Repolarization Of Ventricular Action Potential Research Articles

Insulin-induced recovery in KCNQ1/KCNE1-current accelerates the ventricular action potential repolarization in insulin-resistant aged-rats via affecting β3-adrenergic receptors

Cardiac aging, including cardiometabolic disturbances, is defined as a progressive and intrinsic decline in heart function, which is also a combination of metabolic dysfunctions mainly characterized by insulin resistance and long-QT, via prolonged late-phase repolarizations of ventricular action potentials (APs). So, we hypothesized a contribution of dysfunctional KCNQ1/KCNE1-channels to these abnormal electrical activities in elderly mammal hearts and positive responses to insulin application. Furthermore, taken into consideration β3-AR activation role in insulin-resistant hearts, we aimed to investigate the possible correlations between dysfunctional KCNQ1/KCNE1-channels, β3-AR activation, and the effect of insulin application on these correlations in insulin-resistant aged rat (24-month-old) ventricular cardiomyocytes.

Insulin acts as an atypical KCNQ1/KCNE1‐current activator and reverses long QT in insulin‐resistant aged rats by accelerating the ventricular action potential repolarization through affecting the β3‐adrenergic receptor signaling pathway

Insufficient-heart function is associated with myocardial insulin resistance in the elderly, particularlyassociated with long-QT, in a dependency on dysfunctional KCNQ1/KCNE1-channels. So, we aimed to examine the contribution of alterations in KCNQ1/KCNE1-current (IKs ) to the aging-related remodeling of the heart as well as the role of insulin treatment on IKs in the aged rats. Prolonged late-phase action potential (AP) repolarization of ventricular cardiomyocytes from insulin-resistant 24-month-old rats was significantly reversed by in vitro treatment of insulin or PKG inhibitor (in vivo, as well) via recovery in depressed IKs . Although the protein level of either KCNQ1 or KCNE1 in cardiomyocytes was not affected with aging, PKG level was significantly increased in those cells. The inhibited IKs in β3 -ARs-stimulated cells could be reversed with a PKG inhibitor, indicating the correlation between PKG-activation and β3 -ARs activation. Furthermore, in vivo treatment of aged rats, characterized by β3 -ARs activation, with either insulin or a PKG inhibitor for 2 weeks provided significant recoveries in IKs , prolonged late phases of APs, prolonged QT-intervals, and low heart rates without no effect on insulin resistance. In vivo insulin treatment provided also significant recovery in increased PKG and decreased PIP2 level, without the insulin effect on the KCNQ1 level in β3 -ARs overexpressed cells. The inhibition of IKs in aged-rat cardiomyocytes seems to be associated with activated β3 -ARs dependent remodeling in the interaction between KCNQ1 and KCNE1. Significant recoveries in ventricular-repolarization of insulin-treated aged cardiomyocytes via recovery in IKs strongly emphasize two important issues: (1) IKs can be a novel target in aging-associated remodeling in the heart and insulin may be a cardioprotective agent in the maintenance of normal heart function during the aging process. (2) This study is one of the first to demonstrate insulin'sbenefits on long-QT in insulin-resistant aged rats by accelerating the ventricular AP repolarization through reversing the depressed IKs via affecting the β3 -ARs signaling pathway and particularly affecting activated PKG.

Specific protein kinase C isoform exerts chronic inhibition on the slowly activating delayed-rectifier potassium current by affecting channel trafficking

ABSTRACT The slowly activating delayed rectifier K+ current (I Ks) plays a key role in the repolarization of ventricular action potential in the human heart and is formed by the pore-forming α-subunit encoded by KCNQ1 (Kv7.1) and β-subunit encoded by KCNE1. Evidence suggested that I Ks was regulated through protein kinase C (PKC) pathway, but the mechanism is controversial. This study was designed to identify the specific PKC isoform involved in the long-term regulation of I Ks current. The I Ks current was recorded using whole-cell patch-clamp technique in human embryonic kidney (HEK) 293B cell co-transfected with human KCNQ1/KCNE1 genes. The results revealed that both chronic activation of Ang II and PMA reduced the I Ks current in a long-term regulation (about 24 hours). Further evidence showed that PKCε knockdown by siRNA antagonized the AngII-induced chronic inhibition on the I Ks current, whereas knockdown of cPKC (PKCα and PKCβ) attenuated the inhibition effect of PMA on the current. Moreover, the forward transport inhibition of the channel with brefeldin A alleviated the Ang II-induced chronic inhibition on I Ks current, while the channel endocytosis inhibition with dynasore alleviated both Ang II and PMA-induced chronic inhibition on I Ks current. The above results showed that PKCε activation promoted the channel endocytosis and inhibited the channel forward transport to the plasma membrane, while cPKC activation only promoted the channel endocytosis, which both down regulated the channel current.

