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1.
The proarrhythmic effects of new drugs have been assessed by measuring rapidly activating delayed-rectifier K+ current (IKr) antagonist potency. However, recent data suggest that even drugs thought to be highly specific IKr blockers can be arrhythmogenic via a separate, time-dependent pathway such as late Na+ current augmentation. Here, we report a mechanism for a quinolone antibiotic, sparfloxacin-induced action potential duration (APD) prolongation that involves increase in late L-type Ca2+ current (ICaL) caused by a decrease in Ca2+-dependent inactivation (CDI). Acute exposure to sparfloxacin, an IKr blocker with prolongation of QT interval and torsades de pointes (TdP) produced a significant APD prolongation in rat ventricular myocytes, which lack IKr due to E4031 pretreatment. Sparfloxacin reduced peak ICaL but increased late ICaL by slowing its inactivation. In contrast, ketoconazole, an IKr blocker without prolongation of QT interval and TdP produced reduction of both peak and late ICaL, suggesting the role of increased late ICaL in arrhythmogenic effect. Further analysis showed that sparfloxacin reduced CDI. Consistently, replacement of extracellular Ca2+ with Ba2+ abolished the sparfloxacin effects on ICaL. In addition, sparfloxacin modulated ICaL in a use-dependent manner. Cardiomyocytes from adult mouse, which is lack of native IKr, demonstrated similar increase in late ICaL and afterdepolarizations. The present findings show that sparfloxacin can prolong APD by augmenting late ICaL. Thus, drugs that cause delayed ICaL inactivation and IKr blockage may have more adverse effects than those that selectively block IKr. This mechanism may explain the reason for discrepancies between clinically reported proarrhythmic effects and IKr antagonist potencies.  相似文献   

2.
Cardiovascular complications are leading causes of morbidity and mortality in patients with chronic kidney disease (CKD). CKD significantly affects cardiac calcium (Ca2+) regulation, but the underlying mechanisms are not clear. The present study investigated the modulation of Ca2+ homeostasis in CKD mice. Echocardiography revealed impaired fractional shortening (FS) and stroke volume (SV) in CKD mice. Electrocardiography showed that CKD mice exhibited longer QT interval, corrected QT (QTc) prolongation, faster spontaneous activities, shorter action potential duration (APD) and increased ventricle arrhythmogenesis, and ranolazine (10 µmol/L) blocked these effects. Conventional microelectrodes and the Fluo-3 fluorometric ratio techniques indicated that CKD ventricular cardiomyocytes exhibited higher Ca2+ decay time, Ca2+ sparks, and Ca2+ leakage but lower [Ca2+]i transients and sarcoplasmic reticulum Ca2+ contents. The CaMKII inhibitor KN93 and ranolazine (RAN; late sodium current inhibitor) reversed the deterioration in Ca2+ handling. Western blots revealed that CKD ventricles exhibited higher phosphorylated RyR2 and CaMKII and reduced phosphorylated SERCA2 and SERCA2 and the ratio of PLB-Thr17 to PLB. In conclusions, the modulation of CaMKII, PLB and late Na+ current in CKD significantly altered cardiac Ca2+ regulation and electrophysiological characteristics. These findings may apply on future clinical therapies.  相似文献   

