Effects of ANP on pulmonary vein electrophysiology, Ca2+ homeostasis and adrenergic arrhythmogenesis via PKA.

Atrial fibrillation (AF) is the most common form of arrhythmia and increases the risk of stroke and heart failure (HF). Pulmonary veins (PVs) are important sources of triggers that generate AF, and calcium (Ca2+ ) overload participates in PV arrhythmogenesis. Neurohormonal activation is an important cause of AF. Higher atrial natriuretic peptide (ANP) level predicts paroxysmal AF occurrence in HF patients. However, it is not clear if ANP directly modulates electrophysiological characteristics and Ca2+ homeostasis in the PVs. Conventional microelectrodes, whole-cell patch-clamp, and the Fluo-3 fluorimetric ratio technique were performed using isolated rabbit PV preparations or single isolated PV cardiomyocytes before and after ANP administration. We found that ANP (1, 10, and 100 nmol/L) concentration-dependently decreased spontaneous activity in PV preparations. ANP (100 nmol/L) decreased isoproterenol (1 µmol/L)-induced PV spontaneous activity and burst firing. AP811 (100 nmol/L, NPR-C agonist), H89 (1µmol/L, PKA inhibitor) decreased isoproterenol-induced PV spontaneous activity or burst firing, but successive administration of ANP had no further effect on PV activity. KT5823 (1 µmol/L, PKG inhibitor) decreased isoproterenol-induced PV spontaneous activity but did not change isoproterenol-induced PV burst firing, whereas successive administration of ANP did not change isoproterenol-induced PV burst firing. ANP decreased intracellular Ca2+ transient and sarcoplasmic reticulum Ca2+ content in single PV cardiomyocytes. ANP decreased the late sodium current, L-type Ca2+ current, but did not change nickel-sensitive Na+ -Ca2+ exchanger current in single PV cardiomyocytes. In conclusion, ANP directly regulates PV electrophysiological characteristics and Ca2+ homeostasis and attenuates isoproterenol-induced arrhythmogenesis through NPR-C/cAMP/PKA signal pathway.

Angiotensin 1-7 modulates electrophysiological characteristics and calcium homoeostasis in pulmonary veins cardiomyocytes via MAS/PI3K/eNOS signalling pathway.

BACKGROUND: Atrial fibrillation (AF) is the most common sustained arrhythmia, and pulmonary veins (PVs) play a critical role in triggering AF. Angiotensin (Ang)-(1-7) regulates calcium (Ca2+ ) homoeostasis and also plays a critical role in cardiovascular pathophysiology. However, the role of Ang-(1-7) in PV arrhythmogenesis remains unclear. MATERIALS AND METHODS: Conventional microelectrodes, whole-cell patch-clamp and the fluo-3 fluorimetric ratio technique were used to record ionic currents and intracellular Ca2+ in isolated rabbit PV preparations and in single isolated PV cardiomyocytes, before and after administration of Ang-(1-7). RESULTS: Ang (1-7) concentration dependently (0.1, 1, 10 and 100 nmol/L) decreased PV spontaneous electrical activity. Ang-(1-7) (100 nmol/L) decreased the late sodium (Na+ ), L-type Ca2+ and Na+ -Ca2+ exchanger currents, but did not affect the voltage-dependent Na+ current in PV cardiomyocytes. In addition, Ang-(1-7) decreased intracellular Ca2+ transient and sarcoplasmic reticulum Ca2+ content in PV cardiomyocytes. A779 (a Mas receptor blocker, 3 µmol/L), L-NAME (a NO synthesis inhibitor, 100 µmol/L) or wortmannin (a specific PI3K inhibitor, 10 nmol/L) attenuated the effects of Ang-(1-7) (100 nmol/L) on PV spontaneous electric activity. CONCLUSION: Ang-(1-7) regulates PV electrophysiological characteristics and Ca2+ homoeostasis via Mas/PI3K/eNOS signalling pathway.

Discrepant effects of heart failure on electrophysiological property in right ventricular outflow tract and left ventricular outflow tract cardiomyocytes.

