that have been obtained from clinical studies in the electrophysiology laboratory, noninvasive observations, and genetic studies. We then discuss the information about underlying molecular mechanisms that has been obtained from experimental studies on animal models and patient samples. Finally, we discuss the data available from animal models with spontaneous AF presentation, their relationship to clinical findings, and their relevance to understanding the mechanisms underlying pAF. Our analysis then turns to potential factors governing cases of progression from pAF to persistent AF and the clinical implications of the basic mechanisms we review. We conclude by identifying and discussing questions that we consider particularly important to address through future research in this area. [ABSTRACT FROM AUTHOR]

The complexity of the heart makes an intuitive understanding of the relative contribution of ion channels, transporters and signalling pathways to cardiac electrophysiology challenging. Computational modelling of cardiac cellular electrophysiology has proven useful to integrate experimental findings, extrapolate results obtained in expression systems or animal models to other systems, test quantitatively ideas based on experimental data and provide novel hypotheses that are experimentally testable. While the bulk of computational modelling has traditionally been directed towards ventricular bioelectricity, increasing recognition of the clinical importance of atrial arrhythmias, particularly atrial fibrillation, has led to widespread efforts to apply computational approaches to understanding atrial electrical function. The increasing availability of detailed, atrial-specific experimental data has stimulated the development of novel computational models of atrial-cellular electrophysiology and Ca2+ handling. To date, more than 300 studies have employed mathematical simulations to enhance our understanding of atrial electrophysiology, arrhythmogenesis and therapeutic responses. Future modelling studies are likely to move beyond current whole-cell models by incorporating new data on subcellular architecture, macromolecular protein complexes, and localized ion-channel regulation by signalling pathways. At the same time, more integrative multicellular models that take into account regional electrophysiological and Ca2+ handling properties, mechano-electrical feedback and/or autonomic regulation will be needed to investigate the mechanisms governing atrial arrhythmias. A combined experimental and computational approach is expected to provide the more comprehensive understanding of atrial arrhythmogenesis that is required to develop improved diagnostic and therapeutic options. Here, we review this rapidly expanding area, with a particular focus on Ca2+ handling, and provide ideas about potential future directions. [ABSTRACT FROM AUTHOR]

Aims Previous studies suggested that T-type Ca2+-current (ICaT)-blockers improve cardiac remodelling, but all available ICaT-blockers have non-specific actions on other currents and/or functions. To clarify the role of ICaT in cardiac remodelling, we studied mice with either of the principal cardiac ICaT-subunits (Cav3.1 or Cav3.2) knocked out. Methods and results Adult male Cav3.1- or Cav3.2-knockout (Cav3.1−/−, Cav3.2−/−) mice and respective wild-type (WT) littermate controls were subjected to left anterior descending coronary artery ligation to create myocardial infarction (MI). Echocardiography and programmed electrical stimulation were performed at baseline and 4 weeks post-MI. At baseline, Cav3.1−/− mice had slowed heart rates and longer PR intervals vs. WT, but no other electrophysiological and no haemodynamic differences. Cav3.2−/− showed no differences vs. WT. Contractile indices (left ventricular fractional shortening and ejection fraction) decreased more post-MI in Cav3.1−/− mice than in Cav3.1+/+ (e.g. by 34 and 29% for WT; 50 and 45% for Cav3.1−/−, respectively; P < 0.05 for each). Cav3.1−/− mice had increased ventricular tachycardia (VT) inducibility post-MI (9 of 11, 82%) vs. WT (3 of 10, 30%; P < 0.05). Cav3.2−/− mice were not different in cardiac function or VT inducibility vs. WT. Quantitative polymerase chain reaction showed that Cav3.1 is the major ICaT-subunit and that no compensatory Cav3.2 up-regulation occurs in Cav3.1−/− mice. Cav3.1−/− and Cav3.2−/− mice had no mRNA expression for the knocked-out gene, at baseline or post-MI. Conclusion Our findings suggest that, contrary to suggestions from previous studies with (imperfectly selective) pharmacological agents having T-type Ca2+-channel-blocking actions, elimination of Cav3.1 expression leads to impaired cardiac function and enhanced arrhythmia vulnerability post-MI, whereas Cav3.2 elimination has no effect. [ABSTRACT FROM PUBLISHER]