A computational analysis of the effect of sevoflurane in a human ventricular cell model of long QT syndrome: Importance of repolarization reserve in the QT-prolonging effect of sevoflurane

The slowly and rapidly activating delayed rectifier K+ channels (IKs and IKr, respectively) contribute to the repolarization of ventricular action potential in human heart and thereby determine QT interval on an electrocardiogram. Loss-of-function mutations in genes encoding IKs and IKr cause type 1 and type 2 long QT syndrome (LQT1 and LQT2, respectively), accompanied by a high risk of malignant ventricular arrhythmias and sudden cardiac death. This study was designed to investigate which cardiac electrophysiological conditions exaggerate QT-prolonging and arrhythmogenic effects of sevoflurane. We used the O'Hara-Rudy dynamic model to reconstruct human ventricular action potential and a pseudo-electrocardiogram, and simulated LQT1 and LQT2 phenotypes by decreasing conductances of IKs and IKr, respectively. Sevoflurane, but not propofol, prolonged ventricular action potential duration and QT interval in wild-type, LQT1 and LQT2 models. The QT-prolonging effect of sevoflurane was more profound in LQT2 than in wild-type and LQT1 models. The potent inhibitory effect of sevoflurane on IKs was primarily responsible for its QT-prolonging effect. In LQT2 model, IKs was considerably enhanced during excessive prolongation of ventricular action potential duration by reduction of IKr and relative contribution of IKs to ventricular repolarization was markedly elevated, which appears to underlie more pronounced QT-prolonging effect of sevoflurane in LQT2 model, compared with wild-type and LQT1 models. This simulation study clearly elucidates the electrophysiological basis underlying the difference in QT-prolonging effect of sevoflurane among wild-type, LQT1 and LQT2 models, and may provide important information for developing anesthetic strategies for patients with long QT syndrome in clinical settings.

Pharmacological inhibition of IK1 by PA-6 in isolated rat hearts affects ventricular repolarization and refractoriness.

The inwardly rectifying potassium current (IK 1) conducted through Kir2.X channels contribute to repolarization of the cardiac action potential and to stabilization of the resting membrane potential in cardiomyocytes. Our aim was to investigate the effect of the recently discovered IK 1 inhibitor PA‐6 on action potential repolarization and refractoriness in isolated rat hearts. Transiently transfected HEK‐293 cells expressing IK 1 were voltage‐clamped with ramp protocols. Langendorff‐perfused heart experiments were performed on male Sprague–Dawley rats, effective refractory period, Wenckebach cycle length, and ventricular effective refractory period were determined following 200 nmol/L PA‐6 perfusion. 200 nmol/L PA‐6 resulted in a significant time‐latency in drug effect on the IK 1 current expressed in HEK‐293 cells, giving rise to a maximal effect at 20 min. In the Langendorff‐perfused heart experiments, PA‐6 prolonged the ventricular action potential duration at 90% repolarization (from 41.8 ± 6.5 msec to 72.6 ± 21.1 msec, 74% compared to baseline, P < 0.01, n = 6). In parallel, PA‐6 significantly prolonged the ventricular effective refractory period compared to baseline (from 34.8 ± 4.6 msec to 58.1 ± 14.7 msec, 67%, P < 0.01, n = 6). PA‐6 increased the short‐term beat‐to‐beat variability and ventricular fibrillation was observed in two of six hearts. Neither atrial ERP nor duration of atrial fibrillation was altered following PA‐6 application. The results show that pharmacological inhibition of cardiac IK 1 affects ventricular action potential repolarization and refractoriness and increases the risk of ventricular arrhythmia in isolated rat hearts.