Ventricular arrhythmias commonly arise from the right (RVOT) and left ventricular outflow tracts (LVOT) in patients without structural heart disease. Heart failure (HF) significantly increases the risk of ventricular arrhythmias. The regional differences and how HF affects the electrophysiological characteristics of RVOT and LVOT cardiomyocytes remain unclear. The whole-cell patch-clamp technique was used to investigate the action potentials and ionic currents in isolated single RVOT and LVOT cardiomyocytes from control rabbits and rabbits with HF induced by rapid ventricular pacing. Comparison with control LVOT cardiomyocytes showed that control RVOT cardiomyocytes have a shorter action potential duration (APD), smaller late Na+ currents (INa-late), larger transient outward (Ito) and larger delayed rectifier K+ currents (IKr-tail), but had similar L-type Ca2+ currents (ICa-L) and Na+/Ca2+ exchanger (NCX) current. HF increased APD, INa-late and NCX, but decreased ICa-L and Ito in RVOT cardiomyocytes. In contrast with this, HF decreased APD and ICa-L, but increased Ito and IKr-tail in LVOT cardiomyocytes. In conclusion, RVOT and LVOT cardiomyocytes had distinctive electrophysiological characteristics. HF differentially modulates action potential morphology and ionic currents in RVOT and LVOT cardiomyocytes.

2016 Guidelines of the Taiwan Heart Rhythm Society and the Taiwan Society of Cardiology for the management of atrial fibrillation.

Atrial fibrillation (AF) is the most common sustained arrhythmia. Both the incidence and prevalence of AF are increasing, and the burden of AF is becoming huge. Many innovative advances have emerged in the past decade for the diagnosis and management of AF, including a new scoring system for the prediction of stroke and bleeding events, the introduction of non-vitamin K antagonist oral anticoagulants and their special benefits in Asians, new rhythm- and rate-control concepts, optimal endpoints of rate control, upstream therapy, life-style modification to prevent AF recurrence, and new ablation techniques. The Taiwan Heart Rhythm Society and the Taiwan Society of Cardiology aimed to update the information and have appointed a jointed writing committee for new AF guidelines. The writing committee members comprehensively reviewed and summarized the literature, and completed the 2016 Guidelines of the Taiwan Heart Rhythm Society and the Taiwan Society of Cardiology for the Management of Atrial Fibrillation. This guideline presents the details of the updated recommendations, along with their background and rationale, focusing on data unique for Asians. The guidelines are not mandatory, and members of the writing committee fully realize that treatment of AF should be individualized. The physician's decision remains most important in AF management.

Adenosine monophosphate-regulated protein kinase inhibition modulates electrophysiological characteristics and calcium homeostasis of rabbit right ventricular outflow tract.

BACKGROUND: Metabolic stress predisposes to ventricular arrhythmias and sudden cardiac death. Right ventricular outflow tract (RVOT) is the common origin of ventricular arrhythmias. Adenosine monophosphate-regulated protein kinase (AMPK) activation is an important compensatory mechanism for cardiac remodeling during metabolic stress. OBJECTIVES: The purpose of this study was to access whether AMPK inhibition would modulate RVOT electrophysiology, calcium (Ca2+ ) regulation, and RVOT arrhythmogenesis or not. METHODS: Conventional microelectrodes were used to record electrical activity before and after compound C (10 µM, an AMPK inhibitor) in isoproterenol (1 µM)-treated rabbit RVOT tissue preparations under electrical pacing. Whole-cell patch-clamp and confocal microscopic examinations were performed in baseline and compound C-treated rabbit RVOT cardiomyocytes to investigate ionic currents and intracellular Ca2+ transients in isolated rabbit RVOT cardiomyocytes. RESULTS: Compound C decreased RVOT contractility, and reversed isoproterenol increased RVOT contractility. Compound C decreased the incidence, rate, and duration of isoproterenol-induced RVOT burst firing under rapid pacing. Compared to baseline, compound C-treated RVOT cardiomyocytes had a longer action potential duration, smaller intracellular Ca2+ transients, late sodium (Na+ ), peak L-type Ca2+ current density, Na+ -Ca2+ exchanger, transient outward potassium (K+ ) current, and rapid and slow delayed rectifier K+ currents. CONCLUSION: AMPK inhibition modulates RVOT electrophysiological characteristics and Ca2+ homeostasis, contributing to lower RVOT arrhythmogenic activity. Accordingly, AMPK inhibition might potentially reduce ventricular tachyarrhythmias.

Apelin regulates the electrophysiological characteristics of atrial myocytes.