3.
The L-type Ca current (ICa,L), essential for normal cardiac function, also regulates dynamic action potential (AP) properties that promote ventricular fibrillation. Blocking ICa,L can prevent ventricular fibrillation, but only at levels suppressing contractility. We speculated that, instead of blocking ICa,L, modifying its shape by altering kinetic features could produce equivalent anti-fibrillatory effects without depressing contractility. To test this concept experimentally, we overexpressed a mutant Ca-insensitive calmodulin (CaM1234) in rabbit ventricular myocytes to inhibit Ca-dependent ICa,L inactivation, combined with the ATP-sensitive K current agonist pinacidil or ICa,L blocker verapamil to maintain AP duration (APD) near control levels. Cell shortening was enhanced in pinacidil-treated myocytes, but depressed in verapamil-treated myocytes. Both combinations flattened APD restitution slope and prevented APD alternans, similar to ICa,L blockade. To predict the arrhythmogenic consequences, we simulated the cellular effects using a new AP model, which reproduced flattening of APD restitution slope and prevention of APD/Cai transient alternans but maintained a normal Cai transient. In simulated two-dimensional cardiac tissue, these changes prevented the arrhythmogenic spatially discordant APD/Cai transient alternans and spiral wave breakup. These findings provide a proof-of-concept test that ICa,L can be targeted to increase dynamic wave stability without depressing contractility, which may have promise as an antifibrillatory strategy.  相似文献   

4.
Rate-dependent repolarization (RDR) of action potential (AP) in cardiomyocyte plays a critical role in the genesis of arrhythmias and RDR in atrium has been linked with atrial fibrillation. However, detailed studies focusing on the role of RDR in rabbit atrium are scant. In this study, atrial cells were isolated from rabbit heart and rate-dependent property was explored in single atrial cell to elucidate the underlying mechanism. Our results indicated that rate-dependent prolongation was evident at the action potential duration at 20% (APD20) and 50% (APD50) repolarization but not at 90% repolarization (APD90) under control condition. Using transient outward potassium current (Ito) inhibitor 4-Aminopyridine (4-AP, 2 mM) effectively eliminated the changes in APD20 and APD50, and unmasked the rate-dependent reduction of APD90 which could be diminished by further adding L-type calcium current (ICaL) inhibitor nifedipine (30 μM). However, using the selective late sodium current (INaL) inhibitor GS-458967 (GS967, 1 μM) caused minimal effect on APD90 of atrial cells both in the absence and presence of 4-AP. In consistence with results from APs, Ito and ICaL displayed significant rate-dependent reduction because of their slow reactivation kinetics. In addition, the magnitude of INaL in rabbit atrium was so small that its rate-dependent changes were negligible. In conclusion, our study demonstrated that Ito and ICaL mediate RDR of AP in rabbit atrium, while minimal effect of INaL was seen.  相似文献   

5.
Transient outward K+ current (Ito) plays a crucial role in the early phase of cardiac action potential repolarization. Kv4.3 K+ channel is an important component of Ito. The function and expression of Kv4.3 K+ channel decrease in variety of heart diseases, especially in heart hypertrophy/heart failure. In this review, we summarized the changes of cardiac Kv4.3 K+ channel in heart diseases and discussed the potential role of Kv4.3 K+ channel in heart hypertrophy/heart failure. In heart hypertrophy/heart failure of mice and rats, downregulation of Kv4.3 K+ channel leads to prolongation of action potential duration (APD), which is associated with increased [Ca2+]i, activation of calcineurin and heart hypertrophy/heart failure. However, in canine and human, Kv4.3 K+ channel does not play a major role in setting cardiac APD. So, in addition to Kv4.3 K+ channel/APD/[Ca2+]i pathway, there exits another mechanism of Kv4.3 K+ channel in heart hypertrophy and heart failure: downregulation of Kv4.3 K+ channels leads to CaMKII dissociation from Kv4.3–CaMKII complex and subsequent activation of the dissociated CaMKII, which induces heart hypertrophy/heart failure. Upregulation of Kv4.3 K+ channel inhibits CaMKII activation and its related harmful consequences. We put forward a new point-of-view that Kv4.3 K+ channel is involved in heart hypertrophy/heart failure independently of its electric function, and drugs inhibiting or upregulating Kv4.3 K+ channel might be potentially harmful or beneficial to hearts through CaMKII.  相似文献   

