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Interleukin (IL)-33, a cytokine involved in immune responses, can activate its receptor, suppression of tumorigenicity 2 (ST2), is elevated during atrial fibrillation (AF). However, the role of IL-33/ST2 signaling in atrial arrhythmia is unclear. This study explored the pathological effects of the IL-33/ST2 axis on atrial remodeling and arrhythmogenesis. Patch clamping, confocal microscopy, and Western blotting were used to analyze the electrical characteristics of and protein activity in atrial myocytes (HL-1) treated with recombinant IL-33 protein and/or ST2-neutralizing antibodies for 48 hrs. Telemetric electrocardiographic recordings, Masson's trichrome staining, and immunohistochemistry staining of the atrium were performed in mice receiving tail vein injections with nonspecific immunoglobulin (control), IL-33, and IL-33 combined with anti-ST2 antibody for 2 weeks. IL-33-treated HL-1 cells had a reduced action potential duration, lower L-type Ca2+ current, greater sarcoplasmic reticulum (SR) Ca2+ content, increased Na+/Ca2+ exchanger (NCX) current, elevation of K+ currents, and increased intracellular calcium transient. IL-33-treated HL-1 myocytes had greater activation of the calcium–calmodulin-dependent protein kinase II (CaMKII)/ryanodine receptor 2 (RyR2) axis and nuclear factor kappa B (NF-κB) / NLR family pyrin domain containing 3 (NLRP3) signaling than did control cells. IL-33 treated cells also had greater expression of Nav1.5, Kv1.5, NCX, and NLRP3 than did control cells. Pretreatment with neutralizing anti-ST2 antibody attenuated IL-33-mediated activation of CaMKII/RyR2 and NF-κB/NLRP3 signaling. IL-33-injected mice had more atrial ectopic beats and increased AF episodes, greater atrial fibrosis, and elevation of NF-κB/NLRP3 signaling than did controls or mice treated with IL-33 combined with anti-ST2 antibody. Thus, IL-33 recombinant protein treatment promotes atrial remodeling through ST2 signaling. Blocking the IL-33/ST2 axis might be an innovative therapeutic approach for patients with atrial arrhythmia and elevated serum IL-33. [ABSTRACT FROM AUTHOR]

Chronic obstructive pulmonary disease (COPD) increases the risk of atrial fibrillation (AF), however, its arrhythmogenic mechanisms are unclear. This study investigated the effects of COPD on AF triggers (pulmonary veins, PVs) and substrates (atria), and their potential underlying mechanisms. Electrocardiographic, echocardiographic, and biochemical studies were conducted in control rabbits and rabbits with human leukocyte elastase (0.3 unit/kg)-induced COPD. Conventional microelectrode, Western blotting, and histological examinations were performed on PV, left atrium (LA), right atrium, and sinoatrial node (SAN) preparations from control rabbits and those with COPD. The rabbits with COPD had a higher incidence of atrial premature complexes, PV burst firing and delayed afterdepolarizations, higher sympathetic activity, larger LA, and faster PV spontaneous activity than did the control rabbits; but they exhibited a slower SAN beating rate. The LA of the rabbits with COPD had a shorter action potential duration and longer tachyarrhythmia induced by tachypacing (20 Hz) and isoproterenol (1 μM). Additionally, the rabbits with COPD had higher fibrosis in the PVs, LA, and SAN. H89 (10 μM), KN93 (1 μM), and KB-R7943 (10 μM) significantly suppressed burst firing and delayed afterdepolarizations in the PVs of the rabbits with COPD. Moreover, compared with the control rabbits, those with COPD had lower expression levels of the β1 adrenergic receptor, Cav 1.2, and Na+/Ca2+ exchanger in the PVs; Cav 1.2 in the LA; and hyperpolarization-activated cyclic nucleotide-gated K+ channel 4 in the SAN. COPD increases atrial arrhythmogenesis by modulating the distinctive electrophysiological characteristics of the PVs, LA, and SAN. [ABSTRACT FROM AUTHOR]

Macrophage migration inhibitory factor (MIF), a pleiotropic inflammatory cytokine, is highly expressed in patients with atrial fibrillation (AF). CD74 (major histocompatibility complex, class II invariant chain) is the main receptor for MIF. However, the role of the MIF/CD74 axis in atrial arrhythmogenesis is unclear. In this study, we investigated the effects of MIF/CD74 signaling on atrial electrophysiological characteristics and determined its underlying mechanisms. Confocal fluorescence microscopy, patch clamp, and western blot analysis were used to study calcium homeostasis, ionic currents, and calcium-related signaling in MIF-treated HL-1 atrial cardiomyocytes with or without anti-CD74 neutralized antibodies treatment. Furthermore, electrocardiographic telemetry recording and echocardiography were obtained from mice treated with MIF. Compared with controls, MIF-treated HL-1 myocytes had increased calcium transients, sarcoplasmic reticulum (SR) calcium content, Na+/Ca2+ exchanger (NCX) efflux rate, calcium leak, transient outward potassium current, and ultra-rapid delayed rectifier potassium current. Furthermore, MIF could induce expression of SR Ca2+ATPase, NCX, phosphorylation of ryanodine receptor 2 (RyR2), and activation of calcium/calmodulin kinase II (CaMKII) when compared with control cells. MIF-mediated electrical dysregulation and CaMKII-RyR2 signaling activation were attenuated through blocking of CD74. Moreover, MIF-injected mice had lesser left atrium fractional shortening, greater atrial fibrosis, and atrial ectopic beats than control (nonspecific immunoglobulin treated) or MIF combined with anti-CD74 neutralized antibody-treated mice. Consequently, our study on MIF/CD74 signaling has pointed out a new potential therapeutic intervention of AF patients with MIF elevation. [ABSTRACT FROM AUTHOR]