The article presents a study which explores the cell-signaling that causes action potential duration (APD) and L-type calcium current dowregulation in an in vitro model of atrial tachycardia (AT) remodeling. It notes that AT downregulates the L-type calcium current and causes atrial fibrillation (AF), which promotes electric remodeling. It explains that AF is the most common clinical tachyarrythmia, which causes electrophysiological changes as a result from the rapid atrial activation rates.

CHRIST, T.,et al.: Recording Atrial Monophasic Action Potentials Using Standard Pacemaker Leads: An Alternative Way to Study Electrophysiological Properties of the Human Atrium In Vivo?AF leads to electrophysiological changes, but it is not known if similar alterations also appear before the onset of the first episode of AF because invasive electrophysiological studies are not justified in otherwise symptom-free patients. To address this question requires a safe method of obtaining atrial electrophysiological parameters at no extra risk or discomfort for the patient. The aim of this study was to test if recording of monophasic action potentials (MAPs) is feasible during pacemaker implantation. The study included 22 patients undergoing pacemaker implantation for symptomatic bradycardia without any history of AF. Using a custommade amplifier and a minor modification of the routine procedure for intraoperatively measured P waves, atrial electrograms could be recorded using a standard active pacemaker lead. MAP-like electrograms were obtained in 15 patients. MAP amplitude was 2.6± 0.3 mV, mean action potential duration was 316± 12 ms at a spontaneous heart rate of 67.2± 3.2 beats/min. MAP duration was decreased when atria were stimulated at shorter cycle lengths (249± 12 ms at 150 beats/min, P<0.05 vs sinus rhythm). In about two thirds of patients undergoing pacemaker implantation, recording of MAP-like electrograms was feasible with only minor modification of the atrial electrogram recording technique. The method should allow screening patients for electrophysiological alterations even before the onset of AF.(PACE 2004; 27:1632–1637) [ABSTRACT FROM AUTHOR]

The renin-angiotensin-aldosterone system (RAAS) plays a major role in cardiovascular health and disease. Short-term RAAS activation controls water and salt retention and causes vasoconstriction, which are beneficial for maintaining cardiac output in low blood pressure and early stage heart failure. However, prolonged RAAS activation is detrimental, leading to structural remodeling and cardiac dysfunction. Natriuretic peptides (NPs) are activated to counterbalance the effect of RAAS and sympathetic nervous system by facilitating water and salt excretion and causing vasodilation. Neprilysin is a major NP-degrading enzyme that degrades multiple vaso-modulatory substances. Although the inhibition of neprilysin alone is not sufficient to counterbalance RAAS activation in cardiovascular diseases (e.g., hypertension and heart failure), a combination of angiotensin receptor blocker and neprilysin inhibitor (ARNI) was highly effective in several clinical trials and may modulate the risk of atrial and ventricular arrhythmias. This review summarizes the possible link between ARNI and cardiac arrhythmias and discusses potential underlying mechanisms, providing novel insights about the therapeutic role and safety profile of ARNI in the cardiovascular system. [ABSTRACT FROM AUTHOR]

