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Aims: In mammalian ventricles, electrical gradients establish electrical heterogeneities as essential tissue mechanisms to optimize mechanical efficiency and safeguard electrical stability. Electrical gradients shape mammalian electrocardiographic patterns; disturbance of electrical gradients is proarrhythmic. The zebrafish heart is a popular surrogate model for human cardiac electrophysiology thanks to its remarkable recapitulation of human electrocardiogram and ventricular action potential features. Yet, zebrafish ventricular electrical gradients are largely unexplored. The goal of this study is to define the zebrafish ventricular electrical gradients that shape the QRS complex and T wave patterns at baseline and under oxidative stress., Methods and Results: We performed in vivo electrocardiography and ex vivo voltage-sensitive fluorescent epicardial and transmural optical mapping of adult zebrafish hearts at baseline and during acute H2O2 exposure. At baseline, apicobasal activation and basoapical repolarization gradients accounted for the polarity concordance between the QRS complex and T wave. During H2O2 exposure, differential regional impairment of activation and repolarization at the apex and base disrupted prior to baseline electrical gradients, resulting in either reversal or loss of polarity concordance between the QRS complex and T wave. KN-93, a specific calcium/calmodulin-dependent protein kinase II inhibitor (CaMKII), protected zebrafish hearts from H2O2 disruption of electrical gradients. The protection was complete if administered prior to oxidative stress exposure., Conclusions: Despite remarkable apparent similarities, zebrafish and human ventricular electrocardiographic patterns are mirror images supported by opposite electrical gradients. Like mammalian ventricles, zebrafish ventricles are also susceptible to H2O2 proarrhythmic perturbation via CaMKII activation. Our findings suggest that the adult zebrafish heart may constitute a clinically relevant model to investigate ventricular arrhythmias induced by oxidative stress. However, the fundamental ventricular activation and repolarization differences between the two species that we demonstrated in this study highlight the potential limitations when extrapolating results from zebrafish experiments to human cardiac electrophysiology, arrhythmias, and drug toxicities., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2020. For permissions, please email: journals.permissions@oup.com.)

Despite key advances in the clinical management of life-threatening ventricular arrhythmias, culminating with the development of implantable cardioverter-defibrillators and catheter ablation techniques, pharmacologic/biologic therapeutics have lagged behind. The fundamental issue is that biological targets are molecular factors. Diseases, however, represent emergent properties at the scale of the organism that result from dynamic interactions between multiple constantly changing molecular factors. For a pharmacologic/biologic therapy to be effective, it must target the dynamic processes that underlie the disease. Here we propose a classification of ventricular arrhythmias that is based on our current understanding of the dynamics occurring at the subcellular, cellular, tissue and organism scales, which cause arrhythmias by simultaneously generating arrhythmia triggers and exacerbating tissue vulnerability. The goal is to create a framework that systematically links these key dynamic factors together with fixed factors (structural and electrophysiological heterogeneity) synergistically promoting electrical dispersion and increased arrhythmia risk to molecular factors that can serve as biological targets. We classify ventricular arrhythmias into three primary dynamic categories related generally to unstable Ca cycling, reduced repolarization, and excess repolarization, respectively. The clinical syndromes, arrhythmia mechanisms, dynamic factors and what is known about their molecular counterparts are discussed. Based on this framework, we propose a computational-experimental strategy for exploring the links between molecular factors, fixed factors and dynamic factors that underlie life-threatening ventricular arrhythmias. The ultimate objective is to facilitate drug development by creating an in silico platform to evaluate and predict comprehensively how molecular interventions affect not only a single targeted arrhythmia, but all primary arrhythmia dynamics categories as well as normal cardiac excitation-contraction coupling., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

In the healthy heart, cardiac myocytes form an electrical syncytium embedded in a supportive fibroblast-rich extracellular matrix designed to optimize the electromechanical coupling for maximal contractile efficiency of the heart. In the injured heart, however, fibroblasts are activated and differentiate into myofibroblasts that proliferate and generate fibrosis as a component of the wound-healing response. This review discusses how fibroblasts and fibrosis, while essential for maintaining the structural integrity of the heart wall after injury, have undesirable electrophysiological effects by disrupting the normal electrical connectivity of cardiac tissue to increase the vulnerability to arrhythmias. We emphasize the dual contribution of fibrosis in altering source-sink relationships to create a vulnerable substrate while simultaneously facilitating the emergence of triggers such as afterdepolarization-induced premature ventricular complexes-both factors combining synergistically to promote initiation of reentry. We also discuss the potential role of fibroblasts and myofibroblasts in directly altering myocyte electrophysiology in a pro-arrhythmic fashion. Insight into these processes may open up novel therapeutic strategies for preventing and treating arrhythmias in the setting of heart disease as well as avoiding potential arrhythmogenic consequences of cell-based cardiac regeneration therapy. This article is part of a Special Issue entitled "Myocyte-Fibroblast Signaling in Myocardium.", (© 2013.)