Background: Ivabradine increases the risk of atrial fibrillation (AF). Heart failure (HF) or sinoatrial node (SAN) dysfunction increases the risk of AF, and pulmonary veins (PVs) play a critical role in the pathophysiology of AF. This study investigated the electrophysiologic effects of ivabradine on SANs and PVs in a rabbit model of HF.Methods and Results: Conventional microelectrodes were used to simultaneously record the electrical activities and conduction properties of control and HF rabbit SAN-PV preparations before and after perfusion with ivabradine (0.1, 1, or 10 μmol/L), either alone or with isoproterenol (1 μmol/L). HF SANs exhibited a lower beating rate than the control SANs. SAN automaticity exit blocks and SAN-PV conduction blocks were observed in 25% and 50% of samples, respectively, with P < .05 for HF SANs (n = 8) but not for control SANs (n = 6). Delayed afterdepolarization (DAD) was observed in 37.5% of HF PVs but not in control PVs. HF PVs exhibited a faster beating rate and more severe fibrosis than control PVs. Ivabradine reduced the SAN beating rates and increased the occurrences of SAN-PV conduction blocks and PV DADs in control and HF preparations. However, ivabradine induced SAN automaticity exit blocks only in HF preparations. Isoproterenol induced PV burst firing and shifting electrical conduction in control and HF preparations. A combination of isoproterenol and ivabradine (10 μmol/L) in HF preparations resulted in the highest incidences of PV burst firing and SAN-PV electrical shifting.Conclusions: HF differentially modulates the effects of ivabradine on the electrical activities of SAN and PVs, which may increase PV arrhythmogenesis and contribute to the risk of AF in HF patients. [ABSTRACT FROM AUTHOR]

Cardiac surgery is complicated with atrial fibrillation (AF). Histone deacetylase (HDAC) inhibition reduces AF occurrence. In pericarditis, HDAC inhibition may modulate AF trigger and substrate. We recorded electrocardiograms in control and pericardiotomic (op) rabbits without and with an intraperitoneal injection of MPT0E014 (HDAC inhibitor). Conventional microelectrodes recorded action potentials (APs) in pulmonary veins (PVs), the right and left atrium (LA). Masson's trichrome was used to identify collagen fibers in PVs and the LA. Electrocardiograms showed frequent atrial premature contractions in op rabbits, but not in the other 3 groups. The beating rates in PVs and opPVs were decreased by MPT0E014 treatment. Spontaneous burst firings occurred in opPVs (36.4%), but not in control PVs. H2O2 induced greater burst firings in opPVs (72.7%) than in control PVs (11.1%), MPT0E014-treated PVs (16.7%), and MPT0E014-treated opPVs (12.5%). The AP duration at a repolarization extent of 90% (APD90) was shorter in the opLA than that in the control LA. In the presence of isoproterenol (1 μM), rapid atrial pacing (RAP, 20 Hz) induced a higher incidence of burst firings in the opLA (90%) than in the other groups. In contrast, acetylcholine (5 mM) and RAP induced a lower incidence of burst firing in the MPT0E014-treated LA (33.3%) than in the other groups. Fibrosis prevailed in opPVs and the opLA compared to the respective control PVs and LA, which was attenuated in those that received MPT0E014. In conclusion, a pericardiotomy increased fibrosis and arrhythmogenesis in PVs and the LA, which were prevented by HDAC inhibition. [ABSTRACT FROM AUTHOR]

Glucagon like-peptide-1 (GLP-1) is an incretin hormone with antidiabetic effects through stimulating insulin secretion, β cell neogenesis, satiety sensation, and inhibiting glucagon secretion. Administration of GLP-1 provides cardioprotective effects through attenuating cardiac inflammation and insulin resistance. GLP-1 also modulates the heart rate and systolic pressure, which suggests that GLP-1 may have cardiac electrical effects. Therefore, the purposes of this study were to evaluate whether GLP-1 has direct cardiac effects and identify the underlying mechanisms. Patch clamp, confocal microscopy with Fluo-3 fluorescence, and Western blot analyses were used to evaluate the electrophysiological characteristics, calcium homeostasis, and calcium regulatory proteins in HL-1 atrial myocytes with and without GLP-1 (1 and 10 nM) incubation for 24 h. GLP-1 (1 and 10 nM) and control cells had similar action potential durations. However, GLP-1 at 10 nM significantly increased calcium transients and sarcoplasmic reticular Ca 2+ contents. Compared to the control, GLP-1 (10 nM)—treated cells significantly decreased phosphorylation of the ryanodine receptor at S2814 and total phospholamban, but there were similar protein levels of sarcoplasmic reticular Ca 2+ -ATPase and the sodium–calcium exchanger. Moreover, exendin (9–39) amide (a GLP-1 receptor antagonist, 10 nM) attenuated GLP-1-mediated effects on total SR content and phosphorylated ryanodine receptor S2814. This study demonstrates GLP-1 may regulate HL-1 cell arrhythmogenesis through modulating calcium handling proteins. [ABSTRACT FROM AUTHOR]

