Mechanisms of termination and prevention of atrial fibrillation by drug therapy

Atrial fibrillation (AF) is a disorder of the rhythm of electrical activation of the cardiac atria. It is the most common cardiac arrhythmia, has multiple aetiologies, and increases the risk of death from stroke. Pharmacological therapy is the mainstay of treatment for AF, but currently available anti-arrhythmic drugs have limited efficacy and safety. An improved understanding of how anti-arrhythmic drugs affect the electrophysiological mechanisms of AF initiation and maintenance, in the setting of the different cardiac diseases that predispose to AF, is therefore required. A variety of animal models of AF has been developed, to represent and control the pathophysiological causes and risk factors of AF, and to permit the measurement of detailed and invasive parameters relating to the associated electrophysiological mechanisms of AF. The purpose of this review is to examine, consolidate and compare available relevant data on in-vivo electrophysiological mechanisms of AF suppression by currently approved and investigational anti-arrhythmic drugs in such models. These include the Vaughan Williams class I–IV drugs, namely Na+ channel blockers, β-adrenoceptor antagonists, action potential prolonging drugs, and Ca2+ channel blockers; the “upstream therapies”, e.g., angiotensin converting enzyme inhibitors, statins and fish oils; and a variety of investigational drugs such as “atrial-selective” multiple ion channel blockers, gap junction-enhancers, and intracellular Ca2+-handling modulators. It is hoped that this will help to clarify the main electrophysiological mechanisms of action of different and related drug types in different disease settings, and the likely clinical significance and potential future exploitation of such mechanisms. Keywords: Atrial fibrillation; Cardiac arrhythmia mechanisms: reentry, afterdepolarisations; In-vivo animal models; Pathological electrical remodelling; Pharmacological treatment; Anti-arrhythmic drug mechanisms Abbreviations: ACE, angiotensin-converting enzyme; AF, atrial fibrillation; AFCL, AF cycle length; APD, action potential duration; DAD, delayed afterdepolarisation; EAD, early afterdepolarisation; ERP, effective refractory period; ICaL, L-type Ca2+ current; ICaT, T-type Ca2+ current; If, funny current; IK1, inward rectifier K+ current; IKACh, acetylcholine-activated K+ current; IKr, rapid delayed rectifier K+ current; IKS, slow delayed rectifier K+ current; IKur, ultra-rapid delayed rectifier K+ current; INa, Na+ current; INa/Ca, Na+-Ca2+ exchanger current; INa/H, Na+-H+ exchanger current; INaL, late INa; ISKCa, small conductance Ca2+-activated K+ current; ITO, transient outward K+ curren

Mechanisms of Atrial Arrhythmia: Investigations of the Neuro-Myogenic Interface in the Mouse

Arrhythmia mechanisms rely on multiple factors including structural (myogenic), nervous (neurogenic), and interrelated (the neuro-myogenic interface) factors. I hypothesized that due to this neuro-myogenic interface, the intrinsic cardiac autonomic nervous system (ICANS) is involved in most atrial arrhythmias. This thesis also provides a Threshold Model as a tool to assess the role of different physiological factors influencing arrhythmia. This model allows relative comparison and interpretation of the role of various factors influencing arrhythmogenesis. The mouse allows relatively simple manipulation of genes to determine their role in arrhythmia. This thesis determined what atrial arrhythmias are inducible in the mouse (in vivo) and how to systematically study those arrhythmias. I found that atrial tachycardia/fibrillation (AT/F) and junctional tachycardia (JT) are inducible in the mouse. AF and JT pose significant clinical challenges as many patients do not respond well to current interventions. Neurogenic AF relies on acetylcholine, while myogenic AF relies, in part on gap junctions formed by connexins (Cxs). The atria has muscarinic M2 and M3 receptors. The duration of M2R/M3R G protein signalling is regulated by GTP hydrolysis, a process accelerated by the regulators of G protein signalling (RGS). RGS2 deficient (RGS2-/-) mice had reduced refractory periods that were normalized with a selective M3R blocker (Darifenacin) and increased susceptibility to AT/F induction compared to littermates. For the first time, this showed a role of M3 and RGS in atrial arrhythmia. Cx40 deficient (Cx40-/-) mice were protected from carbachol induced AT/F, while Cx43 G60S mutant (Cx43G60S/+) mice, with an 80% reduction in phospho-Cx43 in the atria were highly susceptible to AT/F that was terminated by darifenacin. This shows a novel neurogenic component to what was previously described as myogenic arrhythmia. Another novel finding was that JT has a neurogenic component, resulting from inappropriate AV nodal pacemaker activation initiated by autonomics. Ivabradine hydrochloride, a selective pacemaker channel blocker, prevented JT and may be useful in patients with JT. In conclusion, this thesis has provided novel findings of the vital role of the neuro-myogenic interface in atrial arrhythmias and has provided the basis for future investigations of potential therapeutic options for patients