6.
Transient outward K+ current (Ito) plays a crucial role in the early phase of cardiac action potential repolarization. Kv4.3 K+ channel is an important component of Ito. The function and expression of Kv4.3 K+ channel decrease in variety of heart diseases, especially in heart hypertrophy/heart failure. In this review, we summarized the changes of cardiac Kv4.3 K+ channel in heart diseases and discussed the potential role of Kv4.3 K+ channel in heart hypertrophy/heart failure. In heart hypertrophy/heart failure of mice and rats, downregulation of Kv4.3 K+ channel leads to prolongation of action potential duration (APD), which is associated with increased [Ca2+]i, activation of calcineurin and heart hypertrophy/heart failure. However, in canine and human, Kv4.3 K+ channel does not play a major role in setting cardiac APD. So, in addition to Kv4.3 K+ channel/APD/[Ca2+]i pathway, there exits another mechanism of Kv4.3 K+ channel in heart hypertrophy and heart failure: downregulation of Kv4.3 K+ channels leads to CaMKII dissociation from Kv4.3–CaMKII complex and subsequent activation of the dissociated CaMKII, which induces heart hypertrophy/heart failure. Upregulation of Kv4.3 K+ channel inhibits CaMKII activation and its related harmful consequences. We put forward a new point-of-view that Kv4.3 K+ channel is involved in heart hypertrophy/heart failure independently of its electric function, and drugs inhibiting or upregulating Kv4.3 K+ channel might be potentially harmful or beneficial to hearts through CaMKII.  相似文献   

7.

Background

The repolarizing cardiac rapid delayed rectifier current, I Kr, is composed of ERG1 channels. It has been suggested that two isoforms of the ERG1 protein, ERG1a and ERG1b, both contribute to I Kr. Marked heterogeneity in the kinetic properties of native I Kr has been described. We hypothesized that the heterogeneity of native I Kr can be reproduced by differential expression of ERG1a and ERG1b isoforms. Furthermore, the functional consequences of differential expression of ERG1 isoforms were explored as a potential mechanism underlying native heterogeneity of action potential duration (APD) and restitution.

Methodology/Principal Findings

The results show that the heterogeneity of native I Kr can be reproduced in heterologous expression systems by differential expression of ERG1a and ERG1b isoforms. Characterization of the macroscopic kinetics of ERG1 currents demonstrated that these were dependent on the relative abundance of ERG1a and ERG1b. Furthermore, we used a computational model of the ventricular cardiomyocyte to show that both APD and the slope of the restitution curve may be modulated by varying the relative abundance of ERG1a and ERG1b. As the relative abundance of ERG1b was increased, APD was gradually shortened and the slope of the restitution curve was decreased.

Conclusions/Significance

Our results show that differential expression of ERG1 isoforms may explain regional heterogeneity of I Kr kinetics. The data demonstrate that subunit dependent changes in channel kinetics are important for the functional properties of ERG1 currents and hence I Kr. Importantly, our results suggest that regional differences in the relative abundance of ERG1 isoforms may represent a potential mechanism underlying the heterogeneity of both APD and APD restitution observed in mammalian hearts.  相似文献   

8.
AimsDiabetes mellitus is associated with changes of α1-adrenoceptor (α1-AR) on heart electrical function and expression. In this study, we investigated the ionic basis underlying abnormal α1-AR mediated QT prolongation in the diabetic rat hearts.Main methodsElectrophysiological and biochemical techniques were used in Streptozotocin (STZ)-induced diabetic and control rat hearts.Key findingsIn both control and diabetic rats, the α1-AR agonist, phenylephrine (PE, 10–100 µM) prolonged the rate-corrected QT intervals (QTc) and action potential durations at 30% (APD30) and 90% (APD90) repolarization levels with the increased QTc and APD90 significantly greater in diabetic rats. PE significantly decreased the transient outward K+ current (Ito) and the steady-state K+ current (Iss) in both control and diabetic rats but had no effects on the delayed rectifier K+ current (Ik). However, PE induced a greater reduction mainly in the Iss, but not Ito, in diabetic rats. Furthermore, using RT–PCR and Western blot analyses, we found that α1A-ARs were over-expressed in the left ventricular tissues of the diabetic rat hearts at both the mRNA and the protein levels.SignificanceThese data suggested that in diabetic hearts, a greater sensitivity of the α1A-AR mediated the larger suppression of Iss and resulted in a more prolonged APD90 and QTc. Thus, higher α1A-AR expression levels in diabetic heart may underlie this type of diabetic cardiomyopathy and suggests that α1A-AR may serve as a therapeutic target.  相似文献   

