Isolated Rabbit Pulmonary Veins Research Articles

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

RationaleAtrial 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.MethodsPatch 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.).ResultsFGF1-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 Ito, IKr−tail in LA myocytes. Protein kinase C (PKC)ε inhibition abolished the effects of FGF1 on the ionic currents of LA and PV myocytes.ConclusionFGF1 changes PV and LA electrophysiological characteristics possibly via modulating oxidative stress, Na+/Ca2+ homeostasis, and the PKCε pathway.

Ceramide modulates electrophysiological characteristics and oxidative stress of pulmonary vein cardiomyocytes.

Ceramide is involved in regulating metabolism and energy expenditure, and its abnormal myocardial accumulation may contribute to heart injury or lipotoxic cardiomyopathy. Whether ceramide can modulate the electrophysiology of pulmonary veins (PVs) remains unknown. We used conventional microelectrodes to measure the electrical activity of isolated rabbit PV tissue preparations before and after treatment with various concentrations of ceramide with or without H2 O2 (2mM), MitoQ, wortmannin or 740 YP. A whole-cell patch clamp and fluorescence imaging were used to record the ionic currents, calcium (Ca2+ ) transients, and intracellular reactive oxygen species (ROS) and sodium (Na+ ) in isolated single PV cardiomyocytes before and after ceramide (1μM) treatment. Ceramide (0.1, 0.3, 1 and 3μM) reduced the beating rate of PV tissues. Furthermore, ceramide (1μM) suppressed the 2mM H2 O2 -induced faster PV beating rate, triggered activities and burst firings, which were further reduced by MitoQ. In the presence of wortmannin, ceramide did not change the PV beating rate. The H2 O2 -induced faster PV beating rate could be counteracted by MitoQ or wortmannin with no additive effect from the ceramide. Ceramide inhibited pPI3K. Ceramide reduced Ca2+ transients, sarcoplasmic reticulum Ca2+ contents, L-type Ca2+ currents, Na+ currents, late Na+ currents, Na+ -hydrogen exchange currents, and intracellular ROS and Na+ in PV cardiomyocytes, but did not change Na+ -Ca2+ exchange currents. C2 ceramide may exert the distinctive electrophysiological effect of modulating PV activities, which may be affected by PI3K pathway-mediated oxidative stress, and might play a role in the pathogenesis of PV arrhythmogenesis.

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.

Effects of phosphodiesterase-1 inhibitor on pulmonary vein electrophysiology and arrhythmogenesis.

Phosphodiesterase (PDE) isoform inhibitors have mechanical and electrical effects on the heart. Inhibition of PDE-1 enzymes is a novel strategy for treating heart failure. However, the electrophysiological effects of PDE-1 inhibition on the heart remain unclear. This study explored the effects of PDE-1 inhibition using ITI-214 on electrical activity in the pulmonary vein (PV), the most common trigger of atrial fibrillation, and investigated the underlying ionic mechanisms. Conventional microelectrodes or whole-cell patch clamps were employed to study the effects of ITI-214 (0.1-10μM) on PV electrical activity, mechanical responses and ionic currents in isolated rabbit PV tissue specimens and isolated single PV cardiomyocytes. ITI-214 at 1μM and 10μM (but not 0.1μM) significantly reduced PV spontaneous beating rate (10±2% and 10±3%, respectively) and PV diastolic tension (11±3% and 17±3%, respectively). ITI-24 (1μM) significantly reduced late sodium current (INa-Late ), L-type calcium current (ICa-L ) and the reverse mode of the sodium-calcium exchanger (NCX), but it did not affect peak sodium currents. ITI-214 reduces PV spontaneous activity and PV diastolic tension by reducing INa-Late , ICa-L and NCX current. Considering its therapeutic potential in heart failure, targeting PDE-1 inhibition may provide a novel strategy for managing atrial arrhythmogenesis.

Klotho modulates electrical activity and calcium homeostasis in pulmonary vein cardiomyocytes via PI3K/Akt signalling.