BACKGROUND: Apelin, a potential agent for treating heart failure, has various ionic effects on ventricular myocytes. However, the effects of apelin on the atrium are not clear. The purpose of this study was to investigate the acute effects of apelin on the electrophysiological characteristics of atrial myocytes. METHOD: Whole-cell patch-clamp techniques were used to investigate the action potential (AP) and ionic currents in isolated rabbit left atrial (LA) myocytes before and after the administration of apelin. RESULT: Apelin reduced LA AP duration measured at 90%, 50% and 20% repolarization of the amplitude by 11 ± 3%, 24 ± 5%, 30 ± 7% at 1 nM (n = 11), and by 14 ± 4%, 36 ± 6% and 45 ± 5% at 10 nM (n = 11), but not at 0·1 nM. Apeline (0·1, 1, 10 nM) did not change the amplitude, or resting membrane potential in LA myocytes. Apelin (1 nM) increased sodium currents, ultra-rapid potassium currents and the reverse mode of sodium-calcium exchanger currents, but decreased late sodium currents and L-type calcium currents and did not change transient outward currents or inward rectifier potassium currents in LA myocytes. CONCLUSIONS: Apelin significantly changed the atrial electrophysiology with a shortening of AP duration, which may be caused by its effects on multiple ionic currents.

Effects of ivabradine on the pulmonary vein electrical activity and modulation of pacemaker currents and calcium homeostasis.

INTRODUCTION: Ivabradine is a novel heart rate decreasing agent with selective and specific antagonist effects on the pacemaker current (I(f)). The aim of this study was to investigate the pharmacological effects of ivabradine on the pulmonary vein (PV) cardiomyocytes. METHODS AND RESULTS: Whole-cell patch-clamp techniques and the indo-1 fluorimetric ratio technique were used to investigate the characteristics of the I(f) and intracellular calcium (Ca(2+)(i)) in single isolated rabbit PV cardiomyocytes with pacemaker activity before and after an ivabradine administration (0.3, 3, 10, and 30 µM). Ivabradine (0.3, 3, 10, and 30 µM) concentration dependently decreased the spontaneous activity by 6 ± 3%, 32 ± 6%, 49 ± 5%, and 85 ± 4%, and decreased the I(f) by 35 ± 8%, 47 ± 9%, 62 ± 5%, and 65 ± 7%, respectively, in PV cardiomyocytes. The decreased extent of the PV beating rate or I(f) by the different concentrations of ivabradine correlated well with the baseline PV beating rates. The IC(50) of the spontaneous activity and I(f) induced by ivabradine were 9.5 and 3.5 µM, respectively. Moreover, ivabradine (30 µM, but not 3 µM) decreased the Ca(2+)(i) transient in the PV cardiomyocytes and ivabradine (30 µM) decreased the L-type calcium current in the PV cardiomyocytes. CONCLUSION: Ivabradine decreased the I(f)s and Ca(2+)(i) transient in the PV cardiomyocytes, which may contribute to its inhibitory effects on the PV spontaneous activity.

A novel finding of the atrial substrate properties and long-term results of catheter ablation in chronic atrial fibrillation patients with left atrial spontaneous echo contrast.

BACKGROUND: The atrial substrate in chronic atrial fibrillation (AF) patients with a left atrial spontaneous echo contrast (LASEC) has not been previously reported. The aim of this study was to investigate the atrial substrate properties and long-term follow-up results in the patients who received catheter ablation of chronic AF. METHODS: Of 36 consecutive patients with chronic AF who received a stepwise ablation approach, 18 patients with an LASEC (group I) were compared with 18 age-gender-left atrial volume matched patients without an LASEC (group II). The atrial substrate properties including the weighted peak-to-peak voltage, total activation time during sinus rhythm (SR), dominant frequency (DF), and complex fractionated electrograms (CFEs) during AF in the bi-atria were evaluated. RESULT: The left atrial weighted bipolar peak-to-peak voltage (1.0 ± 0.6 vs 1.6 ± 0.7 mV, P = 0.04), total activation time (119 ± 20 vs 103 ± 13 ms, P < 0.001) and DF (7.3 ± 1.3 vs 6.6 ± 0.7 Hz, P < 0.001) differed between group I and group II, respectively. Those parameters did not differ in the right atrium. The bi-atrial CFEs (left atrium: 89 ± 24 vs 92 ± 25, P = 0.8; right atrium: 92 ± 25 vs 102 ± 3, P = 0.9) did not differ between group I and group II, respectively. After a mean follow-up of 30 ± 13 month, there were significant differences in the antiarrhythmic drugs (1.1 ± 0.3 vs 0.7 ± 0.5, P = 0.02) needed after ablation, and recurrence as persistent AF (92% vs 50%, P = 0.03) between group I and group II, respectively. After multiple procedures, there were more group II patients that remained in SR, when compared with group I (78% vs 44%, P = 0.04). CONCLUSION: There was a poorer atrial substrate, lesser SR maintenance after catheter ablation and need for more antiarrhythmic drugs in the chronic AF patients with an LASEC when compared with those without an LASEC.