The Effect of Diet on Cardiovascular Disease, Heart Disease and Blood Vessels

Cardiovascular disease (CVD), including coronary artery disease, heart disease, arrhythmias, and other types of vascular diseases, is one of the leading causes of death around the world. It is estimated that approximately half of the variabilities of CVD appear to be attributed to genetics. Therefore, the other half of them have been attributed to acquired factors, including diet. It is of note that even a genetic predisposition to CVD can be canceled out by a healthy lifestyle. In this regard, it is important to acknowledge that acquired factors, including diet, are causally associated with CVD. Based on these facts, important papers are presented in this Special Issue entitled “The Effect of Diet on Cardiovascular Disease, Heart Disease, and Blood Vessels”

Electrophysiological Effects of Lysoplasmenylcholine on Rabbit Ventricular Myocytes

Myocardial ischemia activates a phospholipase A2 that targets plasmalogen phospholipids and liberates 1-0-alkenyl-Iysoplasmenylcholine (LPLC) in preference to 1-0- acyl-Iysophosphatidylcholine (LPC). Although LPC is a pro arrhythmic ischemic metabolite, the effect of LPLC on cardiac electrophysiology is unknown. At the lowest doses investigated, LPLC induced spontaneous contractions in otherwise quiescent rabbit ventricular myocytes significantly faster than LPC. Spontaneous contractions developed with median times of 16.4 (n = 64), 27.4 (n = 36), and \u3e60 min (n = 25) during exposure to 5, 2.5, and 1 JlM LPLC compared with 38.0 (n = 48) and \u3e60 min (n = 29) for 5 and 2.5 JlM LPC, respectively. Median times for 10 JlM lysolipids were not different. To characterize the mechanism of spontaneous activity, membrane potential (Em) and whole-cell currents were measured. LPLC caused an abrupt and sustained depolarization of Em by z 50 m V and culminated in the loss of excitability (n = 7). Voltage-clamp analysis of steady-state currents revealed an inward current at the normal resting Em that reversed at - 18.5 ± 0.9 m V (n = 12). The reversal potential of this current was insensitive to Ca-channel blockade by Cd2+ (n = 3), or by lowering bath [Cl-]. However, a lO-fold reduction in bath [Na+] caused repolarization and reduced the inward current by 56.6 ± 3.6% at -83 m V (n = 4). In contrast, Na-channel blockade by tetrodotoxin (n = 4) or saxitoxin (n =3) failed to inhibit membrane depolarization or the current induced by LPLC. Two lanthanides were studied to determine if the LPLC current was mediated by stretch-activated channels (SACs). Gd3+ ( 100 IlM), a known SAC blocker, and La3+ (100 IlM), devoid of SAC blocking activity, inhibited the LPLC-induced current by 80.2 ± 8.3% (n = 7) and 80.7 ± 8.3 % (n = 6), respectively, at -83 mY. Exposure to hypertonic bathing medium and cell shrinkage failed to restore Em (n = 5) or inhibit the LPLC-induced current (n = 2), which confirmed that lanthanides were not acting through inhibition of SACs. Consistent with the effects on membrane current, pretreatment with 100 IlM Gd3+ or La3+ but not Cd2+, significantly delayed spontaneous activity in 5 IlM LPLC (median times: Gd3+, 55.4 min (n = 35); La3+, 53.0 min (n = 38); Cd2+, 17.4 min (n = 19)). Lanthanides increase phospholipid ordering and may oppose membrane perturbations induced by LPLC. LPLC may contribute to ventricular dysrhythmias during ischemia