Klotho, a potential antiageing protein has remarkable cardiovascular effects, which is lower in the patients with chronic kidney disease (CKD). Chronic kidney disease increases the risk of atrial fibrillation, majorly triggered by pulmonary vein (PV) arrhythmogenesis. This study investigated whether klotho protein can modulate PV electrical activity and the underlying potential mechanisms. A conventional microelectrode and whole-cell patch clamp were used to investigate the action potentials and ionic currents in isolated rabbit PV tissue preparations and single cardiomyocytes before and after klotho administration. Phosphoinositide 3-kinase (PI3K)/Akt signalling was studied using western blotting. Klotho significantly reduced PV spontaneous beating rates in PV tissue preparations at 1.0 and 3.0 ng/mL (but not at 0.1 and 0.3 ng/mL). In the presence of the Akt inhibitor (10 µM), klotho (1.0 and 3.0 ng/mL) did not change PV electrical activities. Klotho (1.0 ng/mL) significantly decreased the late sodium current (INa-Late) and L-type calcium current (ICa-L), similar to the Akt inhibitor (10 µM). Western blots demonstrated that klotho (1.0 ng/mL)-treated PV cardiomyocytes had less phosphorylation of Akt (Ser473) compared with klotho-untreated cardiomyocytes. Compared with control PVs, klotho at relatively lower concentrations (0.1 and 0.3 ng/mL) significantly reduced beating rates and decreased the amplitudes of delay afterdepolarizations in CKD PVs. Klotho modulated PV electrical activity by inhibiting PI3K/Akt signalling, which may provide a novel insight into CKD-induced arrhythmogenesis.

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 100nmol/L) concentration-dependently decreased spontaneous activity in PV preparations. ANP (100nmol/L) decreased isoproterenol (1μmol/L)-induced PV spontaneous activity and burst firing. AP811 (100nmol/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 (1μmol/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.

Factor Xa inhibitors differently modulate electrical activities in pulmonary veins and the sinoatrial node

Factor Xa inhibitors reduce stroke in patients with atrial fibrillation. Pulmonary veins (PVs) and the sinoatrial node (SAN) are crucial for genesis of atrial fibrillation. However, the electrophysiological effects of factor Xa inhibitors (edoxaban and rivaroxaban) on PVs and the SAN remain unclear. Conventional microelectrodes were used to record the action potential in isolated rabbit PVs and SAN preparations before and after administration of edoxaban (0.1, 0.3, and 1 μM) or rivaroxaban (0.01, 0.03, 0.1, and 0.3 μM). A whole-cell patch-clamp was used to record the late sodium current (INa-late) in isolated single rabbit PV cardiomyocytes. Edoxaban significantly reduced PV spontaneous beating rates at 0.3 and 1 μM (N = 6 rabbits, P < 0.05), and reduced SAN beating rates at 1 μM (N = 6, P < 0.05). Similarly, rivaroxaban reduced PV spontaneous beating rates at 0.1 and 0.3 μM (N = 7, P < 0.05), and reduced SAN beating rates at 0.3 μM (N = 6, P < 0.05). However, neither edoxaban (1 μM) nor rivaroxaban (0.3 μM) reduced PV spontaneous beating rates in the presence of 1 μM BMS200261 (an inhibitor of protease-activated receptors type 1, PAR1 inhibitor) or 10 μM ranolazine (an inhibitor of late sodium current, INa-late inhibitor). Edoxaban (0.3 and 1 μM) and rivaroxaban (0.1 and 0.3 μM) respectively decreased the INa-late by 47%, 47%, 36%, and 49% (n = 9 PV cardiomyocytes from 5 rabbits, P < 0.05). In conclusion, Factor Xa inhibitors reduce PV spontaneous activities and may modulate occurrence of atrial fibrillation by inhibiting PAR1 and reducing the INa-late in PVs.

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

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. 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). 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. Ang-(1-7) regulates PV electrophysiological characteristics and Ca2+ homoeostasis via Mas/PI3K/eNOS signalling pathway.

Mitochondrial dysfunction on sinoatrial node and pulmonary vein electrophysiological activities.