9.
Cholesterol affects diverse biological processes, in many cases by modulating the function of integral membrane proteins. In this study we have investigated the role of cholesterol in the adenosine-dependent regulation of ion transport in colonic epithelial cells. We observed that methyl-β-cyclodextrin (MβCD), a cholesterol-sequestering molecule, enhanced adenosine A2A receptor-activated transepithelial short circuit current (Isc), but only from the basolateral side. Cholesterol is a major constituent of membrane microdomains, called lipid rafts that also contain sphingolipids. However, studies with the sphingomyelin-degrading enzyme, sphingomyelinase, and the cholesterol-binding agent, filipin, indicated that the change in the level of cholesterol alone was sufficient to control the adenosine-modulated Isc. Cholesterol depletion had a major effect on the functional selectivity of A2A receptors. Under control conditions, adenosine activated Isc more potently than the specific A2A agonist, CGS-21680, and the current was inhibited by XE991, an inhibitor of cAMP-dependent K+ channels. Following cholesterol depletion, CGS-21680 activated Isc more potently than adenosine, and the current was inhibited by clotrimazole, an inhibitor of Ca2+-activated K+ (IK1) channels. Co-immunoprecipitation experiments revealed that A2A receptors associate with IK1 channels following cholesterol depletion. These results suggest that cholesterol content in colonic epithelia affects adenosine-mediated anion secretion by controlling agonist-selective signaling.  相似文献   

10.
Persistent atrial fibrillation (PeAF) in humans is characterized by shortening of action potential duration (APD) and attenuation of APD rate-adaptation. However, the quantitative influences of particular ionic current alterations on rate-dependent APD changes, and effects on patterns of reentry in atrial tissue, have not been systematically investigated. Using mathematical models of human atrial cells and tissue and performing parameter sensitivity analysis, we evaluated the quantitative contributions to action potential (AP) shortening and APD rate-adaptation of ionic current remodeling seen with PeAF. Ionic remodeling in PeAF was simulated by reducing L-type Ca2+ channel current (ICaL), increasing inward rectifier K+ current (IK1) and modulating five other ionic currents. Parameter sensitivity analysis, which quantified how each ionic current influenced APD in control and PeAF conditions, identified interesting results, including a negative effect of Na+/Ca2+ exchange on APD only in the PeAF condition. At high pacing rate (2 Hz), electrical remodeling in IK1 alone accounts for the APD reduction of PeAF, but at slow pacing rate (0.5 Hz) both electrical remodeling in ICaL alone (-70%) and IK1 alone (+100%) contribute equally to the APD reduction. Furthermore, AP rate-adaptation was affected by IKur in control and by INaCa in the PeAF condition. In a 2D tissue model, a large reduction (-70%) of ICaL becomes a dominant factor leading to a stable spiral wave in PeAF. Our study provides a quantitative and unifying understanding of the roles of ionic current remodeling in determining rate-dependent APD changes at the cellular level and spatial reentry patterns in tissue.  相似文献   