Background: Hypokalemia and hyponatremia increase the occurrence of atrial fibrillation. Sinoatrial nodes (SANs) and pulmonary veins (PVs) play a critical role in the pathophysiology of atrial fibrillation.Objective: The purpose of this study was to evaluate whether electrolyte disturbances with low concentrations of potassium ([K(+)]) or sodium ([Na(+)]) modulate SAN and PV electrical activity and arrhythmogenesis, and to investigate potential underlying mechanisms.Methods: Conventional microelectrodes were used to record electrical activity in rabbit SAN and PV tissue preparations before and after perfusion with different low [K(+)] or [Na(+)], interacting with the Na(+)-Ca(2+) exchanger inhibitor KB-R7943 (10 μΜ).Results: Low [K(+)] (3.5, 3, 2.5, and 2 mM) decreased beating rates in PV cardiomyocytes with genesis of delayed afterdepolarizations (DADs), burst firing, and increased diastolic tension. Low [K(+)] (3.5, 3, 2.5, and 2 mM) also decreased SAN beating rates, with genesis of DADs. Low [Na(+)] increased PV diastolic tension, DADs, and burst firing, which was attenuated in the co-superfusion with low [K(+)] (2 mM). In contrast, low [Na(+)] had little effect on SAN electrical activities. KB-R7943 (10 μΜ) reduced the occurrences of low [K(+)] (2 mM)- or low [Na(+)] (110 mM)-induced DAD and burst firing in both PVs and SANs.Conclusion: Low [K(+)] and low [Na(+)] differentially modulate SAN and PV electrical properties. Low [K(+)]- or low [Na(+)]-induced slowing of SAN beating rate and genesis of PV burst firing may contribute to the high occurrence of atrial fibrillation during hypokalemia or hyponatremia. [ABSTRACT FROM AUTHOR]

Background Renal failure (RF) increases the risk of atrial fibrillation (AF), but arrhythmogenic mechanism is unclear. The present study investigated the electrophysiological effects of RF on AF trigger (pulmonary veins, PVs) and substrate (atria) and evaluated potential underlying mechanisms. Methods Electrocardiographic, echocardiographic, and biochemical studies were conducted in rabbits with and without antibiotic-induced mild (creatinine = 1.5–6.0 mg/dl) and advanced (creatinine > 6.0 mg/dl) RF. Conventional microelectrode techniques, western blotting, and histological examinations were performed using the isolated rabbit PV, left atrium (LA), right atrium (RA) and sinoatrial node (SAN). Results Advanced RF rabbits (n = 18) had a higher incidence (33.3% vs. 11.1% and 0%, p < 0.05) of atrial arrhythmia than mild RF (n = 18) and control (n = 18) rabbits. Advanced RF rabbits exhibited faster PV spontaneous activities, longer action potential duration (APD) in the LA, higher fibrosis in the LA, and slower SAN beating rates than control rabbits, but had a similar APD and fibrosis in the RA. Caffeine (1 mM) increased advanced RF PV arrhythmogenesis, which is blocked by flecainide (10 μM), or KB-R7943 (10 μM). Moreover, advanced RF rabbits had a higher expression of the Na + /Ca 2 + exchanger, protein kinase A, phosphorylated ryanodine receptor (Serine 2808), and phosphorylated phospholamban (Serine 16) in PVs, and a higher expression of Ca v 1.2 in the LA, and a lower expression of hyperpolarization-activated cyclic nucleotide-gated potassium channel 4 in the SAN. Conclusions Advanced RF increases atrial arrhythmia by modulating the distinctive electrophysiological characteristics of the PV, LA, and SAN. [ABSTRACT FROM AUTHOR]

Background Resveratrol has shown benefits in reducing ventricular remodeling and arrhythmias. Objective This study aimed to assess the therapeutic efficacy of resveratrol in reducing atrial fibrillation (AF) in a heart failure (HF) model and to explore the underlying mechanisms. Methods HF rabbits were created 4 weeks after undergoing coronary ligation. Group 1 (n = 6) was divided into subgroups of (a) normal rabbits, (b) HF sham rabbits, and (c) HF rabbits treated for 1 week with intraperitoneal injections of resveratrol, (d) resveratrol plus wortmannin, or (e) resveratrol plus diphenyleneiodonium chloride (DPI). All rabbits underwent epicardial catheter stimulation. Collagen content, messenger RNA and protein expression in ion channels, and phosphoinositide 3-kinase (PI3K)/AKT/endothelial nitric oxide synthase (eNOS) signaling pathways were studied in left atrial appendage (LAA) preparations. To investigate acute drug effects on left atrial electrophysiology, groups 2 a through 2e (n = 6 per group) were subjected to Langendorff perfusion. Results Higher AF inducibility was found in the HF group and groups that were given PI3K and eNOS inhibitors than in the normal and resveratrol-treated groups ( P < .001). Histologic analysis of the LAA revealed a decrease in fibrosis in resveratrol-treated groups compared with the HF group (8.95% ± 1.53% vs 26.62% ± 2.19%, P < .001). In real-time polymerase chain reaction analysis, ion channels including Kv1.4, Kv1.5, KvLQT1, Kir2.1, Nav1.5, Cav1.2, NCX, SERCA2a, and phospholamban were upregulated by resveratrol. PI3K, AKT, and eNOS messenger RNA and protein expression were upregulated by resveratrol but were inhibited by the coadministration of wortmannin and DPI. Conclusion Resveratrol decreases left atrial fibrosis and regulates variation in ion channels to reduce AF through the PI3K/AKT/eNOS signaling pathway. [ABSTRACT FROM AUTHOR]