Acute excessive ethyl alcohol (ethanol) consumption alters cardiac electrophysiology and can evoke cardiac arrhythmias, e.g., in 'holiday heart syndrome'. Ethanol acutely modulates numerous targets in cardiomyocytes, including ion channels, Ca2+-handling proteins and gap junctions. However, the mechanisms underlying ethanol-induced arrhythmogenesis remain incompletely understood and difficult to study experimentally due to the multiple electrophysiological targets involved and their potential interactions with preexisting electrophysiological or structural substrates. Here, we employed cellular- and tissue-level in-silico analyses to characterize the acute effects of ethanol on cardiac electrophysiology and arrhythmogenesis. Acute electrophysiological effects of ethanol were incorporated into human atrial and ventricular cardiomyocyte computer models: reduced I Na , I Ca,L , I to , I Kr and I Kur , dual effects on I K1 and I K,ACh (inhibition at low and augmentation at high concentrations), and increased I NCX and SR Ca2+ leak. Multiscale simulations in the absence or presence of preexistent atrial fibrillation or heart-failure-related remodeling demonstrated that low ethanol concentrations prolonged atrial action-potential duration (APD) without effects on ventricular APD. Conversely, high ethanol concentrations abbreviated atrial APD and prolonged ventricular APD. High ethanol concentrations promoted reentry in tissue simulations, but the extent of reentry promotion was dependent on the presence of altered intercellular coupling, and the degree, type, and pattern of fibrosis. Taken together, these data provide novel mechanistic insight into the potential proarrhythmic interactions between a preexisting substrate and acute changes in cardiac electrophysiology. In particular, acute ethanol exposure has concentration-dependent electrophysiological effects that differ between atria and ventricles, and between healthy and diseased hearts. Low concentrations of ethanol can have anti-fibrillatory effects in atria, whereas high concentrations promote the inducibility and maintenance of reentrant atrial and ventricular arrhythmias, supporting a role for limiting alcohol intake as part of cardiac arrhythmia management. Unlabelled Image • Ethanol has region-specific concentration-dependent electrophysiological effects. • Low ethanol concentrations have an I K1 -mediated protective effect in the atrium. • High ethanol concentrations promote reentrant atrial and ventricular arrhythmias. • Ethanol-induced gap-junction remodeling is key for increased reentry vulnerability. • The degree, type and pattern of fibrosis define the exact proarrhythmic risk of ethanol. [ABSTRACT FROM AUTHOR]

The identification of disturbances in the cellular structure, electrophysiology and calcium handling of atrial cardiomyocytes is crucial to the understanding of common pathologies such as atrial fibrillation. Human right atrial specimens can be obtained during routine cardiac surgery and may be used for isolation of atrial myocytes. These samples provide the unique opportunity to directly investigate the effects of human disease on atrial myocytes. However, atrial myocytes vary greatly between patients, there is little if any access to truly healthy controls and the challenges associated with assessing the in vivo effects of drugs or devices in man are considerable. These issues highlight the need for animal models. Large mammalian models are particularly suitable for this purpose as their cardiac structure and electrophysiology are comparable with humans. Here, we review techniques for obtaining atrial cardiomyocytes. We start with background information on solution composition. Agents shown to increase viable cell yield will then be explored followed by a discussion of the use of tissue-dissociating enzymes. Protocols are detailed for the perfusion method of cell isolation in large mammals and the chunk digest methods of cell isolation in humans. [ABSTRACT FROM AUTHOR]

Calcium (Ca2+) plays a central role in cardiomyocyte excitation-contraction coupling. To ensure an optimal electrical impulse propagation and cardiac contraction, Ca2+ levels are regulated by a variety of Ca2+-handling proteins. In turn, Ca2+ modulates numerous electrophysiological processes. Accordingly, Ca2+-handling abnormalities can promote cardiac arrhythmias via various mechanisms, including the promotion of afterdepolarizations, ion-channel modulation and structural remodeling. In the last 30 years, significant improvements have been made in the computational modeling of cardiomyocyte Ca2+ handling under physiological and pathological conditions. However, numerous questions involving the Ca2+-dependent regulation of different macromolecular complexes, cross-talk between Ca2+-dependent regulatory pathways operating over a wide range of time scales, and bidirectional interactions between electrophysiology and mechanics remain to be addressed by in vitro and in silico studies. A better understanding of disease-specific Ca2+-dependent proarrhythmic mechanisms may facilitate the development of improved therapeutic strategies. In this review, we describe the fundamental mechanisms of cardiomyocyte Ca2+ handling in health and disease, and provide an overview of currently available computational models for cardiomyocyte Ca2+ handling. Finally, we discuss important uncertainties and open questions about cardiomyocyte Ca2+ handling and highlight how synergy between in vitro and in silico studies may help to answer several of these issues. • Ca2+-handling abnormalities play important roles in cardiac arrhythmogenesis. • Computer models enable studies of cardiac electrophysiology in health and disease. • Cardiomyocyte models can investigate complex Ca2+-dependent signaling pathways. • Cardiomyocyte models provide insight in bidirectional electromechanical coupling. • Interactions between in silico and in vitro studies may improve data accuracy. [ABSTRACT FROM AUTHOR]