Long-term follow-up in patients with arrhythmogenic right ventricular cardiomyopathy.

INTRODUCTION: Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a major cause of sudden cardiac death due to tachyarrhythmias. The purpose of this study was to investigate the long-term prognosis in patients with ARVC and the incidence of rapid ventricular arrhythmias during follow-up. METHODS: Thirty ARVC patients (19 male, 63.3%, mean age 48 ± 15 years) fulfilling modified Task Force criteria 2010 were included. Of them, 13 patients (43.3%) received implantable cardioverter-defibrillator (ICD) implantation. Rapid ventricular arrhythmia was defined as electrical storm or the occurrence of ventricular tachycardia (VT) or ventricular fibrillation (VF) with a cycle length of 240 ms or less that necessitate shock delivery to 2 or more times within a 24-hour period. RESULTS: With a mean follow-up of 68 ± 10 months, 6 patients (20%) with ICD implantation had recurrent rapid VT/VF. One (3.3%) of them died of multiple shocks and SCD, and 5 (16.7%) had multiple ICD therapies due to VT/VF and electrical storm. The interval between the diagnosis of ARVC and occurrence of rapid VT/VF was 13.4 ± 4.9 months. Most (5/6, 83.3%) events of recurrent rapid VT/VF occurred within 2 years. Ablated patients who did not receive an ICD implant were totally free of rapid VT/VF. CONCLUSIONS: For patients with ARVC, long-term prognosis is favorable. During a long-term follow-up, patients meeting the criteria for ICD implantation have a higher rate of rapid and potentially life-threatening arrhythmias. However, early and clustered recurrence of rapid VT/VF in patients with an ICD is common, whereas late occurrence of rapid VT/VF is very rare.

Long-term outcome of catheter ablation in patients with atrial fibrillation originating from the superior vena cava.

UNLABELLED: Long-Term Outcome of SVC AF Ablation. INTRODUCTION: Data of the long-term clinical outcome after superior vena cava (SVC) isolation are limited. We aimed to evaluate the long-term outcome in patients with atrial fibrillation (AF) who had triggers originating from the SVC and received catheter ablation of AF. METHODS AND RESULTS: The study consisted of 68 patients (age 56 ± 12 years old, 32 males) who underwent the ablation procedure for drug-refractory, symptomatic paroxysmal AF originating from the SVC since 1999. Group 1 consisted of 37 patients with AF initiated from the SVC only, and group 2 consisted of 31 patients with both SVC and pulmonary vein (PV) triggers. During a follow-up period of 88 ± 50 months, the AF recurrence rate was 35.3% after a single procedure. The freedom-from-AF rates were 85.3% at 1 year and 73.3% at 5 years. In the baseline study, group 2 had larger left atrium (38 ± 4 mm vs 36 ± 5 mm, P = 0.04), left ventricle (50 ± 5 mm vs 46 ± 5 mm, P = 0.003), and PV diameters. Kaplan-Meier survival analysis showed a higher AF recurrence rate in group 2 compared to that in group 1 (P = 0.012). The independent predictor of an AF recurrence was a larger SVC diameter (P = 0.02, HR 1.4, 95% CI 1.1-1.8). CONCLUSION: Among the patients with paroxysmal AF originating from the SVC, 73% remained free of AF for 5 years after a single catheter ablation procedure. Superior vena cava isolation without PV isolation is an acceptable therapeutic strategy in those patients with AF originating from the SVC only. The SVC diameter was an independent predictor of AF recurrence. (J Cardiovasc Electrophysiol, Vol. 23, pp. 955-961, September 2012).

ATX-II-induced pulmonary vein arrhythmogenesis related to atrial fibrillation and long QT syndrome.