Background: Sinoatrial node (SAN) dysfunction increases the occurrences of atrial fibrillation (AF). The pulmonary veins (PVs) play a critical role in the pathophysiology of AF. The purpose of this study was to evaluate whether SAN electrical activity can modulate PV arrhythmogenesis. Methods: Conventional microelectrodes and multi-electrode array system were used to simultaneously record the electrical activity and conduction properties of rabbit SAN and PV tissue preparations with and without SAN–PV interruptions before and after perfusion with Anemonia sulcata toxin (ATX)-II (100nM) or isoproterenol (1μM). Results: ATX-II significantly increased PV beating rates, which overdrove SAN electrical activity with the occurrences of PV burst firings in 5 (56%) of 9 tissue preparations, and induced SAN–PV conduction block in 6 (67%) of 9 preparations. After SAN–PV disconnection, ATX-II induced burst firing and early afterdepolarizations in 8 (89%) of 9 PVs. Moreover, the multi-electrode array found that ATX-II reversed the electrical conduction between the SAN and PV with an increase in electrical activity from 1.8±0.6 to 2.9±0.6Hz (P <0.05) in SAN–PV preparations (n =7). In contrast, isoproterenol did not reverse electrical conduction between the SAN and PV with an increase in electrical activity from 1.8±0.2 to 3.0±0.3Hz (P <0.005) in SAN–PV preparations (n =7). Conclusions: SAN electrical activity modulates PV arrhythmogenesis. SAN–PV conduction blocks can increase PV arrhythmogenesis. [ABSTRACT FROM AUTHOR]

Abstract: Background: Sex and sodium/calcium regulation play critical roles in cardiac electrophysiology and atrial arrhythmogenesis. We investigated whether sodium and calcium contributed to sex differences in atrial electrophysiology. Methods: Whole-cell patch clamp techniques and the indo-1 fluorometric ratio technique were used to investigate the ionic current and intracellular calcium in single isolated male and female rabbit myocytes from the left atrium posterior wall (LAPW) and right atrium (RA). Results: Female LAPW (n =95) and RA (n =49) myocytes had larger cell widths (15.1±0.4 vs. 13.8±0.4μm, p <0.05; 14.9±0.6 vs. 13.5±0.4μm, p <0.05) than male LAPW (n =142) and RA (n =57) myocytes. Male LAPW myocytes (n =26) had a higher incidence (57 vs. 16%, p <0.05) of delayed afterdepolarizations (DADs) than female LAPW myocytes (n =24) but there were similar incidences (20 vs. 20%, p >0.05) of DADs in male and female RA myocytes. The late sodium current, calcium transients, and sarcoplasmic reticulum calcium contents were larger in male than female LAPW myocytes but were similar in male and female RA myocytes. However, the I Ca-L and nickel-sensitive sodium/calcium exchanger currents were similar between two groups. Different from those in female myocytes, ouabain (10μM) only induced repeated atrial beats (0 to 45%, p <0.05) in male myocytes (n =11). Moreover, ranolazine (3μM) perfusion (4.5±0.6 vs. 1min, p <0.05) was required to decrease the amplitude of DADs in male but not female LAPW myocytes. Conclusions: Increased late sodium currents and calcium contents may contribute to higher arrhythmogenesis in male LAPW myocytes. [Copyright &y& Elsevier]

Abstract: Background: Geranylgeranylacetone (GGA) has been reported up-regulating heat shock protein (HSP) expression, and protecting against atrial remodeling. This study aimed to investigate the effects of GGA on atrial electrophysiology and inducibility of atrial fibrillation (AF) in heart failure (HF) model. Methods and results: HF rabbits were created 4weeks after coronary artery ligation. Monophasic action potential recordings and multielectrode array were used to record the electrophysiological characteristics of left atrium (LA) in normal, or HF rabbits with (HF-GGA) and without (HF-control) oral administration of GGA (200mg/kg, 24h before experiments). The mRNA and protein expressions of ionic channels were measured by Western blot and PCR. HF-GGA LA (n=10), similar to normal LA (n=10) had a shorter action potential duration (APD) and effective refractory period than HF-control LA (n=10). HF-GGA LA had less triggered activity and APD alternans (20% vs. 100%, P=0.001), lower maxima slope of restitution curve of APD (0.94±0.04 vs.1.69±0.04, P<0.001), and less inducibility of AF (50% vs. 100%, P=0.033) than HF-control LA. HF-GGA LA had a shorter activation time and higher conduction velocity than HF-control LA. HF-GGA LA had a higher mRNA expression of Cav1.2, Nav1.5, Kir2.1, Kv1.4, Kv7.1, Kv11.1, sarcoplasmic reticulum Ca2+-ATPase, and higher phosphorylation of phospholamban than HF-control LA. Conclusions: GGA decreases triggered activity, dispersion of APD and inducibility of AF in failing heart through induction of HSP, and modulation of ionic channels and calcium homeostasis. [Copyright &y& Elsevier]