Serine/threonine protein phosphatases control dephosphorylation of numerous cardiac proteins, including a variety of ion channels and calcium-handling proteins, thereby providing precise post-translational regulation of cardiac electrophysiology and function. Accordingly, dysfunction of this regulation can contribute to the initiation, maintenance and progression of cardiac arrhythmias. Atrial fibrillation (AF) is the most common heart rhythm disorder and is characterized by electrical, autonomic, calcium-handling, contractile, and structural remodeling, which include, among other things, changes in the phosphorylation status of a wide range of proteins. Here, we review AF-associated alterations in the phosphorylation of atrial ion channels, calcium-handling and contractile proteins, and their role in AF-pathophysiology. We highlight the mechanisms controlling the phosphorylation of these proteins and focus on the role of altered dephosphorylation via local type-1, type-2A and type-2B phosphatases (PP1, PP2A, and PP2B, also known as calcineurin, respectively). Finally, we discuss the challenges for phosphatase research, potential therapeutic significance of altered phosphatase-mediated protein dephosphorylation in AF, as well as future directions. [ABSTRACT FROM AUTHOR]

Introduction Pharmacological rhythm control of atrial fibrillation (AF) in patients with structural heart disease is limited. Ranolazine in combination with low dose dronedarone remarkably reduced AF-burden in the phase II HARMONY trial. We thus aimed to investigate the possible mechanisms underlying these results. Methods and results Patch clamp experiments revealed that ranolazine (5 μM), low-dose dronedarone (0.3 μM), and the combination significantly prolonged action potential duration (APD 90 ) in atrial myocytes from patients in sinus rhythm (prolongation by 23.5 ± 0.1%, 31.7 ± 0.1% and 25.6 ± 0.1% respectively). Most importantly, in atrial myocytes from patients with AF ranolazine alone, but more the combination with dronedarone, also prolonged the typically abbreviated APD 90 (prolongation by 21.6 ± 0.1% and 31.9 ± 0.1% respectively). It was clearly observed that neither ranolazine, dronedarone nor the combination significantly changed the APD or contractility and twitch force in ventricular myocytes or trabeculae from patients with heart failure (HF). Interestingly ranolazine, and more so the combination, but not dronedarone alone, caused hyperpolarization of the resting membrane potential in cardiomyocytes from AF. As measured by confocal microscopy (Fluo-3), ranolazine, dronedarone and the combination significantly suppressed diastolic sarcoplasmic reticulum (SR) Ca 2 + leak in myocytes from sinus rhythm (reduction by ranolazine: 89.0 ± 30.7%, dronedarone: 75.6 ± 27.4% and combination: 78.0 ± 27.2%), in myocytes from AF (reduction by ranolazine: 67.6 ± 33.7%, dronedarone: 86.5 ± 31.7% and combination: 81.0 ± 33.3%), as well as in myocytes from HF (reduction by ranolazine: 64.8 ± 26.5% and dronedarone: 65.9 ± 29.3%). Conclusions Electrophysiological measurements during exposure to ranolazine alone or in combination with low-dose dronedarone showed APD prolongation, cellular hyperpolarization and reduced SR Ca 2 + leak in human atrial myocytes. The combined inhibitory effects on various currents, in particular Na + and K + currents, may explain the anti-AF effects observed in the HARMONY trial. Therefore, the combination of ranolazine and dronedarone, but also ranolazine alone, may be promising new treatment options for AF, especially in patients with HF, and merit further clinical investigation. [ABSTRACT FROM AUTHOR]