BACKGROUND: Long QT syndrome (LQTS) is associated with a high incidence of atrial fibrillation (AF), but the underlying mechanisms are unclear. Pulmonary veins (PVs) play a critical role in AF genesis. Type 3 LQTS increases late sodium current (I(Na,L) ), which may increase PV arrhythmogenesis and AF. Therefore, this study examines PV arrhythmogenesis in anemonia sulcata toxin II (ATX-II)-induced type 3 LQTS and evaluates whether the I(Na,L) inhibitor ranolazine can suppress PV arrhythmogenesis. MATERIALS AND METHODS: Conventional microelectrodes were used to record the action potentials (AP) and contractility in isolated rabbit PV specimens before and after ATX-II administration with or without ranolazine. RESULTS: Anemonia sulcata toxin II (100 nM) increased the PV spontaneous rates from 2·0 ± 0·1 to 2·9 ± 0·2 Hz (n = 7), induced PV burst firing (100%) with the genesis of early afterdepolarization (EAD) (86%) and prolonged the AP duration. Ranolazine (0·1, 1 and 10 µM) dose dependently reduced the PV spontaneous rates from 2·5 ± 0·2 to 2·3 ± 0·2 Hz, 1·9 ± 0·2 and 1·5 ± 0·3 Hz (P < 0·05) and decreased the diastolic tension by 40 ± 19%, 87 ± 26% and 113 ± 28%. In the presence of ranolazine (10 µM), ATX-II (100 nM) further increased the AP duration. However, ATX-II neither increased the PV spontaneous rates (1·6 ± 0·1 vs. 1·7 ± 0·2 Hz, n = 7) nor induced PV burst firing or EAD. Moreover, ranolazine (10 µM) reduced ATX-II-induced PV acceleration and EAD. CONCLUSIONS: The I(Na,L) enhancer ATX-II can increase PV arrhythmogenesis, which can be attenuated or blocked by ranolazine. This suggests that AF may be related to type 3 LQTS through increased I(Na,L) .

Hypoxia and reoxygenation modulate the arrhythmogenic activity of the pulmonary vein and atrium.

Ischaemia and reperfusion contribute to the genesis of AF (atrial fibrillation). PVs (pulmonary veins) and the atria are important foci for AF initiation and maintenance. However, the effect of ischaemia and reperfusion on PVs and the atria has not yet been fully elucidated. In the present study, conventional microelectrodes were used to record the APs (action potentials) in isolated rabbit PV, LA (left atrium) and RA (right atrium) specimens during hypoxia and reoxygenation, and pharmacological interventions. Hypoxia reduced the PV beating rates from 1.8±0.1 to 1.3±0.2 and 0.8±0.1 Hz at 30 and 60 min respectively (n=8, P<0.005), and induced EAD (early after depolarization) in three (37.5%) of the PVs and DAD (delayed after depolarization) in one (12.5%) of the PVs. Reoxygenation increased the PV spontaneous rate to 1.4±0.2 Hz (P<0.05) and induced PV burst firings (3.5±0.1 Hz, P<0.001) in six (75%) of the PVs. Hypoxia shortened the AP duration in the LA and PVs, but not in the RA. Pretreatment with glibenclamide attenuated hypoxia-induced decreases in the PV spontaneous activity and the shortening of the LA and PV AP duration. Similar to those in hypoxia, the K(ATP) (ATP-sensitive potassium) channel opener pinacidil (30 µM) decreased PV spontaneous activity and shortened the AP duration. Pretreatment with 5 mM N-MPG [N-(mercaptopropionyl)glycine; a hydroxyl (•OH) free-radical scavenger] or 300 µM chloramphenicol [a cytochrome P450 inhibitor that reduces ROS (reactive oxygen species)] attenuated the rate changes induced by hypoxia and reoxygenation, and also decreased the burst firing incidence. In conclusion, hypoxia and reoxygenation significantly increased PV arrhythmogenesis and induced different electrophysiological responses in the RA and LA, which may play a role in the pathophysiology of AF.

Hydralazine-induced promoter demethylation enhances sarcoplasmic reticulum Ca2+ -ATPase and calcium homeostasis in cardiac myocytes.