11.
Beat-to-beat variability of repolarization duration (BVR) is an intrinsic characteristic of cardiac function and a better marker of proarrhythmia than repolarization prolongation alone. The ionic mechanisms underlying baseline BVR in physiological conditions, its rate dependence, and the factors contributing to increased BVR in pathologies remain incompletely understood. Here, we employed computer modeling to provide novel insights into the subcellular mechanisms of BVR under physiological conditions and during simulated drug-induced repolarization prolongation, mimicking long-QT syndromes type 1, 2, and 3. We developed stochastic implementations of 13 major ionic currents and fluxes in a model of canine ventricular-myocyte electrophysiology. Combined stochastic gating of these components resulted in short- and long-term variability, consistent with experimental data from isolated canine ventricular myocytes. The model indicated that the magnitude of stochastic fluctuations is rate dependent due to the rate dependence of action-potential (AP) duration (APD). This process (the “active” component) and the intrinsic nonlinear relationship between membrane current and APD (“intrinsic component”) contribute to the rate dependence of BVR. We identified a major role in physiological BVR for stochastic gating of the persistent Na+ current (INa) and rapidly activating delayed-rectifier K+ current (IKr). Inhibition of IKr or augmentation of INa significantly increased BVR, whereas subsequent β-adrenergic receptor stimulation reduced it, similar to experimental findings in isolated myocytes. In contrast, β-adrenergic stimulation increased BVR in simulated long-QT syndrome type 1. In addition to stochastic channel gating, AP morphology, APD, and beat-to-beat variations in Ca2+ were found to modulate single-cell BVR. Cell-to-cell coupling decreased BVR and this was more pronounced when a model cell with increased BVR was coupled to a model cell with normal BVR. In conclusion, our results provide new insights into the ionic mechanisms underlying BVR and suggest that BVR reflects multiple potentially proarrhythmic parameters, including increased ion-channel stochasticity, prolonged APD, and abnormal Ca2+ handling.  相似文献   

12.
Experimental evidence suggests that regional differences in action potential (AP) morphology can provide a substrate for initiation and maintenance of reentrant arrhythmias in the right atrium (RA), but the relationships between the complex electrophysiological and anatomical organization of the RA and the genesis of reentry are unclear. In this study, a biophysically detailed three-dimensional computer model of the right atrial tissue was constructed to study the role of tissue heterogeneity and anisotropy in arrhythmogenesis. The model of Lindblad et al. for a rabbit atrial cell was modified to incorporate experimental data on regional differences in several ionic currents (primarily, INa, ICaL, IK1, Ito, and Isus) between the crista terminalis and pectinate muscle cells. The modified model was validated by its ability to reproduce the AP properties measured experimentally. The anatomical model of the rabbit RA (including tissue geometry and fiber orientation) was based on a recent histological reconstruction. Simulations with the resultant electrophysiologically and anatomically detailed three-dimensional model show that complex organization of the RA tissue causes breakdown of regular AP conduction patterns at high pacing rates (>11.75 Hz): as the AP in the crista terminalis cells is longer, and electrotonic coupling transverse to fibers of the crista terminalis is weak, high-frequency pacing at the border between the crista terminalis and pectinate muscles results in a unidirectional conduction block toward the crista terminalis and generation of reentry. Contributions of the tissue heterogeneity and anisotropy to reentry initiation mechanisms are quantified by measuring action potential duration (APD) gradients at the border between the crista terminalis and pectinate muscles: the APD gradients are high in areas where both heterogeneity and anisotropy are high, such that intrinsic APD differences are not diminished by electrotonic interactions. Thus, our detailed computer model reconstructs complex electrical activity in the RA, and provides new insights into the mechanisms of transition from focal atrial tachycardia into reentry.  相似文献   

13.
Background Gender disparities in the incidence of torsade de pointes (TdP) ventricular tachycardia exist, but the mechanisms in humans are unresolved. We addressed this issue using a mathematical model of a human ventricular cell. MethodsWe implemented gender differences in the Priebe-Beuckelmann model cell by modifying the amplitudes of the L-type Ca2+ current (ICa,L), transient outward K+ current (Ito), and rapid component of the delayed rectifier K+current (IKr), according to experimental data from animal male and female hearts. Gender disparities in electrical heterogeneity between transmural layers (subepicardium, midmyocardium, subendocardium) were implemented by modifying various ion currents according to experimental data. ResultsAction potentials in female cells have longer durations and steeper duration versus frequency relationships than male cells. In the female cells, electrical heterogeneity between transmural layers is larger and the susceptibility to early afterdepolarisations is higher than in male cells. ConclusionGender-related differences in ICa,L, Ito, and IKr may explain the gender disparities in human cardiac electrophysiology. Female cells have an increased susceptibility to early afterdepolarisations following mild reductions in net repolarising forces. Combined with their greater electrical heterogeneity, this renders them more vulnerable to TdP. (Neth Heart J 2007;15:405-11.)  相似文献   