Investigating the protective role of the natural hormone Melatonin, in reducing drug-induced cardiotoxicity in the therapy of chronic diseases

Heart failure (HF) is a highly complex disorder and a major end-point of cardiovascular diseases (CVD). The pathogenesis of HF is mostly unresolved but involves interplay between cardiac structural and electrical remodelling, metabolic alterations, cell death and altered gene expression. Mitochondrial dysfunction and HF are common complications of chronic treatment from diverse groups of drugs, in particular anticancer drugs such as doxorubicin (DOX). Treatment of animals and cardiomyocytes with cardiotoxic chemicals such as β-adrenergic receptor agonists (such as isoproterenol) induces cardiac dysfunction and HF. Previous work done by the group have identified the pineal hormone melatonin was protective against stress-induced cardiac arrhythmias and simulated heart failure in cardiomyocytes in vitro. Melatonin synthesis is also dramatically decreased with age and in patients with CVD. The aim of the present project was to better understand the pathogenesis of druginduced cardiac dysfunction and delineate the role of melatonin in cardioprotection in H9c2, a model rat cell line in vitro. Using the Seahorse XF analyser method, it was demonstrated that commonly used medication for chronic diseases such as amiodarone, amitriptyline, and statins all caused altered mitochondrial dysfunction. In addition, cardiotoxic chemicals (isoproterenol, hydrogen peroxide, DOX) altered oxidative phosphorylation and glycolysis in living cardiomyocyte-derived H9c2 cells; these deleterious metabolic changes were ameliorated by melatonin. Flowcytometry and Alamar Blue staining methods demonstrated that DOX robustly induced apoptosis in H9c2 cells (~30%) which was reversed by melatonin. Doxorubicin-induced stress in H9c2 cells dramatically altered gene expression in several key signalling pathways integral in cardiac function and disease. These included mitochondrial metabolism (UCP2, PPARɣ, Drp1, Mfn1, Parp 1, Parp2, Sirt3 and Cav3), apoptosis (Bcl2 and Bcl-xL), cardiac electrophysiology and arrhythmia (Scn5a, SERCA2a), calcium handling (SERCA2a) and cardiac remodelling (Myh7, ms1). Melatonin pre-treatment attenuated or completely blocked this DOX-induced alteration in gene expression in cardiomyocytes. In conclusion, the present result demonstrated for the first time that melatonin is cardioprotective against drug-induced cardiotoxicity and apoptosis via modifying diverse heart failure-related signalling pathways. This provides novel insight on the possible use of melatonin as an adjunct intervention in several therapies including anti-cancer