New micro waveforms firstly recorded on electrocardiogram in human

In our study, not only the P-QRS-T waves but also the micro-wavelets before QRS complex (in P wave and PR segment) and after QRS complex (ST segment and upstroke of T wave) were first to be identified on surface electrocardiogram in human by the “new electrocardiogram” machine (model PHS-A10) according to conventional 12-lead electrocardiogram connection methods. By comparison to the conventional electrocardiogram in 100 cases of healthy individuals and several patients with arrhythmias, we have found that the wavelets before P wave theoretically reflected electrical activity of sinus node and the micro-wavelets before QRS complex may be related to atrioventricular conduction system (atrioventricular node, His bundle and bundle branch) potentials. Noninvasive atrioventricular node and His bundle potential tracing will contribute to differentiation of the origin of wide QRS and the location of the atrioventricular block. We also have found that the wavelets after QRS complex may be associated with phase 2 and 3 repolarization of ventricular action potential, which will further reveal ventricular repolarization changes.

Long QT Syndromes Are Heterogeneous Disease Entities Presenting not only QT Prolongation but Multiple Phenotypes—Cases with Compound Heterozygous Mutations

Long QT syndromes (LQTS) consist of heterogeneous disease entities characterized by a remarkable prolongation of QT interval on ECG and a peculiar polymorphic ventricular tachycardia called “torsade de pointes” , which often degenerates into ventricular fibrillation and causing cardiogenic syncope or sudden death. Because of extensive studies on familial LQTS patients using the molecular genetics since early 1990s, it has been shown that genetic variants in genes encoding cardiac ion channels or their modulating proteins cause the delayed repolarization of ventricular action potential and thereby QT prolongation as a phenotype. Up to date, a variety of mutations in 13 different genes have been shown to be associated with LQTS. In the clinical setting, however, the first three subtypes (LQT1-3) are most frequently seen and presenting quite different phenotypes. Because the gene responsible for each subtype encodes a distinct ion current (IKs, IKr and INa), it is no wonder that the malfunction of specific channel leads to different clinical features. On a thorough screen for candidate genes in our multicenter study, in addition, we recognized that there were a substantial number of compound mutation cases (26 among 310 probands, 8.4%). In this symposium, we would like to make a brief overview on Japanese LQTS.

Evolution of ventricular myocyte electrophysiology

The relative importance of regulatory versus structural evolution for the evolution of different biological systems is a subject of controversy. The primacy of regulatory evolution in the diversification of morphological traits has been promoted by many evolutionary developmental biologists. For physiological traits, however, the role of regulatory evolution has received less attention or has been considered to be relatively unimportant. To address this issue for electrophysiological systems, we examined the importance of regulatory and structural evolution in the evolution of the electrophysiological function of cardiac myocytes in mammals. In particular, two related phenomena were studied: the change in action potential morphology in small mammals and the scaling of action potential duration across mammalian phylogeny. In general, the functional properties of the ion channels involved in ventricular action potential repolarization were found to be relatively invariant. In contrast, there were large changes in the expression levels of multiple ion channel and transporter genes. For the Kv2.1 and Kv4.2 potassium channel genes, which are primary determinants of the action potential morphology in small mammals, the functional properties of the proximal promoter regions were found to vary in concordance with species-dependent differences in mRNA expression, suggesting that evolution of cis-regulatory elements is the primary determinant of this trait. Scaling of action potential duration was found to be a complex phenomenon, involving changes in the expression of a large number of channels and transporters. In this case, it is concluded that regulatory evolution is the predominant mechanism by which the scaling is achieved.