Sarcoplasmic reticulum (SR) Ca(2+)-ATPase (SERCA2a) plays an essential role in Ca(2+) homeostasis and cardiac functions. The promoter region of SERCA2a has a high content of CpG islands; thus, epigenetic modification by inhibiting methylation can enhance SERCA2a expression in cardiomyocytes. Hydralazine, a drug frequently used in heart failure, is a potential DNA methylation inhibitor. We evaluated whether hydralazine can modulate Ca(2+) handling through an increase in SERCA2a expression via regulating methylation. We used indo-1 fluorescence, real-time RT-PCR, immunoblotting, and methylation-specific PCR to investigate intracellular Ca(2+), the expressions of RNA and protein, and methylation of SERCA2a in HL-1 cardiomyocytes with and without (control) the administration of hydralazine (1, 10, and 30 µM) for 72 h. Hydralazine (10 and 30 µM) increased the intracellular Ca(2+) transients and SR Ca(2+) contents. Hydralazine (10 and 30 µM) decreased methylation in the SERCA2a promoter region and increased the RNA and protein expressions of SERCA2a. Additionally, hydralazine (10 and 30 µM) decreased the expression of DNA methyltransferase 1. Moreover, treatment with hydralazine in isoproterenol-induced heart failure rats decreased the promoter methylation of SERCA2a and increased SERCA2a RNA expression. In conclusion, hydralazine-induced promoter demethylation may improve cardiac function through increasing SERCA2a and modulating calcium homeostasis in cardiomyocytes.

Discrepant electrophysiological characteristics and calcium homeostasis of left atrial anterior and posterior myocytes.

The left atrial (LA) posterior wall has been demonstrated to have regional electrophysiological differences with a higher arrhythmogenic potential leading to atrial fibrillation (AF). However, the ionic characteristics and calcium regulation in the LA anterior and posterior myocytes have not been fully elucidated. The purpose of this study was to investigate the electrical characteristics of the LA anterior and posterior myocytes. Whole-cell patch-clamp techniques and the indo-1 fluorimetric ratio technique were used to investigate the characteristics of the ionic currents, action potentials, and intracellular calcium in single isolated rabbit myocytes in the LA anterior and posterior walls. The expression of the Na(+)-Ca(2+) exchanger (NCX) and ryanodine receptor (RyR) were evaluated by a Western blot. The LA posterior myocytes (n = 15) had a higher incidence (53 vs. 19%, P < 0.05) of delayed afterdepolarizations than the LA anterior myocytes (n = 16). The LA posterior myocytes had larger sodium currents and late sodium currents, but smaller inward rectifier potassium currents than the LA anterior myocytes. The LA posterior myocytes had larger intracellular Ca(2+) transient and sarcoplasmic reticulum Ca(2+) contents as compared with the LA anterior myocytes. However, the NCX currents in the LA posterior myocytes were smaller than those in the LA anterior myocytes. The LA posterior myocytes had a smaller protein expression of NCX, but a larger protein expression of RyR than the LA anterior myocytes. In conclusion, LA posterior myocytes contain a high arrhythmogenic potential and distinctive electrophysiological characteristics, which may contribute to the pathophysiology of AF.

Heat-stress responses modulate beta-adrenergic agonist and angiotensin II effects on the arrhythmogenesis of pulmonary vein cardiomyocytes.

INTRODUCTION: Heat stress-induced responses reduce the occurrence of atrial fibrillation (AF). Pulmonary vein (PV) cardiomyocytes with pacemaker activity play a critical role in the pathophysiology of AF. In this study, we examined whether heat-stress responses alter the electrophysiological characteristics of PV cardiomyocytes and protect the PV against angiotensin II- or isoproterenol-induced arrhythmogenesis. METHODS AND RESULTS: We used whole-cell patch clamp techniques to investigate the spontaneous activity and ionic currents in single isolated rabbit PV pacemaker cardiomyocytes with or without (control) exposure to heat stress (43°C, 15 minutes) 5 ± 1 hours before the experiments. Compared to control cardiomyocytes, heat-stressed PV cardiomyocytes had slower beating rates. Heat-stressed PV cardiomyocytes had larger L-type calcium currents, transient outward currents, smaller inward rectifier potassium currents, but similar sodium-calcium exchanger currents. Additionally, heat-stressed PV cardiomyocytes had a lower incidence of pacemaker currents than control PV cardiomyocytes. Moreover, isoproterenol increased the beating rate of control cardiomyocytes but not heat-stressed PV cardiomyocytes. Similarly, angiotensin II also increased the beating rate of control cardiomyocytes, but not heat-stressed PV cardiomyocytes, in association with decreased expression of the angiotensin II type 1 receptor. CONCLUSION: Heat-stress responses altered the electrophysiological characteristics of PV cardiomyocytes and attenuated the effects of isoproterenol and angiotensin II on PV arrhythmogenesis, which may play a role in the protective potential of heat-stress responses.