The Role of MicroRNA Regulation of Cardiac Ion Channel in Arrhythmia

La fibrillation auriculaire (FA) est le trouble du rythme le plus fréquemment observé en pratique clinique. Elle constitue un risque important de morbi-mortalité. Le traitement de la FA reste un défi majeur en lien avec les nombreux effets secondaires associés aux approches thérapeutiques actuelles. Dans ce contexte, une meilleure compréhension des mécanismes sous-jacents à la FA est essentielle pour le développement de nouvelles thérapies offrant un meilleur rapport bénéfice/risque pour les patients. La FA est caractérisée par i) un remodelage électrique délétère associé le plus souvent ii) à un remodelage structurel du myocarde favorisant la récurrence et le maintien de l’arythmie. La diminution de la période réfractaire effective au sein du tissu auriculaire est un élément clef du remodelage électrique. Le remodelage structurel, quant à lui, se manifeste principalement par une fibrose tissulaire qui altère la propagation de l’influx électrique dans les oreillettes. Les mécanismes moléculaires impliqués dans la mise en place de ces deux substrats restent mal connus. Récemment, le rôle des microARNs (miARNs) a été pointé du doigt dans de nombreuses pathologies notamment cardiaques. Dans ce contexte les objectifs principaux de ce travail ont été i) d'acquérir une compréhension approfondie du rôle des miARNs dans la régulation de l’expression des canaux ioniques et ii) de mieux comprendre le rôle de ces molécules dans l’installation d’un substrat favorable a la FA. Nous avons, dans un premier temps, effectué une analyse bio-informatique combinée à des approches expérimentales spécifiques afin d’identifier clairement les miARNs démontrant un fort potentiel de régulation des gènes codant pour l’expression des canaux ioniques cardiaques humains. Nous avons identifié un nombre limité de miARNs cardiaques qui possédaient ces propriétés. Sur la base de ces résultats, nous avons démontré que l’altération de l'expression des canaux ioniques, observée dans diverse maladies cardiaques (par exemple, les cardiomyopathies, l’ischémie myocardique, et la fibrillation auriculaire), peut être soumise à ces miARNs suggérant leur implication dans l’arythmogénèse. La régulation du courant potassique IK1 est un facteur déterminant du remodelage électrique auriculaire associée à la FA. Les mécanismes moléculaires sous-jacents sont peu connus. Nous avons émis l’hypothèse que l'altération de l’expression des miARNs soit corrélée à l’augmentation de l’expression d’IK1 dans la FA. Nous avons constaté que l’expression de miR-26 est réduite dans la FA et qu’elle régule IK1 en modulant l’expression de sa sous-unité Kir2.1. Nous avons démontré que miR-26 est sous la répression transcriptionnelle du facteur nucléaire des lymphocytes T activés (NFAT) et que l’activité accrue de NFATc3/c4, aboutit à une expression réduite de miR-26. En conséquence IK1 augmente lors de la FA. Nous avons enfin démontré que l’interférence in vivo de miR-26 influence la susceptibilité à la FA en régulant IK1, confirmant le rôle prépondérant de miR-26 dans le remodelage auriculaire électrique. La fibrose auriculaire est un constituant majeur du remodelage structurel associé à la FA, impliquant l'activation des fibroblastes et l’influx cellulaire du Ca2 +. Nous avons cherché à déterminer i) si le canal perméable au Ca2+, TRPC3, jouait un rôle dans la fibrose auriculaire en favorisant l'activation des fibroblastes et ii) étudié le rôle potentiel des miARNs dans ce contexte. Nous avons démontré que les canaux TRPC3 favorisent l’influx du Ca2 +, activant la signalisation Ca2 +-dépendante ERK et en conséquence activent la prolifération des fibroblastes. Nous avons également démontré que l’expression du TRPC3 est augmentée dans la FA et que le blocage in vivo de TRPC3 empêche le développement de substrats reliés à la FA. Nous avons par ailleurs validé que miR-26 régule les canaux TRPC3 en diminuant leur expression dans les fibroblastes. Enfin, nous avons montré que l'expression réduite du miR-26 est également due à l’activité augmentée de NFATc3/c4 dans les fibroblastes, expliquant ainsi l’augmentation de TRPC3 lors de la FA, confirmant la contribution de miR-26 dans le processus de remodelage structurel lié à la FA. En conclusion, nos résultats mettent en évidence l'importance des miARNs dans la régulation des canaux ioniques cardiaques. Notamment, miR-26 joue un rôle important dans le remodelage électrique et structurel associé à la FA et ce, en régulant IK1 et l’expression du canal TRPC3. Notre étude démasque ainsi un mécanisme moléculaire de contrôle de la FA innovateur associant des miARNs. miR-26 en particulier représente apres ces travaux une nouvelle cible thérapeutique prometteuse pour traiter la FA.Atrial fibrillation (AF) is the most frequently-encountered arrhythmia in clinical practice and constitutes a major cause of cardiac morbidity and mortality. The management of AF remains a major challenge as current therapeutic approaches are limited by potential adverse effects and high rate of AF recurrence/persistence. A better understanding of the mechanisms underlying AF is of great importance to improve AF therapy. AF is characterized by impaired electrical and structural remodeling, both of which favors the recurrence and maintenance of the arrhythmia. A key feature in electrical remodeling is the reduced atrial effective refractory period, due to ion channel alteration. Structural remodeling, on the other hand, mainly results from atrial fibrosis. However, the precise molecular mechanisms underlying these remodeling processes are still incompletely understood. The importance of microRNAs (miRNAs) in various pathophysiological conditions of the heart has been well established, but little is known with regard to cardiac arrhythmias. Emerging evidence suggests that dysregulation of miRNAs may underlie heart rhythm disturbances. The aim of the present work was to acquire a comprehensive understanding of miRNA-mediated regulation of ion channels in cardiac arrhythmias. Notably, we will focus on the mechanistic insights of miRNAs related to the control of AF. Currently available experimental approaches do not permit thorough characterization of miRNA targeting. For this purpose, we performed bioinformatic analyses in conjunction with experimental approaches to identify miRNAs from the database that potentially regulate human cardiac ion channel genes. We found that only a subset of miRNAs target cardiac ion channel genes. Based on these results, we further demonstrated that the dysregulation of ion channel gene expression observed in various cardiac disorders (e.g. cardiomyopathy, myocardial ischemia, and atrial fibrillation) can be explained by the dysregulation of miRNAs. These findings further support the potential implication of miRNAs in arrhythmogenesis under these cardiac conditions. The upregulation of the cardiac inward rectifying potassium current, IK1, is a key determinant of adverse atrial electrical remodeling associated with AF. The molecular mechanisms underlying this ionic remodeling are poorly understood. We hypothesized that altered miRNA expression is responsible for IK1 upregulation in AF. We found that miR-26 is significantly downregulated in AF and regulates IK1 by controlling the expression of its underlying subunit Kir2.1. Moreover, we demonstrated that miR-26 is under the transcriptional repression of the nuclear factor of activated T cells (NFAT) and enhanced activities of members of the NFAT family, NFATc3/c4, results in miR-26 downregulation, which accounts for IK1 enhancement in AF. Furthermore, we observed that in vivo interference of miR-26 affects AF susceptibility via the regulation of IK1, suggesting an important role of miR-26 in atrial electrical remodeling. Atrial fibrosis is a major constituent in AF-associated adverse atrial structural remodeling, involving the activation of fibroblasts and cellular Ca2+ entry. Here, we sought to determine whether the Ca2+ permeable channel, TRPC3, plays a role in AF-induced fibrosis by promoting fibroblast activation. Furthermore, we investigated the potential role of miRNAs in this context. We found that TRPC3 channels promote Ca2+-entry, which results in activation of Ca2+-dependent ERK-signaling and consequently fibroblast activation. We also demonstrated that TRPC3 is upregulated in AF and in vivo TRPC3 blockade suppresses the development of AF-promoting substrate. Furthermore, we observed that miR-26 regulates TRPC3 channels via controlling the expression of the underlying channel subunit and is downregulated in AF-fibroblasts. Finally, we showed that the reduced expression of miR-26 is also due to the enhanced NFATc3/c4 activities in AF-fibroblasts and accounts for AF-induced upregulation of TRPC3, suggesting the potential contribution of miR-26 in AF-related adverse structural remodeling process. In conclusion, our findings emphasize the importance of miRNAs in the regulation of cardiac ion channels. Notably, miR-26 plays a crucial role in AF-associated electrical and structural remodeling via the regulation of IK1 and TRPC3 channel genes. Thus, our study unravels a novel molecular control mechanism of AF at the miRNA level, suggesting miR-26 as a new and promising therapeutic target for AF