Atrial fibrillation (AF) is associated with mitochondrial dysfunction. Sinoatrial node (SAN) dysfunction increases arrhythmogenesis of pulmonary veins (PVs), which is the most important trigger of AF; however, it is not clear whether mitochondrial dysfunction differentially regulates electrical activity of SANs and PVs. In the present study, conventional microelectrodes were used to record the action potentials (APs) in isolated rabbit PVs, SANs, left atrium (LA) and right atrium (RA) before and after application of trifluorocarbonylcyanide phenylhydrazone (FCCP; a mitochondrial uncoupling agent) at 10, 100 and 300 nM. FCCP application at 100 and 300 nM decreased spontaneous rates in PVs and in SANs at 10, 100 and 300 nM. FCCP shortened the 20, 50 and 90% AP durations in the LA, and shortened only the 20% AP duration in the RA. FCCP caused a greater rate reduction in SANs than in PVs; however, in the presence of coenzyme-Q10 (10 µM), FCCP reduced the beating rate in PVs and SANs to a similar extent. In SAN-PV preparations with intact electrical connections, FCCP (100 nM) application shifted the SAN-PV electrical conduction into PV-SAN conduction in 5 (62.5%) of 8 preparations. In conclusion, mitochondrial dysfunction modulates PV and SAN electrical activities, which may contribute to atrial arrhythmogenesis.

Latrunculin B modulates electrophysiological characteristics and arrhythmogenesis in pulmonary vein cardiomyocytes.

AF (atrial fibrillation) is the most common sustained arrhythmia, and the PVs (pulmonary veins) play a critical role in triggering AF. Stretch causes structural remodelling, including cytoskeleton rearrangement, which may play a role in the genesis of AF. Lat-B (latrunculin B), an inhibitor of actin polymerization, is involved in Ca(2+) regulation. However, it is unclear whether Lat-B directly modulates the electrophysiological characteristics and Ca(2+) homoeostasis of the PVs. Conventional microelectrodes, whole-cell patch-clamp, and the fluo-3 fluorimetric ratio technique were used to record ionic currents and intracellular Ca(2+) within isolated rabbit PV preparations, or within isolated single PV cardiomyocytes, before and after administration of Lat-B (100 nM). Langendorff-perfused rabbit hearts were exposed to acute and continuous atrial stretch, and we studied PV electrical activity. Lat-B (100 nM) decreased the spontaneous electrical activity by 16±4% in PV preparations. Lat-B (100 nM) decreased the late Na(+) current, L-type Ca(2+) current, Na(+)/Ca(2+) exchanger current, and stretch-activated BKCa current, but did not affect the Na(+) current in PV cardiomyocytes. Lat-B reduced the transient outward K(+) current and ultra-rapid delayed rectifier K(+) current, but increased the delayed rectifier K(+) current in isolated PV cardiomyocytes. In addition, Lat-B (100 nM) decreased intracellular Ca(2+) transient and sarcoplasmic reticulum Ca(2+) content in PV cardiomyocytes. Moreover, Lat-B attenuated stretch-induced increased spontaneous electrical activity and trigger activity. The effects of Lat-B on the PV spontaneous electrical activity were attenuated in the presence of Y-27632 [10 μM, a ROCK (Rho-associated kinase) inhibitor] and cytochalasin D (10 μM, an actin polymerization inhibitor). In conclusion, Lat-B regulates PV electrophysiological characteristics and attenuates stretch-induced arrhythmogenesis.