Vascular endothelial growth factor modulates pulmonary vein arrhythmogenesis via vascular endothelial growth factor receptor 1/NOS pathway.

Atrial fibrillation (AF) is a common form of arrhythmia with serious public health impacts, but its underlying mechanisms are not yet fully understood. Vascular endothelial growth factor (VEGF) is highly expressed in the atrium of patients with AF, but whether VEGF affects AF pathogenesis remains unclear. Pulmonary veins (PVs) are important sources for the genesis of atrial tachycardia or AF. Therefore, this study assessed the effects of VEGF on PV electrophysiological properties and evaluated its underlying mechanisms. Conventional microelectrodes and whole-cell patch clamps were performed using isolated rabbit PV preparations or single isolated PV cardiomyocytes before and after VEGF or VEGF receptor (VEGFR), Akt, NOS inhibitor administration. We found that VEGF (0.1, 1, and 10 ng/mL) reduced the PV beating rate in a dose-dependent manner. Furthermore, VEGF (10 ng/mL) reduced late diastolic depolarization and diastolic tension. Isoproterenol increased PV beating and burst firing, which was attenuated by VEGF (1 ng/mL). In the presence of VEGFR-1 inhibition (ZM306416 at 10 µM) and L-NAME (100 µM), VEGF (1 ng/mL) did not alter PV spontaneous activity. In isolated PV cardiomyocytes, VEGF (1 ng/mL) decreased L-type calcium, sodium/calcium exchanger, and late sodium currents. In conclusion, we found that VEGF reduces PV arrhythmogenesis by modulating sodium/calcium homeostasis through VEGFR-1/NOS signaling pathway.

Fibroblast Growth Factor 1 Reduces Pulmonary Vein and Atrium Arrhythmogenesis via Modification of Oxidative Stress and Sodium/Calcium Homeostasis.

RATIONALE: Atrial fibrillation is a critical health burden. Targeting calcium (Ca2+) dysregulation and oxidative stress are potential upstream therapeutic strategies. Fibroblast growth factor (FGF) 1 can modulate Ca2+ homeostasis and has antioxidant activity. The aim of this study was to investigate whether FGF1 has anti-arrhythmic potential through modulating Ca2+ homeostasis and antioxidant activity of pulmonary vein (PV) and left atrium (LA) myocytes. METHODS: Patch clamp, western blotting, confocal microscopy, cellular and mitochondrial oxidative stress studies were performed in isolated rabbit PV and LA myocytes treated with or without FGF1 (1 and 10 ng/mL). Conventional microelectrodes were used to record electrical activity in isolated rabbit PV and LA tissue preparations with and without FGF1 (3 µg/kg, i.v.). RESULTS: FGF1-treated rabbits had a slower heart rate than that observed in controls. PV and LA tissues in FGF1-treated rabbits had slower beating rates and longer action potential duration than those observed in controls. Isoproterenol (1 µM)-treated PV and LA tissues in the FGF1-treated rabbits showed less changes in the increased beating rate and a lower incidence of tachypacing (20 Hz)-induced burst firing than those observed in controls. FGF1 (10 ng/mL)-treated PV and LA myocytes had less oxidative stress and Ca2+ transient than those observed in controls. Compared to controls, FGF1 (10 ng/mL) decreased INa-L in PV myocytes and lowered I to, I Kr-tail in LA myocytes. Protein kinase C (PKC)ε inhibition abolished the effects of FGF1 on the ionic currents of LA and PV myocytes. CONCLUSION: FGF1 changes PV and LA electrophysiological characteristics possibly via modulating oxidative stress, Na+/Ca2+ homeostasis, and the PKCε pathway.

Heat stress responses modulate calcium regulations and electrophysiological characteristics in atrial myocytes.