Background--Recent evidence points to functional Ca2+-dependent K+ (SK) channels in the heart that may govern atrial fibrillation (AF) risk, but the underlying mechanisms are unclear. This study addressed the role of SK channels in atrial repolarization and AF persistence in a canine AF model. Methods and Results--Electrophysiological variables were assessed in dogs subjected to atrial remodeling by 7-day atrial tachypacing (AT-P), as well as controls. Ionic currents and single-channel properties were measured in isolated canine atrial cardiomyocytes by patch clamp. NS8593, a putative selective SK blocker, suppressed SK current with an IC50 of ≈5 μmol/L, without affecting Na+, Ca2+, or other K+ currents. Whole-cell SK current sensitive to NS8593 was significantly larger in pulmonary vein (PV) versus left atrial (LA) cells, without a difference in SK single-channel open probability (Po), whereas AT-P enhanced both whole-cell SK currents and single-channel Po. SK-current block increased action potential duration in both PV and LA cells after AT-P; but only in PV cells in absence of AT-P. SK2 expression was more abundant at both mRNA and protein levels for PV versus LA in control dogs, in both control and AT-P; AT-P upregulated only SK1 at the protein level. Intravenous administration of NS8593 (5 mg/kg) significantly prolonged atrial refractoriness and reduced AF duration without affecting the Wenckebach cycle length, left ventricular refractoriness, or blood pressure. Conclusions--SK currents play a role in canine atrial repolarization, are larger in PVs than LA, are enhanced by atrial-tachycardia remodeling, and appear to participate in promoting AF maintenance. These results are relevant to the potential mechanisms underlying the association between SK single-nucleotide polymorphisms and AF and suggest SK blockers as potentially interesting anti-AF drugs. [ABSTRACT FROM AUTHOR]

Background--Electrical, structural, and Ca2+-handling remodeling contribute to the perpetuation/progression of atrial fibrillation (AF). Recent evidence has suggested a role for spontaneous sarcoplasmic reticulum Ca2+-release events in long-standing persistent AF, but the occurrence and mechanisms of sarcoplasmic reticulum Ca2+-release events in paroxysmal AF (pAF) are unknown. Method and Results--Right-atrial appendages from control sinus rhythm patients or patients with pAF (last episode a median of 10-20 days preoperatively) were analyzed with simultaneous measurements of [Ca2+]j (fluo-3-acetoxymethyl ester) and membrane currents/action potentials (patch-clamp) in isolated atrial cardiomyocytes, and Western blot. Action potential duration, L-type Ca2+ current, and Na+/Ca2+-exchange current were unaltered in pAF, indicating the absence of AF-induced electrical remodeling. In contrast, there were increases in SR Ca2+ leak and incidence of delayed after-depolarizations in pAF. Ca2+-transient amplitude and sarcoplasmic reticulum Ca2+ load (caffeine-induced Ca2+-transient amplitude, integrated Na+/Ca2+-exchange current) were larger in pAE Ca2+-transient decay was faster in pAF, but the decay of caffeine-induced Ca2+ transients was unaltered, suggesting increased SERCA2a function. In agreement, phosphorylation (inactivation) of the SERCA2a-inhibitor protein phospholamban was increased in pAF. Ryanodine receptor fractional phosphorylation was unaltered in pAl, whereas ryanodine receptor expression and single-channel open probability were increased. A novel computational model of the human atrial cardiomyocyte indicated that both ryanodine receptor dysregulation and enhanced SERCA2a activity promote increased sarcoplasmic reticulum Ca2+ leak and sarcoplasmic reticulum Ca2+-release events, causing delayed after-depolarizations/triggered activity in pAF. Conclusions--Increased diastolic sarcoplasmic reticulum Ca2+ leak and related delayed after-depolarizations/triggered activity promote cellular arrhythmogenesis in pAF patients. Biochemical, functional, and modeling studies point to a combination of increased sarcoplasmic reticulum Ca2+ load related to phospholamban hyperphosphorylation and ryanodine receptor dysregulation as underlying mechanisms. [ABSTRACT FROM AUTHOR]