Background: Obesity is an important risk factor for atrial fibrillation (AF) and heart failure (HF). The effects of epicardial fat on atrial electrophysiology were not clear. This study was to evaluate whether HF may modulate the effects of epicardial fat on atrial electrophysiology. Methods: Conventional microelectrodes recording was used to record the action potential in left (LA) and right (RA) atria of healthy (control) rabbits before and after application of epicardial fat from control or HF (ventricular pacing of 360–400bpm for 4weeks) rabbits. Adipokine profiles were checked in epicardial fat of control and HF rabbits. Results: The LA 90% of AP duration was prolonged by control epicardial fat (from 77±6 to 87±7ms, p <0.05, n =7), and by HF epicardial fat (from 78±3 to 98±4ms, p <0.001, n =9). However, control or HF epicardial fat did not change the AP morphology in RA. HF epicardial fat increased the contractility in LA (61±11 vs. 35±6mg, p =0.001), but not in RA. Control fat did not change the LA or RA contractility. Moreover, control and HF epicardial fat induced early and delayed afterdepolarizations in LA and RA, but only HF epicardial fat provoked spontaneous activity and burst firing in LA (n =3/9, 33.3% vs. n =0/7, 0%, n =0/9, 0%, p <0.05). Compared to control fat, HF epicardial fat, had lower resistin, C-reactive protein and serum amyloid A, but similar interluekin-6, leptin, monocyte chemotactic protein-1, adiponectin and adipsin. Conclusions: HF epicardial fat increases atrial arrhythmogenesis, which may contribute to the higher atrial arrhythmia in obesity. [ABSTRACT FROM AUTHOR]

Abstract: Background: Diabetes and hypertension have significant effects on cardiac calcium (Ca2+) regulation, which plays an essential role in determining cardiac function. The effect of peroxisome proliferator-activated receptor (PPAR)-γ agonists on Ca2+ regulation in the cardiomyocytes is unclear. Objective: The purpose of this study was to investigate the effects of hypertension, diabetes, and PPAR-γ agonist-rosiglitazone on the regulation of Ca2+ and the electrophysiological characteristics of isolated ventricular myocytes. Methods: The indo-1 fluorometric ratio technique and whole-cell patch clamp were used to investigate intracellular Ca2+ (Ca2+ i), action potentials, and ionic currents in ventricular myocytes from rats of Wistar–Kyoto (WKY), diabetic WKY (induced by streptozotocin), diabetic WKY treated with rosiglitazone (5mg/kg), spontaneously hypertensive rats (SHR), diabetic SHR, and diabetic SHR treated with rosiglitazone. Western blot was used to evaluate protein expressions of sarcoplasmic reticulum ATPase (SERCA2a), Na+–Ca2+ exchanger (NCX), and ryanodine receptor (RyR). Results: Diabetic WKY and diabetic SHR had smaller sarcoplasmic reticulum Ca2+ contents, and Ca2+ i transients with a prolonged decay portion, down-regulated SERCA2a, NCX, and RyR protein expressions and smaller L-type Ca2+ currents than non-diabetic WKY and SHR, respectively. The Ca2+ dysregulations in diabetes were attenuated in rats treated with rosiglitazone. Diabetes and hypertension both prolonged the action potential duration which were enhanced by the use of rosiglitazone, and induced the genesis of triggered activity. Conclusions: Diabetes and hypertension modulate Ca2+ handling. Rosiglitazone significantly changed the Ca2+ regulation and electrophysiological characteristics, and may contain an arrhythmogenic potential in diabetes with hypertension. [Copyright &y& Elsevier]

Abstract: Dronedarone and amiodarone are anti-atrial fibrillation agents with different potency. Pulmonary veins play a critical role in the genesis of atrial fibrillation. Oxidative stress can enhance pulmonary vein arrhythmogenesis. This study was done to compare the effects of dronedarone and amiodarone on pulmonary vein electrophysiological and mechanical properties, and oxidative stress-induced arrhythmogenecity. Conventional microelectrodes were used to record action potentials in isolated rabbit pulmonary vein specimens before and after dronedarone and amiodarone with or without the presence of H2O2 (2mM). Dronedarone (0.1, 1 and 10μM) concentration-dependently decreased pulmonary vein beating rates (from 2.2±0.1 to 1.9±0.1, 1.8±0.1 and 1.7±0.1Hz, n=8, P<0.01). Amiodarone (0.1, 1 and 10μM) also concentration-dependent decreased pulmonary vein beating rates (from 2.5±0.2 to 2.3±0.2, 2.2±0.2 and 2.0±0.2Hz, n=7, P<0.01). However, dronedarone decreased pulmonary vein beating rates to a greater extent at 0.1μM (12% versus 4%, P<0.005) and 1μM (17% versus 9%, P<0.005). Dronedarone or amiodarone (0.1, 1 and 10μM) did not change the pulmonary vein contractility. However, dronedarone (1 and 10μM) concentration-dependent reduced pulmonary vein diastolic tension by 13±2mg (P<0.005) and 18±3mg (P<0.005). In contrast, amiodarone did not change pulmonary vein diastolic tension. Dronedarone (10μM) and amiodarone (10μM) attenuated H2O2-induced pulmonary vein burst firings from 100% to 33.3% (P<0.01), and to 0% (P<0.005), respectively. In conclusion, amiodarone and dronedarone both significantly reduced pulmonary vein spontaneous beating rates and H2O2-induced pulmonary vein arrhythmogenesis. However, only dronedarone produced pulmonary vein vasodilation. [Copyright &y& Elsevier]