Heat stress-induced responses change the ionic currents and calcium homeostasis. However, the molecular insights into the heat stress responses on calcium homeostasis remain unclear. The purposes of this study were to examine the mechanisms of heat stress responses on calcium handling and electrophysiological characteristics in atrial myocytes. We used indo-1 fluorimetric ratio technique and whole-cell patch clamp to investigate the intracellular calcium, action potentials, and ionic currents in isolated rabbit single atrial cardiomyocytes with or without (control) exposure to heat stress (43 degrees C, 15 min) 5+/-1 h before experiments. The expressions of sarcoplasmic reticulum ATPase (SERCA2a), and Na(+)-Ca(2+) exchanger (NCX) in the control and heat stress-treated atrial myocytes were evaluated by Western blot and real-time PCR. As compared with control myocytes, the heat stress-treated myocytes had larger sarcoplasmic reticulum calcium content and larger intracellular calcium transient with a shorter decay portion. Heat stress-treated myocytes also had larger L-type calcium currents, transient outward potassium currents, but smaller NCX currents. Heat stress responses increased the protein expressions, SERCA2a, NCX, and heat shock protein. However, heat stress responses did not change the RNA expression of SERCA2a and NCX. In conclusion, heat stress responses change calcium handling through protein but not RNA regulation.

Tumor necrosis factor-alpha decreases sarcoplasmic reticulum Ca2+-ATPase expressions via the promoter methylation in cardiomyocytes.

OBJECTIVES: Sarcoplasmic reticulum Ca-ATPases (SERCA2a) plays an essential role in the Ca homeostasis and cardiac functions. Tumor necrosis factor-alpha (TNF-alpha) decreases the SERCA2a, which may underlie cardiac dysfunction during sepsis and heart failure. Because the promoter region of SERCA2a contains CpG islands, gene methylation should be critical in regulating SERCA2a. The present study was to evaluate whether TNF-alpha can modulate SERCA2a via enhancing methylation and to investigate the underlying mechanisms. DESIGN: Controlled laboratory experiment. SETTING: University research laboratory. SUBJECTS: HL-1 cardiomyocytes. INTERVENTIONS: TNF-alpha (1-50 ng/mL) was administered in HL-1 cardiomyocytes with and without co-administration of an NF-kappaB inhibitor (SN-50, 50 microg/mL), antioxidant agents (ascorbic acid, 100 microM, or coenzyme Q10, 10 microM), or methylation inhibitor (5-aza-2'-deoxycytidine, 0.1, 1 microM). MEASUREMENTS AND MAIN RESULTS: TNF-alpha (50 ng/mL) decreased the SERCA2a RNA and protein by quantitative polymerase chain reaction and immunoblot. Furthermore, TNF-alpha (50 ng/mL) increased the methylation in the SERCA2a promoter region, which was not influenced by the co-administration of SN-50, ascorbic acid, or coenzyme Q10, but was attenuated by 5-aza-2'-deoxycytidine (0.1 microM). Additionally, TNF-alpha (50 ng/mL) increased the expression of DNA methyltransferase 1. CONCLUSIONS: TNF-alpha increased DNA methyltransferase levels, thus enhancing the methylation in the SERCA2a promoter region with a result of reducing SERCA2a. These findings suggest that inhibition of hypermethylation may be a novel treatment strategy for cardiac dysfunction.

Oxidative stress on pulmonary vein and left atrium arrhythmogenesis.

BACKGROUND: Oxidative stress and pulmonary veins (PVs) play critical roles in the pathophysiology of atrial fibrillation. The purpose of the present study was to investigate whether oxidative stress and antioxidant agents can change the electrophysiological characteristics of the left atrium (LA) and PVs. METHODS AND RESULTS: Conventional microelectrodes were used to record the action potentials (APs) in isolated rabbit PV and LA specimens before and after H(2)O(2) administration with or without ascorbic acid or N-mercaptopropionyl-glycine (N-MPG, a free radical .OH scavenger). H(2)O(2) (0.02 and 0.2 mmol/L) decreased the PV spontaneous rates from 2.0+/-0.1 Hz to 1.6+/-0.1 Hz, and 1.7+/-0.1 Hz (n=10, P<0.05), but H(2)O(2) (2 mmol/L) increased PV spontaneous rates from 2.0+/-0.1 Hz to 2.8+/-0.2 Hz. H(2)O(2) easily induced PV burst firing and early afterdepolarizations, but not in the LA. H(2)O(2) shortened the AP duration and increased the contractile force to a greater extent in the LA than in PVs. In addition, the H(2)O(2)-induced PV burst firing and increasing spontaneous rates were suppressed or attenuated by pretreatment with ascorbic acid (1 mmol/L) or N-MPG (10 mmol/L). CONCLUSIONS: H(2)O(2) significantly changed the electrophysiological characteristics of PV and LA through activation of free radicals and may facilitate the occurrence of atrial fibrillation.