Unlike young hearts, aged hearts are highly susceptible to early afterdepolarization (EAD)-mediated ventricular fibrillation (VF). This differential may result from age-related structural remodeling (fibrosis) or electrical remodeling of ventricular myocytes or both. We used optical mapping and microelectrode recordings in Langendorff-perfused hearts and patch-clamp recordings in isolated ventricular myocytes from aged (24-26 mo) and young (3-4 mo) rats to assess susceptibility to EADs and VF during either oxidative stress with ANG II (2 μM) or ionic stress with hypokalemia (2.7 mM). ANG II caused EAD-mediated VF in 16 of 19 aged hearts (83%) after 32 ± 7 min but in 0 of 9 young hearts (0%). ANG II-mediated VF was suppressed with KN-93 (Ca(2+)/calmodulin-dependent kinase inhibitor) and the reducing agent N-acetylcysteine. Hypokalemia caused EAD-mediated VF in 11 of 11 aged hearts (100%) after 7.4 ± 0.4 min. In 14 young hearts, however, VF did not occur in 6 hearts (43%) or was delayed in onset (31 ± 22 min, P < 0.05) in 8 hearts (57%). In patch-clamped myocytes, ANG II and hypokalemia (n = 6) induced EADs and triggered activity in both age groups (P = not significant) at a cycle length of >0.5 s. When myocytes of either age group were coupled to a virtual fibroblast using the dynamic patch-clamp technique, EADs arose in both groups at a cycle length of <0.5 s. Aged ventricles had significantly greater fibrosis and reduced connexin43 gap junction density compared with young hearts. The lack of differential age-related sensitivity at the single cell level in EAD susceptibility indicates that increased ventricular fibrosis in the aged heart plays a key role in increasing vulnerability to VF induced by oxidative and ionic stress.

Aims: Fibrosis is known to promote cardiac arrhythmias by disrupting myocardial structure. Given recent evidence that myofibroblasts form gap junctions with myocytes at least in co-cultures, we investigated whether myofibroblast-myocyte coupling can promote arrhythmia triggers, such as early afterdepolarizations (EADs), by directly influencing myocyte electrophysiology., Methods and Results: Using the dynamic voltage clamp technique, patch-clamped adult rabbit ventricular myocytes were electrotonically coupled to one or multiple virtual fibroblasts or myofibroblasts programmed with eight combinations of capacitance, membrane resistance, resting membrane potential, and gap junction coupling resistance, spanning physiologically realistic ranges. Myocytes were exposed to oxidative (1 mmol/L H(2)O(2)) or ionic (2.7 mmol/L hypokalaemia) stress to induce bradycardia-dependent EADs. In the absence of myofibroblast-myocyte coupling, EADs developed during slow pacing (6 s), but were completely suppressed by faster pacing (1 s). However, in the presence of myofibroblast-myocyte coupling, EADs could no longer be suppressed by rapid pacing, especially when myofibroblast resting membrane potential was depolarized (-25 mV). Analysis of the myofibroblast-myocyte virtual gap junction currents revealed two components: an early transient-outward I(to)-like current and a late sustained current. Selective elimination of the I(to)-like component prevented EADs, whereas selective elimination of the late component did not., Conclusion: Coupling of myocytes to myofibroblasts promotes EAD formation as a result of a mismatch in early vs. late repolarization reserve caused by the I(to)-like component of the gap junction current. These cellular and ionic mechanisms may contribute to the pro-arrhythmic risk in fibrotic hearts.

Mutations in SCN5A encoding the principal Na+ channel alpha-subunit expressed in human heart (Na(V)1.5) have recently been linked to an inherited form of dilated cardiomyopathy with atrial and ventricular arrhythmia. We compared the biophysical properties of 2 novel Na(V)1.5 mutations associated with this syndrome (D2/S4--R814W; D4/S3--D1595H) with the wild-type (WT) channel using heterologous expression in cultured tsA201 cells and whole-cell patch-clamp recording. Expression levels were similar among WT and mutant channels, and neither mutation affected persistent sodium current. R814W channels exhibited prominent and novel defects in the kinetics and voltage dependence of activation characterized by slower rise times and a hyperpolarized conductance-voltage relationship resulting in an increased "window current." This mutant also displayed enhanced slow inactivation and greater use-dependent reduction in peak current at fast pulsing frequencies. By contrast, D1595H channels exhibited impaired fast inactivation characterized by slower entry into the inactivated state and a hyperpolarized steady-state inactivation curve. Our findings illustrate the divergent biophysical defects caused by 2 different SCN5A mutations associated with familial dilated cardiomyopathy. Retrospective review of the published clinical data suggested that cardiomyopathy was not common in the family with D1595H, but rather sinus bradycardia was the predominant clinical finding. However, for R814W, we speculate that an increased window current coupled with enhanced slow inactivation and rate-dependent loss of channel availability provided a unique substrate predisposing myocytes to disordered Na+ and Ca2+ homeostasis leading to myocardial dysfunction.