Abstract: Cardiomyocytes consist of single- and bi-nucleus myocytes. However, the electrophysiological characteristics of mononucleated and binucleated myocytes have never been elucidated. Left atrium (LA) and pulmonary veins (PVs) are important substrate and initiators of atrial fibrillation. The purposes of this study were to evaluate the electrical properties and calcium homeostasis in mononucleated and binucleated cardiomyocytes in the LA and PVs. A whole-cell clamp, fluo-4 fluorescence, and immunocytostaining were used to investigated mononucleated and binucleated cardiomyocytes in the LA and PVs. Both mononucleated PV and LA cardiomyocytes had more positive resting membrane potential than respective PV and LA binucleated cardiomyocytes. Additionally, mononucleated PV cardiomyocytes (n = 36) had faster beating rates (2.1 ± 0.2 Hz versus 1.0 ± 0.2 Hz, P < 0.05) than binucleated (n = 10) PV cardiomyocytes. The PV (n = 18) and LA (n = 15) mononucleated cardiomyocytes had larger [Ca2+]i transients (F/F 0 1.64 ± 0.09 versus 1.20 ± 0.03 IU, P < 0.05; 1.52 ± 0.06 versus 1.19 ± 0.05 IU, P < 0.05) than the binucleated PV (n = 10) and LA (n = 10) cardiomyocytes. The immunostaining showed that mononucleated cardiomyocytes had lower Kir 2.3 and higher ryanodine receptor densities than did binucleated cardiomyocytes both in the PV and LA. In conclusions, mononucleated PV and LA cardiomyocytes contain distinctive electrophysiological characteristics with a higher arrhythmogenic activity, which indicates that cell nucleus number may potentially determine the electrical activity and calcium handling in cardiomyocytes. [Copyright &y& Elsevier]

Abstract: Aims: Omega-3 polyunsaturated fatty acids can modulate cardiac electrophysiology and reduce the genesis of atrial fibrillation. This study investigates the potential mechanisms through which eicosapentaenoic acid (EPA) reduces pulmonary vein (PV) arrhythmogenesis. Main methods: Conventional microelectrodes were used to record the action potentials (APs), before and after the EPA (0.1μM and 1.0μM) administration with and without the presence of a nitric oxide (NO) synthase inhibitor (L-NAME, 100μM) in isolated rabbit PV tissue preparations. Furthermore, indo-1 fluorimetric ratio technique was used to evaluate intracellular calcium in isolated single PV cardiomyocytes with or without incubation of EPA (1.0μM, 30min). Key findings: EPA concentration-dependently reduced the PV spontaneous beating rate (P<0.05). EPA (1.0μM) also reduced the amplitude of delayed afterdepolarizations (P<0.05). EPA hyperpolarized the maximal diastolic potential (MDP), shortened AP duration, increased AP amplitude (APA), and reduced diastolic tension and contractility. However, EPA in the presence of L-NAME or omega-9 fatty acids (oleic acid, 1.0μM) did not have any effect on PV spontaneous activity, AP morphology, or contractile force. A linear regression shows that the decrease in PV spontaneous beating rates induced by EPA correlated well with the changes of MDP, APA, diastolic tension, and contractile force of PVs. In addition, intracellular Ca2+ transient and sarcoplasmic reticulum Ca2+ content were significantly more decreased in the EPA-treated cardiomyocytes than in control PV cardiomyocytes as observed by indo-1 fluorescence. Significance: EPA reduces PV arrhythmogenesis through the mechanoelectrical feedback generated by NO production. [Copyright &y& Elsevier]

Abstract: Background: Myocardial fibrosis plays a critical role in heart failure, resulting in cardiac structural and electrical remodeling which can induce atrial arrhythmias. Collagen is the major element of fibrosis. However, it is not clear whether collagen can directly regulate the calcium homeostasis and the electrophysiologic characteristics of cardiomyocytes. The aim of this study was to determine the effects of collagen on calcium homeostasis and the electrical properties of atrial cardiomyocytes. Methods and Results: HL-1 cardiomyocytes were cultured with and without collagen type I (1 or 10 μg/mL) or losartan (10 μmol/L). Whole-cell clamp, indo-1 fluorescence, and Western blotting were used to evaluate the action potential (AP) and ionic currents, intracellular calcium homeostasis, and calcium regulatory proteins. Compared with the control samples, there was no significant difference in collagen (1 μg/mL)–treated HL-1 cardiomyocytes. However, collagen (10 μg/mL)–treated HL-1 cardiomyocytes exhibited larger intracellular calcium ([Ca2+]i) transients by 113% and a larger sarcoplasmic reticulum calcium content by 86%. Collagen (10 μg/mL)–treated HL-1 cardiomyocytes had higher expression of sarcoplasmic reticulum ATPase (SERCA2a) and Thr17-phosphorylated phospholamban but similar protein expressions of the Na+/Ca2+ exchanger and ryanodine receptor. Collagen (10 μg/mL)–treated HL-1 cardiomyocytes (n = 11) had larger AP amplitude (104 ± 5 vs 83 ± 7 mV; P < .05), and shorter 90% of AP duration (25 ± 2 vs 33 ± 2 ms, P < .05) than control cells (n = 11). Moreover, collagen (10 μg/mL)–treated HL-1 cells had larger Ito and IKsus values than control cells. The administration of losartan (10 μmol/L) attenuated collagen-induced changes in [Ca2+]i transients, [Ca2+]i stores, AP morphology, ionic currents, SERCA2a, and Thr17-phosphorylated phospholamban expressions. Conclusions: This study demonstrates that collagen can directly modulate the calcium dynamics and electrical activities of atrial cardiomyocytes, which are associated with the renin-angiotensin system. These findings suggest a critical role of collagen in electrical remodeling during fibrosis. [Copyright &y& Elsevier]