14.
15.
KV2.1 is the prominent somatodendritic sustained or delayed rectifier voltage-gated potassium (Kv) channel in mammalian central neurons, and is a target for activity-dependent modulation via calcineurin-dependent dephosphorylation. Using hanatoxin-mediated block of KV2.1 we show that, in cultured rat hippocampal neurons, glutamate stimulation leads to significant hyperpolarizing shifts in the voltage-dependent activation and inactivation gating properties of the KV2.1-component of delayed rectifier K+ (IK) currents. In computer models of hippocampal neurons, these glutamate-stimulated shifts in the gating of the KV2.1-component of IK lead to a dramatic suppression of action potential firing frequency. Current-clamp experiments in cultured rat hippocampal neurons showed glutamate-stimulation induced a similar suppression of neuronal firing frequency. Membrane depolarization also resulted in similar hyperpolarizing shifts in the voltage-dependent gating properties of neuronal IK currents, and suppression of neuronal firing. The glutamate-induced effects on neuronal firing were eliminated by hanatoxin, but not by dendrotoxin-K, a blocker of KV1.1-containing channels. These studies together demonstrate a specific contribution of modulation of KV2.1 channels in the activity-dependent regulation of intrinsic neuronal excitability.  相似文献   

16.
17.

Aim

Hydrogen sulfide (H2S) is a promising cardioprotective agent and a potential modulator of cardiac ion currents. Yet its cardiac effects on humans are poorly understood due to lack of functional cardiomyocytes. This study investigates electrophysiological responses of human pluripotent stem cells (hPSCs) derived cardiomyocytes towards H2S.

Methods and Results

Cardiomyocytes of ventricular, atrial and nodal subtypes differentiated from H9 embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs) were electrophysiologically characterized. The effect of NaHS, a donor of H2S, on action potential (AP), outward rectifier potassium currents (I Ks and I Kr), L-type Ca2+ currents (I CaL) and hyperpolarization-activated inward current (I f) were determined by patch-clamp electrophysiology and confocal calcium imaging. In a concentration-dependent manner, NaHS (100 to 300 µM) consistently altered the action potential properties including prolonging action potential duration (APD) and slowing down contracting rates of ventricular-and atrial-like cardiomyocytes derived from both hESCs and hiPSCs. Moreover, inhibitions of slow and rapid I K (I Ks and I Kr), I CaL and I f were found in NaHS treated cardiomyocytes and it could collectively contribute to the remodeling of AP properties.

Conclusions

This is the first demonstration of effects of H2S on cardiac electrophysiology of human ventricular-like, atrial-like and nodal-like cardiomyocytes. It reaffirmed the inhibitory effect of H2S on I CaL and revealed additional novel inhibitory effects on I f, I Ks and I Kr currents in human cardiomyocytes.  相似文献   