Adenosine and its role in cardioplegia : experimental evaluation in the isolated rat heart and in an-vivo primate model

This study was designed to investigate the role of adenosine, an endogenous cardioprotectant agent, without high potassium and as cardioplegic additive to high potassium solutions. Adenosine cardioplegia and potassium cardioplegia supplemented by adenosine (K + ADO) were investigated in terms of hemodynamic, metabolic and ultrastructural recovery in the isolated rat heart and in the in-vivo baboon model during periods of global myocardial ischemia, simulating the clinical situation during open heart surgery. The results obtained in both models show that adenosine improved postischemic hemodynamic function when used without high potassium cardioplegia. The combination of adenosine and high potassium was less effective in both models in terms of hemodynamic recovery; however, improved rhythm stability and coronary vasodilatation were still present. In addition adenosine alone was able to induce fast electromechanical arrest in the isolated rat heart. However, failure of even high concentrations of adenosine to limit ventricular fibrillation in the baboon exclude its use as cardioplegic agent on its own without additional interventions. It appears likely that adenosine without high potassium is cardioprotective via activation of A₁ receptors and opening of ATP-sensitive potassium channels, a mechanism which is probably non-functional in a high potassium environment. In view of the limited cardioprotection achieved with the combination of adenosine and high potassium further studies should aim for additional interventions to induce cardioplegia with adenosine and normokalemic solutions