Abstract: Aims: Lactic acidosis causes atrial fibrillation (AF), and pulmonary veins (PVs) are the most important focus for the generation of AF. Cariporide (HOE642), a Na+/H+ blocker, can prevent atrial tachycardia-induced electrical remodeling. The purpose of this study was to investigate whether cariporide can prevent lactic acidosis-induced PV arrhythmogenesis. Main methods: Conventional microelectrodes were used to record the action potentials (APs) before and after the administration of lactic acid (10 and 20 mM) in the absence and presence of cariporide (10 µM) pretreatment in isolated rabbit PV and atrial tissue preparations. Key findings: Lactic acidosis of 10 mM (pH 7.0±0.1) and 20 mM (pH 6.7±0.1) reduced PV (n =6) spontaneous rates from 2.5±0.3 to 1.6±0.4 (by 36±1%) and 1.1±0.4 Hz (by 56±2%), respectively, but lactic acidosis (10 and 20 mM) induced 12 episodes (3.9±0.2 Hz) and 23 episodes (4.0±0.3 Hz) of non-sustained burst firings in 4 PV specimens. Lactic acidosis (10 and 20 mM) decreased the AP amplitude (APA) and velocity of depolarization (Vmax), but increased the resting membrane potential (RMP), AP duration, and strength-response interval (SRI) in the PV and atrium. In the presence of cariporide (10 µM), lactic acidosis (10 and 20 mM) only reduced PV spontaneous rates from 2.4±0.2 to 1.8±0.4 (by 25±1%) and 1.6±0.4 Hz (by 33±1%), respectively, and prevented lactic acidosis-induced PV burst firings. Cariporide also reduced the effects of lactic acidosis on the RMP, AP duration, APA, Vmax, and SRI. Significance: Lactic acidosis has significant arrhythmogenic effects on PVs, which may be attenuated by Na+/H+ blockers. [Copyright &y& Elsevier]

Abstract: Pulmonary veins (PVs) contain cardiomyocytes with spontaneous activity that may be responsible for PV arrhythmia. Abnormal Ca2+ regulation is known to contribute to PV arrhythmogenesis. The purpose of this study was to investigate whether PV cardiomyocytes with spontaneous activity have different intracellular Ca2+ ([Ca2+]i) transients, Ca2+ sparks and responses to isoproterenol and ryanodine receptor modulators (magnesium and FK506) than do PV cardiomyocytes without spontaneous activity and left atrial (LA) cardiomyocytes. Through fluorescence and confocal microscopy, we evaluated the [Ca2+]i transients and Ca2+ sparks in isolated rabbit PV and LA cardiomyocytes. PV cardiomyocytes with spontaneous activity had larger [Ca2+]i transients and sarcoplasmic reticulum (SR) Ca2+ stores than PV cardiomyocytes without spontaneous activity or LA cardiomyocytes. PV cardiomyocytes with spontaneous activity also had a higher incidence and frequency of Ca2+ sparks, and had Ca2+ sparks with larger amplitudes than other cardiomyocytes. Magnesium (5.4 mM) reduced the [Ca2+]i transient amplitude and beating rate in PV cardiomyocytes with spontaneous activity. However, in contrast with other cardiomyocytes, low doses (1.8 mM) of magnesium did not reduce the [Ca2+]i transients amplitude in PV cardiomyocytes with spontaneous activity. FK506 (1 μM) diminished the SR Ca2+ stores in PV cardiomyocytes with spontaneous activity to a lesser extent than that in other cardiomyocytes. Isoproterenol (10 nM) increased the [Ca2+]i transient amplitude to a lesser extent in LA cardiomyocytes than in PV cardiomyocytes with or without spontaneous activity. In conclusion, our results suggest that enhanced [Ca2+]i transients, increased Ca2+ sparks and SR Ca2+ stores may contribute to the spontaneous activity of PV cardiomyocytes. [Copyright &y& Elsevier]