18.
Lysophosphatidic acid (LPA) has diverse actions on the cardiovascular system and is widely reported to modulate multiple ion currents in some cell types. However, little is known about its electrophysiological effects on cardiac myocytes. This study investigated whether LPA has electrophysiological effects on isolated rabbit myocardial preparations. The results indicate that LPA prolongs action potential duration at 90% repolarization (APD90) in a concentration- and frequency-dependent manner in isolated rabbit ventricular myocytes. The application of extracellular LPA significantly increases the coefficient of APD90 variability. LPA increased L-type calcium current (ICa,L) density without altering its activation or deactivation properties. In contrast, LPA has no effect on two other ventricular repolarizing currents, the transient outward potassium current (Ito) and the delayed rectifier potassium current (IK). In arterially perfused rabbit left ventricular wedge preparations, the monophasic action potential duration, QT interval, and Tpeak-end are prolonged by LPA. LPA treatment also significantly increases the incidence of ventricular tachycardia induced by S1S2 stimulation. Notably, the effects of LPA on action potentials and ICa,L are PTX-sensitive, suggesting LPA action requires a Gi-type G protein. In conclusion, LPA prolongs APD and increases electrophysiological instability in isolated rabbit myocardial preparations by increasing ICa,L in a Gi protein-dependent manner.  相似文献   

19.
We aimed to investigate the role of activin receptor-like kinase (ALK7) in regulating cardiac electrophysiology. Here, we showed that Alk7-/- mice exhibited prolonged QT intervals in telemetry ECG recordings. Furthermore, Langendorff-perfused Alk7-/- hearts had significantly longer action potential duration (APD) and greater incidence of ventricular arrhythmia (AV) induced by burst pacing. Using whole-cell patch clamp, we found that the densities of repolarizing K+ currents Ito and IK1 were profoundly reduced in Alk7-/- ventricular cardiomyocytes. Mechanistically, the expression of Kv4.2 (a major subunit of Ito carrying channel) and KCHIP2 (a key accessory subunit of Ito carrying channel), was markedly decreased in Alk7-/- hearts. These findings suggest that endogenous expression of ALK7 is necessary to maintain repolarizing K+ currents in ventricular cardiomyocytes, and finally prevent action potential prolongation and ventricular arrhythmia.  相似文献   

20.
Drug-mediated or medical condition-mediated disruption of hERG function accounts for the main cause of acquired long-QT syndrome (acLQTs), which predisposes affected individuals to ventricular arrhythmias (VA) and sudden death. Many Chinese herbal medicines, especially alkaloids, have risks of arrhythmia in clinical application. The characterized mechanisms behind this adverse effect are frequently associated with inhibition of cardiac hERG channels. The present study aimed to assess the potent effect of Rutaecarpine (Rut) on hERG channels. hERG-HEK293 cell was applied for evaluating the effect of Rut on hERG channels and the underlying mechanism. hERG current (IhERG) was measured by patch-clamp technique. Protein levels were analysed by Western blot, and the phosphorylation of Sp1 was determined by immunoprecipitation. Optical mapping and programmed electrical stimulation were used to evaluate cardiac electrophysiological activities, such as APD, QT/QTc, occurrence of arrhythmia, phase singularities (PSs), and dominant frequency (DF). Our results demonstrated that Rut reduced the IhERG by binding to F656 and Y652 amino acid residues of hERG channel instantaneously, subsequently accelerating the channel inactivation, and being trapped in the channel. The level of hERG channels was reduced by incubating with Rut for 24 hours, and Sp1 in nucleus was inhibited simultaneously. Mechanismly, Rut reduced threonine (Thr)/ tyrosine (Tyr) phosphorylation of Sp1 through PI3K/Akt pathway to regulate hERG channels expression. Cell-based model unables to fully reveal the pathological process of arrhythmia. In vivo study, we found that Rut prolonged QT/QTc intervals and increased induction rate of ventricular fibrillation (VF) in guinea pig heart after being dosed Rut for 2 weeks. The critical reasons led to increased incidence of arrhythmias eventually were prolonged APD90 and APD50 and the increase of DF, numbers of PSs, incidence of early after-depolarizations (EADs). Collectively, the results of this study suggest that Rut could reduce the IhERG by binding to hERG channels through F656 and Y652 instantaneously. While, the PI3K/Akt/Sp1 axis may play an essential role in the regulation of hERG channels, from the perspective of the long-term effects of Rut (incubating for 24 hours). Importantly, the changes of electrophysiological properties by Rut were the main cause of VA.  相似文献   

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