Refining Insights into High-Affinity Drug Binding to the Human Ether-à-go-go-Related Gene Potassium Channel: Fig. 1.

hERG (human ether-à-go-go-related gene) potassium (K(+)) channels play a crucial role in electrophysiological activity in the heart, exerting a profound influence on ventricular action potential repolarization and on the duration of the QT interval of the electrocardiogram. hERG channels are strongly implicated in the acquired form of long QT syndrome in that they exhibit a unique susceptibility to pharmacological inhibition by therapeutically and chemically diverse drugs. Investigations over a number of years provide compelling evidence that a comparatively large inner cavity and the presence of particular aromatic amino acid residues (Tyr652 and Phe656) on the inner (S6) helices of the channel are important features that allow hERG to accommodate and bind disparate drugs. However, whereas functional hERG channels are composed of four identical subunits, blocking molecules may not interact equally with aromatic residues from each of the four subunits. In this issue of Molecular Pharmacology, Myokai et al. (p. 1643) report for the first time the use of tandem dimers incorporating mutations to Tyr652 and Phe656 to elucidate asymmetric binding of the high affinity hERG inhibitor cisapride. Not only has this approach provided increased information on spatial arrangements involved in cisapride binding to the channel, but it offers a powerful means of refining the wider understanding of hERG channel structure-function in relation to drug binding.

Calcium-activated transient outward chloride current and phase 1 repolarization of swine ventricular action potential.

It is unknown whether 4-aminopyridine- (4-AP-) sensitive transient outward K(+) current (I(to1)) and/or Ca(2+)-activated transient outward Cl(-) current (I(Ca.Cl) or I(to2)) contribute(s) to phase 1 repolarization of pig ventricular action potential (AP). The purpose of the present study was to determine ionic contribution of the phase 1 repolarization of AP in pig ventricle. We used whole-cell patch techniques to record APs and membrane currents, and Western immunoblotting analysis to detect expression of I(to1) protein (Kv4.2 or Kv4.3) in pig ventricular myocytes. A transient outward current (I(to)) was activated upon depolarization voltage steps to between -10 and +60 mV from -50 mV in pig ventricular cells, and the I(to) was resistant to 4-AP application, but sensitive to the inhibition by ryanodine (10 micromol/l) and the Ca(2+) channel blockade, and the Cl(-) channel blocker 4,4'-diisothiocyanostilben-2,2'disulfonic acid (DIDS, 150 micromol/l). The current was diminished by external Cl(-) (Cl(-)(o)) replacement and showed a 'bell-shaped' I-V relationship at room temperature, typical of I(to2). No difference in I(to2) was observed in the regional cells from epicardium, midmyocardium, and endocardium of left ventricle. APs showed significant phase 1 and 'spike and dome' in pig ventricular myocytes. The phase 1 and 'spike and dome' of APs were not affected by 4-AP (3 mmol/l), but abolished by replacing Cl(-)(o) and by application of 100 micromol/l DIDS, suggesting I(to2) contribution. Western immunoblotting analysis showed no evidence for the expression of 4-AP-sensitive I(to1) channel protein (Kv4.2 or Kv4.3) in pig ventricular cells. The results indicate that 4-AP-sensitive I(to1) is not expressed, and only Ca(2+)-activated I(to2) is present in pig cardiac cells, which contributes importantly to the phase 1 repolarization of ventricular APs in this species.

Block of recombinant KCNQ1/KCNE1 K+ channels (IKs) by intracellular Na+ and its implications on action potential repolarization.

I(Ks), the slow component of delayed rectifier K+ current, plays an important role for the repolarization of ventricular action potential. We investigated the block of I(Ks) by intracellular Na+ ([Na+](i)), using a heterologous expression system (KCNQ1/KCNE1 expressed in COS7 cells), since this well-known blocking action on various K+ channels has not been fully or quantitatively characterized in I(Ks) current. The Na+ block of I(Ks) was concentration- and voltage-dependent and was described by a conventional binding-site model (Woodhull AM: J Gen Physiol 61: 687-708, 1973). In physiological ionic conditions, the blocking action was operating noticeably with Delta ("electrical" distance of the block site) approximately 0.6 and K(d)(0) (apparent dissociation constant at 0 mV) approximately 300 mM. Because K(d)(0) was a function of intra- and extracellular K+ concentrations, changes in ionic environments not only of [Na+](i), but also of [K+](o), affected the amplitude of I(Ks) through the modulation of the Na+ block. Based on these experimental data, we analyzed the effects of Na+ block on action potentials by a computer simulation study, using the Luo-Rudy model. In a physiological ionic environment, the Na+ block of I(Ks) contributed little to modifying action potentials. However, when action potential duration (APD) was marginally prolonged because of decreased I(Ks), as observed in M cells under the conditions of bradycardia and low [K+](o), the Na+ block of I(Ks) may contribute to arrhythmogenesis through the facilitation of early afterdepolarizations (EADs).