Abstract: Protein phosphorylation is a major control mechanism of a wide range of physiological processes and plays an important role in cardiac pathophysiology. Serine/threonine protein phosphatases control the dephosphorylation of a variety of cardiac proteins, thereby fine-tuning cardiac electrophysiology and function. Specificity of protein phosphatases type-1 and type-2A is achieved by multiprotein complexes that target the catalytic subunits to specific subcellular domains. Here, we describe the composition, regulation and target substrates of serine/threonine phosphatases in the heart. In addition, we provide an overview of pharmacological tools and genetic models to study the role of cardiac phosphatases. Finally, we review the role of protein phosphatases in the diseased heart, particularly in ventricular arrhythmias and atrial fibrillation and discuss their role as potential therapeutic targets. [Copyright &y& Elsevier]

Abstract: Atrial dilatation is an independent risk factor for thromboembolism in patients with and without atrial fibrillation (AF). In many patients, atrial dilatation goes along with depressed contractile function of the dilated atria. While some mechanisms causing atrial contractile dysfunction in fibrillating atria have been addressed previously, the cellular and molecular mechanisms of atrial contractile remodeling in dilated atria are unknown. This study characterized in vivo atrial contractile function in a goat model of atrial dilatation and compared it to a goat model of AF. Differences in the underlying mechanisms were elucidated by studying contractile function, electrophysiology and sarcoplasmic reticulum (SR) Ca2+ load in atrial muscle bundles and by analyzing expression and phosphorylation levels of key Ca2+-handling proteins, myofilaments and the expression and activity of their upstream regulators. In 7 chronically instrumented, awake goats atrial contractile dysfunction was monitored during 3 weeks of progressive atrial dilatation after AV-node ablation (AV block goats (AVB)). In open chest experiments atrial work index (AWI) and refractoriness were measured (10 goats with AVB, 5 goats with ten days of AF induced by repetitive atrial burst pacing (AF), 10 controls). Isometric force of contraction (FC), transmembrane action potentials (APs) and rapid cooling contractures (RCC, a measure of SR Ca2+ load) were studied in right atrial muscle bundles. Total and phosphorylated Ca2+-handling and myofilament protein levels were quantified by Western blot. In AVB goats, atrial size increased by 18% (from 26.6±4.4 to 31.6±5.5 mm, n =7 p <0.01) while atrial fractional shortening (AFS) decreased (from 18.4±1.7 to 12.8±4.0% at 400 ms, n =7, p <0.01). In open chest experiments, AWI was reduced in AVB and in AF goats compared to controls (at 400 ms: 8.4±0.9, n =7, and 3.2±1.8, n =5, vs 18.9±5.3 mm×mmHg, n =7, respectively, p <0.05 vs control). FC of isolated right atrial muscle bundles was reduced in AVB (n =8) and in AF (n =5) goats compared to controls (n =9) (at 2 Hz: 2.3±0.5 and 0.7±0.2 vs 5.5±1.0 mN/mm2, respectively, p <0.05). APs were shorter in AF, but unchanged in AVB goats. RCCs were reduced in AVB and AF versus control (AVB, 3.4±0.5 and AF, 4.1±1.4 vs 12.2±3.2 mN/mm2, p <0.05). Protein levels of protein kinase A (PKA) phosphorylated phospholamban (PLB) were reduced in AVB (n =8) and AF (n =8) vs control (n =7) by 37.9±12.4% and 29.7±10.1%, respectively (p <0.01), whereas calmodulin-dependent protein kinase II (CaMKII) phosphorylated ryanodine channels (RyR2) were increased by 166±55% in AVB (n =8) and by 146±56% in AF (n =8) goats (p <0.01). PKA-phosphorylated myosin-binding protein-C and troponin-I were reduced exclusively in AVB goat atria (by 75±10% and 55±15%, respectively, n =8, p <0.05). Atrial dilatation developing during slow ventricular rhythm after complete AV block as well as AF-induced remodeling are associated with atrial contractile dysfunction. Both AVB and AF goat atria show decreased SR Ca2+ load, likely caused by PLB dephosphorylation and RYR2 hyperphosphorylation. While shorter APs further compromise contractility in AF goat atria, reduced myofilament phosphorylation may impair contractility in AVB goat atria. Thus, atrial hypocontractility appears to have distinct molecular contributors in different types of atrial remodeling. [Copyright &y& Elsevier]