Background: Aging and pulmonary veins (PVs) play a critical role in the pathophysiology of atrial fibrillation. Abnormal Ca2+ regulation and ryanodine receptors are known to contribute to PV arrhythmogenesis. Objective: The purpose of this study was to investigate whether aging alters PV electrophysiology, Ca2+ regulation proteins, and responses to rapamycin, FK-506, ryanodine, and ouabain. Methods: Conventional microelectrodes were used to record action potential and contractility in isolated PV tissue samples in 15 young (age 3 months) and 16 aged (age 3 years) rabbits before and after drug administration. Expression of sarcoplasmic reticulum Ca2+ ATPase (SERCA2a), ryanodine receptor, and Na+/Ca2+ exchanger was evaluated by western blot. Results: Aged PVs had larger amplitude of delayed afterdepolarizations, greater depolarized resting membrane potential, longer action potential duration, and higher incidence of action potential alternans and contractile alternans with increased expression of Na+/Ca2+ exchanger and ryanodine receptor and decreased expression of SERCA2a. Rapamycin (1,10,100 nM), FK-506 (0.01, 0.1, 1 μM), ryanodine (0.1, 1 μM), and ouabain (0.1, 1 μM) concentration-dependently increased PV spontaneous rates and the incidence of delayed afterdepolarizations in young and aged PVs. Compared with results in young PVs, rapamycin and FK-506 in aged PVs increased PV spontaneous rates to a greater extent and exhibited a larger delayed afterdepolarization amplitude. In PVs without spontaneous activity, rapamycin and FK-506 induced spontaneous activity only in aged PVs, but ryanodine and ouabain induced spontaneous activity in both young and aged PVs. Conclusion: Aging increases PV arrhythmogenesis via abnormal Ca2+ regulation. These findings support the concept that ryanodine receptor dysfunction may result in high PV arrhythmogenesis and aging-related arrhythmogenic vulnerability. [Copyright &y& Elsevier]

Abstract: Inflammation and abnormal calcium homeostasis play important roles in atrial fibrillation. Tumor necrosis factor-α (TNFα), a proinflammatory cytokine, can induce cardiac arrhythmias. Pulmonary veins (PVs) are critical in initiating paroxysmal atrial fibrillation. This study was designed to investigate whether TNFα may change the calcium handling and arrhythmogenic activity of PV cardiomyocytes. We used whole-cell patch clamp and indo-1 fluorimetric ratio technique to investigate the action potentials, ionic currents and intracellular calcium in isolated rabbit single PV cardiomyocytes with and without (control) incubation with TNFα (25 ng/ml) for 7–10 h. The expression of sarcoplasmic reticulum ATPase in the control and TNFα-treated PV cardiomyocytes was evaluated by confocal micrographs and Western blot. We found that the spontaneous beating rates were similar between the control (n =45) and TNFα-treated (n =28) PV cardiomyocytes. Compared with the control PV cardiomyocytes, the TNFα-treated PV cardiomyocytes had significantly a larger amplitude of the delayed afterdepolarizations (6.0±1.7 vs. 2.6±0.8 mV, P <0.05), smaller L-type calcium currents, larger transient inward currents, larger Na+–Ca2+ exchanger currents, a smaller intracellular calcium transient, smaller sarcoplasmic reticulum calcium content, larger diastolic intracellular calcium, a longer decay portion of the calcium transient (Tau), and a decreased sarcoplasmic reticulum ATPase expression. In conclusion, TNFα can increase the PV arrhythmogenicity and induce an abnormal calcium homeostasis, thereby causing inflammation-related atrial fibrillation. [Copyright &y& Elsevier]

: ObjectivesThis study was conducted to investigate the effects of thyroid hormone on the electrophysiological characteristics of pulmonary vein (PV) cardiomyocytes.: BackgroundHyperthyroidism is an important etiology of paroxysmal atrial fibrillation (AF). Pulmonary veins are known to initiate paroxysmal AF.: MethodsThe action potential and ionic currents were investigated in single rabbit PV and atrial cardiomyocytes with (hyperthyroid) and without (control) incubation of L-triiodothyronine using the whole-cell clamp technique.: ResultsCompared with the control cardiomyocytes, hyperthyroid PV and atrial cardiomyocytes had shorter action potential duration. Hyperthyroid PV cardiomyocytes had faster beating rates (1.82 ± 0.13 Hz vs. 1.03 ± 0.15 Hz, p < 0.005) and a higher incidence of delayed afterdepolarization (beating: 92% vs. 6%, p < 0.0001; non-beating: 45% vs. 3%, p < 0.005). However, only hyperthyroid PV beating cardiomyocytes had a higher incidence of early afterdepolarization (46% vs. 0%, p < 0.0001). The ionic current experiments showed that hyperthyroid PV beating cardiomyocytes had larger densities of overall slow inward (2.72 ± 0.21 pA/pF vs. 2.07 ± 0.19 pA/pF, p < 0.05), overall transient outward (1.39 ± 0.21 pA/pF vs. 0.48 ± 0.08 pA/pF, p < 0.001) and steady state outward currents (0.78 ± 0.06 pA/pF vs. 0.58 ± 0.04 pA/pF, p < 0.05) on depolarization and larger transient inward (0.021 ± 0.004 pA/pF vs. 0.005 ± 0.001 pA/pF, p < 0.001) on repolarization. By contrast, the hyperthyroid PV non-beating cardiomyocytes had larger densities of overall transient outward (1.01 ± 0.14 pA/pF vs. 0.37 ± 0.07 pA/pF, p < 0.001), steady state outward (0.61 ± 0.06 pA/pF vs. 0.44 ± 0.04 pA/pF, p < 0.05) and transient inward currents (0.011 ± 0.002 pA/pF vs. 0.003 ± 0.001 pA/pF, p < 0.05).: ConclusionsThyroid hormone changes the electrophysiological activity of the PV cardiomyocytes. Increased automaticity and enhanced triggered activity may increase the arrhythmogenic activity of PVs in hyperthyroidism. [Copyright &y& Elsevier]