Histone Deacetylase Inhibition Reduces Pulmonary Vein Arrhythmogenesis through Calcium Regulation

Pulmonary veins (PVs) play a critical role in the pathophysiology of atrial fibrillation (AF). Histone deacetylases (HDACs) are vital to calcium homeostasis and AF genesis. However, the electrophysiological effects of HDAC inhibition were unclear. This study evaluated whether HDAC inhibition can regulate PV electrical activity through calcium modulation.Whole-cell patch-clamp, confocal microscopic with fluorescence, and Western blot were used to evaluate electrophysiological characteristics and Ca2+ dynamics in isolated rabbit PV cardiomyocytes with and without MPT0E014 (a pan HDAC inhibitor), MS-275 (HDAC1 and 3 inhibitor), and MC-1568 (HDAC4 and 6 inhibitor) for 5~8h. Atrial electrical activity and induced-AF (rapid atrial pacing and acetylcholine infusion) were measured in rabbits with and without MPT0E014 (10mg/kg treated for 5hours) in vivo.MPT0E014 (1μM)-treated PV cardiomyocytes (n=12) had slower beating rates (2.1±0.2 vs. 2.8±0.1Hz, p<0.05) than control PV cardiomyocytes. However, control (n=11) and MPT0E014 (1μM)-treated (n=12) SAN cardiomyocytes had similar beating rates (3.2±0.2 vs. 2.9±0.3Hz). MS-275-treated PV cardiomyocytes (n=12, 2.3±0.2Hz), but not MC-1568-treated PV cardiomyocytes (n=14, 3.1±0.3Hz) had slower beating rates than control PV cardiomocytes. MPT0E014-treated PV cardiomyocytes (n=14) had a lower frequency (2.4±0.6 vs. 0.3±0.1 spark/mm/s, p<0.05) of Ca2+ sparks than control PV (n=17) cardiomyocytes. As compared to control, MPT0E014-treated PV cardiomyocytes had reduced Ca2+ transient amplitudes, sodium-calcium exchanger currents, and ryanodine receptor expressions. Moreover, MPT0E014-treated rabbits had less AF and shorter AF duration than control rabbits.In conclusions, HDAC inhibition reduced PV arrhythmogenesis and AF inducibility with modulation on calcium homeostasis.

Apamin modulates electrophysiological characteristics of the pulmonary vein and the Sinoatrial Node

Small-conductance calcium-activated potassium (SK) channels play an important role in atrial electrophysiology. Blocking SK channels prolongs action potential (AP) duration and attenuate electrical remodelling. The effects of SK blocking on the pulmonary vein (PV) and the sinoatrial node (SAN) remain unclear. Conventional microelectrodes were used to record the AP in SANs and in isolated rabbit PVs with and without denudation before and after treatment with apamin (1, 10 nM). Using the whole-cell patch-clamp technique, the SK current was investigated in isolated single PV and SAN myocytes. In nondenudated PVs (n = 6), apamin (1 and 10 nM) increased PV spontaneous activity (from 1.8 ± 0.2 Hz to 2.4 ± 0.2 Hz and 2.3 ± 0.2 Hz, P < 0.05) and increased the PV tension by 42 ± 14% and 37 ± 11%. Conversely, in denudated PVs (n = 6), apamin (1 and 10 nM) decreased spontaneous activity (from 2.2 ± 0.3 Hz to 1.9 ± 0.2 Hz and 1.8 ± 0.2 Hz, P < 0.05) and prolonged AP duration without changing PV tensions. Additionally, apamin (1 and 10 nM) decreased spontaneous activity (from 2.8 ± 0.1 Hz to 2.4 ± 0.1 Hz, and 2.4 ± 0.1 Hz, P < 0.05) and prolonged AP duration in SAN (n = 6). SAN myocytes had larger SK currents than did PV cardiomyocytes. Treatment with apamin (10 nM) resulted in a greater decrease (25 ± 8 vs. 20 ± 13%, P < 0.05) in spontaneous activity in isolated single SAN (n = 17) vs. PV cardiomyocytes (n = 16). Small-conductance calcium-activated potassium channels play a key role in SAN and PV spontaneous activity and AP morphology. Apamin may modulate PV electrical activity with and without endothelium dependence.

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

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. 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. 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. 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) .