Cardiac arrhythmias are a global health burden, contributing significantly to morbidity and mortality worldwide. Despite technological advances in catheter ablation therapy, antiarrhythmic drugs (AADs) remain a cornerstone for the management of cardiac arrhythmias. Experimental and translational studies have shown that commonly used AADs exert multiple effects in the heart, the manifestation of which strongly depends on the exact experimental or clinical conditions. This diversity makes the optimal clinical application of AADs challenging. Here, we present a novel computational tool designed to facilitate a better understanding of the complex mechanisms of action of AADs (the Maastricht Antiarrhythmic Drug Evaluator, MANTA). In this tool, we integrated published computational cardiomyocyte models from different species (mouse, guinea pig, rabbit, dog, and human), regions (atrial, ventricular, and Purkinje cells) and disease conditions (atrial fibrillation- and heart failure-related remodeling). Subsequently, we investigated the effects of clinically available AADs (Vaughan-Williams Classes I, III, IV and multi-channel blockers) on action potential (AP) properties and the occurrence of proarrhythmic effects such as early afterdepolarizations. Steady-state drug effects were simulated based on a newly compiled overview of published IC 50 values for each cardiac ion channel and by integrating state-dependent block of the cardiac Na+-current by Class I AADs using a Markov-model approach. Using MANTA, we demonstrated and characterized important species-, rate-, cell-type-, and disease-state-specific AAD effects, including 1) a stronger effect of Class III AADs in large mammals than in rodents; 2) a rate-dependent decrease in upstroke velocity with Class I AADs and reverse rate-dependent effects of Class III AADs on action potential duration; 3) ventricular-predominant effects of pure I Kr blockers; 4) preferential reduction in atrial AP upstroke velocity with vernakalant; and 5) excessive AP prolongation with Class III AADs other than amiodarone under heart failure conditions. In conclusion, the effects of AADs are highly complex and strongly dependent on the experimental or clinical conditions. MANTA is a powerful and freely available tool reproducing a wide range of AAD characteristics that enables analyses of the underlying ionic mechanisms. Use of MANTA is expected to improve our understanding of AAD effects on cellular electrophysiology under a wide range of conditions, which may provide clinically-relevant information on the safety and efficacy of AAD treatment. [ABSTRACT FROM AUTHOR]