Reduced repolarization reserve in ventricular myocytes from female mice

Cardiac repolarization is prolonged and repolarization reserve (RR) is diminished in female rabbits and humans, compared to males. Reduced RR is evidenced by the relatively greater increase in ventricular action potential duration (APD) in myocytes from females in response to drugs that block repolarizing K(+) currents. Mice are an increasingly important experimental model animal for cardiovascular research, but gender-dependent differences have not been reported for repolarization in murine ventricular myocytes. APD and repolarizing K(+) currents were measured in isolated ventricular myocytes from adult littermate male and female mice. Repolarizing K(+) currents were dissected into transient (I(to)) and sustained (I(sus)) components and the selective I(sus) antagonist FK506 was used to probe for differences in RR. Under control conditions APD at 50% (APD(50)) and at 90% (APD(90)) repolarization was significantly longer in females (APD(50)=15 +/- 3 ms, n=6 and APD(90)=63 +/- 6 ms, n=6) compared to males (APD(50)=8 +/- 2 ms, n=7 and APD(90)=42 +/- 9 ms, n=7) at 1.0 Hz. At 0.3 Hz stimulation frequency APD(90), but not APD(50), was significantly longer in females (APD(50)=12 +/- 2 ms and APD(90)=54 +/- 5 ms, n=10) compared to males (APD(50)=11 =/- 2 ms and APD(90)=47 +/- 7 ms, n=10). FK506 treatment (25 microM) selectively and equally inhibited I(sus) in all cells, and significantly increased APD(50) and APD(90) in males and females at 0.3 and 1.0 Hz. However, increases in APD(50) and APD(90) (0.3 and 1.0 Hz) in response to FK506 were significantly greater in myocytes from females compared to males. Voltage clamp measurement of I(to) and I(sus) revealed that males had a relatively more prominent I(to) while females exhibited a more prominent I(sus). Ventricular action potential repolarization is prolonged in myocytes from female compared to male mice. Female mice have reduced RR that is unmasked by FK506. These findings suggest that gender is an important variable for cardiovascular studies using mice.

Familial and acquired long qt syndrome and the cardiac rapid delayed rectifier potassium current.

1. Long QT syndrome (LQTS) is a cardiac disorder characterized by syncope, seizures and sudden death; it can be congenital, idiopathic, or iatrogenic. 2. Long QT syndrome is so-named because of the connection observed between the distinctive polymorphic ventricular tachycardia torsade de pointes and prolongation of the QT interval of the electrocardiogram, reflecting abnormally slowed ventricular action potential (AP) repolarization. Acquired LQTS has many similar clinical features to congenital LQTS, but typically affects older individuals and is often associated with specific pharmacological agents. 3. A growing number of drugs associated with QT prolongation and its concomitant risks of arrhythmia and sudden death have been shown to block the 'rapid' cardiac delayed rectifier potassium current (IKr) or cloned channels encoded by the human ether-a-go-go-related gene (HERG; the gene believed to encode native IKr). Because IKr plays an important role in ventricular AP repolarization, its inhibition would be expected to result in prolongation of both the AP and QT interval of the electrocardiogram. 4. The drugs that produce acquired LQTS are structurally heterogeneous, including anti-arrhythmics, such as quinidine, non-sedating antihistamines, such as terfenadine, and psychiatric drugs, such as haloperidol. In addition to heterogeneity in their structure, the electrophysiological characteristics of HERG/IKr inhibition differ between agents. 5. Here, clinical observations are associated with cellular data to correlate acquired LQTS with the IKr/HERG potassium (K+) channel. One strategy for developing improved compounds in those drug classes that are currently associated with LQTS could be to design drug structures that preserve clinical efficacy but are modified to avoid pharmacological interactions with IKr. Until such time, awareness of the QT-prolongation risk of particular agents is important for the clinician.