Amyloid peptide regulates calcium homoeostasis and arrhythmogenesis in pulmonary vein cardiomyocytes

Amyloid peptides modulate cardiac calcium homoeostasis and play an important role in the pathophysiology of atrial fibrillation. Pulmonary veins (PVs) are critical in the genesis of atrial fibrillation and contain abundant amyloid peptides. Therefore, the purpose of this study is to investigate whether amyloid peptides may change the PV electrical activity through regulating calcium homoeostasis. The channel and calcium-handling protein expressions, intracellular calcium and ionic currents were studied in isolated rabbit PV cardiomyocytes in the presence and absence (control) of beta-amyloid (Aβ(25-35) ) for 4-6 h, using Western blot analysis, indo-1 fluorimetric ratio and whole-cell patch clamp techniques. Aβ(25-35) decreased the expressions of Ca(V) 1.2, total or Ser16-phosphorylated phospholamban (p-PLB), p-PLB/PLB ratio, sodium/calcium exchanger, but did not change ryanodine receptor, sarcoplasmic reticulum (SR) ATPase and K(+) channel proteins (Kir2.1, Kir2.3, Kv1.4, Kv1.5 and Kv4.2). Aβ(25-35) -treated cardiomyocytes had smaller calcium transient, SR calcium store, L-type calcium current and sodium/calcium exchanger current than control cardiomyocytes. Moreover, Aβ(25-35) -treated cardiomyocytes (n = 20) had shorter 90% of the action potential duration (82 ± 3 vs. 93 ± 5 ms, P < 0·05) than control cardiomyocytes (n = 16). Aβ(25-35) has direct electrophysiological effects on PV cardiomyocytes.

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.

Effects of Ivabradine on the Pulmonary Vein Electrical Activity and Modulation of Pacemaker Currents and Calcium Homeostasis

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. 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. 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.

Electromechanical effects of the direct renin inhibitor (aliskiren) on the pulmonary vein and atrium

Activation of the atrial renin-angiotensin system plays an important role in the pathophysiology of atrial fibrillation (AF). The pulmonary vein (PV) and left atrium (LA) are important trigger and substrate for the genesis of AF. We investigate the effects of a direct renin inhibitor, aliskiren, on the PV and LA arrhythmogenic activity and the underlying electromechanical mechanisms. Conventional microelectrodes were used to record action potentials and contractility in isolated rabbit PVs and LA tissues before and after the administration of aliskiren (0.1, 1, 3 and 10μM). By the whole-cell patch clamp and indo-1 fluorimetric ratio techniques, ionic currents and intracellular calcium transient were studied in isolated single PV and LA cardiomyocyte before and after the administration of aliskiren (3μM). Aliskiren (0.1, 1, 3 and 10μM) reduced PV firing rate in a concentration-dependent manner (6, 10, 14 and 17%) and decreased PV diastolic tension, which could be attenuated in the presence of 100μM L-N(G)-Nitroarginine Methyl Ester (L-NAME). Aliskiren induced PV automatic rhythm exit block causing slow and irregular PV activity with variable pauses. Aliskiren increased PV and LA contractility, which could be abolished by pre-treating with 0.1μM ryanodine. Aliskiren (3μM) decreased L-type calcium currents, but increased reverse-mode of Na( + )/Ca(2+ ) exchanger currents, intracellular calcium transients, and sarcoplasmic reticulum calcium content in PV and LA cardiomyocytes. Pretreatment with renin, losartan or angiotensin II did not alter the effect of aliskiren on sarcolemmal calcium flux. In conclusion, aliskiren reduces PV arrhythmogenic activity with a direct vasodilatory property and has a positive inotropic effect on cardiomyocytes. These findings may reveal the anti-arrhythmic and anti-heart failure potentials of aliskiren.

Eicosapentaenoic acid reduces the pulmonary vein arrhythmias through nitric oxide

AimsOmega-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 methodsConventional 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 findingsEPA 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. SignificanceEPA reduces PV arrhythmogenesis through the mechanoelectrical feedback generated by NO production.

Heat-Stress Responses Modulate Beta-Adrenergic Agonist and Angiotensin II Effects on the Arrhythmogenesis of Pulmonary Vein Cardiomyocytes

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. 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. 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.

Oxidative Stress on Pulmonary Vein and Left Atrium Arrhythmogenesis

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. 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). 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.