Introduction: Nucleoplasmic (Nuc) Ca2+ regulates gene expression; nuclear Ca2+ changes in AF are poorly understood. Hypothesis: AF modulates atrial cardiomyocyte (ACM) nuclear Ca2+ handling via IP3 receptor (IP3R) changes. Methods: ACMs were isolated from control dogs and dogs kept in AF for 1 wk by atrial pacing at 600 bpm. Nuc and cytoplasmic (cyto) Ca2+ were recorded via confocal imaging with Fluo-4 AM. Immunostaining (IS) and western blot (WB) were used to detect protein expression. Results: Diastolic [Ca2+]nuc was higher than [Ca2+]cytoat baseline and increased further in AF. In intact and saponin-permeabilized ACMs, ET-1 (100 nM) and IP3 (20 μM) increased diastolic [Ca2+]nuc and [Ca2+ ]cyto, with significantly larger changes in AF. IS and WB showed increased IP3R type 1 (IP3R1) expression in AF. ACMs paced at 3 Hz (P3) for 24 hours to mimic AF showed AF-like [Ca2+]nucchanges vs 1 Hz paced cells (P1, Figure). To evaluate the role of IP3R, we incubated P1 and P3 cells with siRNA targeting IP3R1 or IP3R2. IP3R1 knockdown (KD) suppressed P3-induced changes in nuclear Ca2+-handling (Fig A); IP3R2 KD produced no significant changes (Fig B). IS showed that IP3R1 KD prevented P3-induced IP3R1 fluorescence intensity (FI) upregulation (Fig C). KD reduced diastolic [Ca2+]nuc from 212±5 to 146±5 nM (n/N=96-140/6, cells/dogs; P<0.001) in P1 cells and 383±9 to 245±9 nM (n/N=91-104/6, P<0.001) in P3 cells. Like AF, P3 increased nuclear CaMKIIδ phosphorylation-ratio and activated HDAC4 nuclear export vs P1 (Fig C). Autocamtide CaMKII inhibitory peptide (AIP) suppressed P3 effects on nuclear CaMKII phosphorylation and HDAC4 translocation. IP3R1 KD reversed P3 effects on nuclear CaMKIIδ phosphorylation-ratio and prevented nuclear HDAC4 export (Fig C). Conclusions: AF modulates ACM nuc Ca2+ handling to increase nuclear Ca2+ load by IP3R1 upregulation, leading to enhanced nuclear IP3R1-CaMKII-HDAC4 signaling and potential gene-dysregulation via altered epigenetic control. [ABSTRACT FROM AUTHOR]

Background: Electrical storm (ES) is a life-threatening complication of defibrillator (ICD)-treated arrhythmic syndromes, including long QT syndrome (LQTS). Hypothesis: ICD-shocks for recurrent ventricular tachyarrhythmias cause accentuated action potential duration (APD) heterogeneities, a substrate of severe ES in LQTS. Methods: A model of ES was created by inducing chronic complete AV block in ICD-implanted rabbits, causing LQTS, repetitive Torsades de Pointes (TdP) and frequent ventricular fibrillation (VF) episodes. Results: Optical mapping in perfused hearts from 13 rabbits with ES events (defined as 3 or more VF episodes per 24 h) showed APD prolongation, particularly at the LV base (e.g. by 47% at 400 ms cycle length, p<0.05, vs. control (CTL)) and detected a large island-like long-APD region (Fig A) in 10 rabbits. The number of VF episodes correlated with the APD and its dispersion ex vivo (B). Epinephrine exacerbated the island by remarkably prolonging APDs, leading to strikingly increased APD dispersion, which disappeared in response to 10 μM ranolazine. TdP was initiated by an ectopic beat that failed to enter the refractory island and formed a reentrant wave, and maintained by meandering rotors in the periphery of the island (C). Late Na+ current (INa-L) was increased in ES rabbit myocytes (0.94±0.21% of peak current, n=8) vs. CTLs (0.21±0.05%, n=7, p<0.05). Na+ channel NaV1.5 was hyperphosphorylated at Ser571 by 140% (n=9, p<0.05) along with upregulation of phosphorylated and oxidized Ca2+/calmodulin-dependent protein kinase II by 185 and 195%, respectively, in ES rabbit LV tissues (n=9, p<0.01). Epinephrine infusion evoked TdP in vivo. Lidocaine (2 mg/kg i.v.) ameliorated QT-U abnormalities and completely suppressed TdP in 4 epinephrine-loaded rabbits. Conclusions: A tissue-island with enhanced refractoriness created by increased INa,Lcontributes to the generation of meandering rotors in a unique manner, which underlies LQTS-associated ES. [ABSTRACT FROM AUTHOR]