Effects of chromanol 293B on transient outward and ultra-rapid delayed rectifier potassium currents in human atrial myocytes

It is unclear whether chromanol 293B, a selective inhibitor of slow component of delayed rectifier K(+) current (I(Ks)), may affect other K(+) currents in human atrium. With whole-cell patch configuration, we evaluated effects of 293B on transient outward K(+) current (I(to1)) and ultra-rapid delayed rectifier K(+) current (I(Kur)) in isolated human atrial myocytes. It was found that 293B inhibited I(to1) and I(Kur) in a concentration-dependent manner. At 10 microM 293B suppressed I(to1) to 3.4 +/- 0.4 from 5.1 +/- 0.3 pA/pF (P < 0.01), and I(Kur) to 1.5 +/- 0.2 from 2.1 +/- 0.3 pA/pF (P < 0.01) at +50 mV. The inhibition of I(to1) and I(Kur) was independent of depolarizing voltage, and the concentration of 50% inhibition was 31.2 microM for I(to1), and 30.9 microM for I(Kur). 293B blocked I(to1) and I(Kur) with the same concentration range, and the significant effect was observed from the concentration of 1 microM. The maximum inhibitive effect was 88% for I(to1) and 96% for I(Kur) at 250 microM. Voltage dependence of activation and inactivation, and time-dependent recovery from inactivation of I(to1) were not altered by 293B; however, time to peak and time-dependent inactivation of I(to1) was significantly accelerated. The results indicate that 293B significantly inhibits the major repolarization K(+) currents I(to1) and I(Kur) in human atrial myocytes.     

Demonstration of calcium-activated transient outward chloride current and delayed rectifier potassium currents in Swine atrial myocytes

Cellular electrophysiology is not fully understood in the atrium of pig heart. The objective of the present study was to determine whether transient outward current (I(to)), ultra-rapid delayed rectifier potassium current (I(Kur)), and rapid and slow delayed rectifier K(+) currents (I(Kr) and I(Ks)) were present in pig atrium. The whole-cell patch technique was applied to record membrane currents and action potentials in myocytes isolated from pig atrium. It was found that an I(to) was activated upon depolarization voltage steps to between -10 and +60 mV from -50 mV in pig atrial cells, and the I(to) was sensitive to the inhibition by the blockade of L-type calcium (Ca(2+)) current, showed a "bell-shaped" I-V relationship, typical of I(to2) (i.e. I(Cl.Ca)). The I(to2) was inhibited by the chloride (Cl(-)) channel blocker anthracene-9-carboxylic acid (9-AC, 200 micromol/l) or 4,4'-diisothiocyanostilben-2,2'disulfonic acid (200 micromol/l), and by Cl(-) substitution in the superfusate. I(Kur) was found in pig atrial myocytes, and the current showed properties of weak inward rectification and use- and frequency-dependent reduction. I(Kur) was resistant to tetraethylammonium, but sensitive to inhibition by 4-aminopyridine (4-AP) (IC(50) = 71.7 +/- 3.5 micromol/l). In addition, E-4031-sensitive I(Kr) and chromanol 293B-sensitive I(Ks) were observed in pig atrial myocytes. Blockade of I(to2), I(Kur), I(Kr) or I(Ks) with corresponding blockers significantly prolonged atrial action potentials. These results indicate that Ca(2+)-activated I(to2), 4-AP-sensitive I(Kur), E-4031-sensitive I(Kr), and 293B-sensitive I(Ks) are present in pig atrial myocytes, and these currents play important roles in action potential repolarization of pig atria.     

Postnatal development of atrial repolarization in the mouse

OBJECTIVES AND METHODS: This study examines postnatal development of action potential duration (APD) and voltage-dependent K(+) currents in mouse atrial myocytes and compares the expression levels of corresponding K(+) channels between adult and neonatal mouse atrial tissues. APD and K(+) currents were compared between atrial myocytes isolated from postnatal Day-1, Day-7, Day-20, and adult mice. RESULTS: All K(+) currents examined underwent significant up-regulation during postnatal life in mouse atrium, resulting in a dramatic shortening of the APD. The ultrarapid delayed rectifier (I(Kur)) was absent in the developing mouse heart and only contributed to repolarization in the adult mouse atrium, whereas the density of the other K(+) currents increased earlier during the developmental period. Indeed, the major changes in the expression of the inward rectifier current (I(K1)) occurred within the first week of life, the density of the Ca(2+)-independent transient outward K(+) current (I(to)) gradually increased while the development of the steady-state outward K(+) current (I(ss)) was completed within the first 3 weeks of life. Results of RNase protection assay and Western blot analysis confirmed that the postnatal development of the mouse atrial K(+) currents correlates with an increase in expression levels of underlying K(+) channel isoforms. CONCLUSION: These findings indicate that in mouse atrium, each K(+) current exhibits a specific postnatal development, suggesting that regulatory factors taking place during development are major determinants of the functional role of K(+) channels in cardiac repolarization. The mouse atrium is, therefore, a very interesting model to gain information on the mechanisms regulating K(+) channel activity.     

Characterization of ventricular repolarization in male and female guinea pigs

Since both components of the delayed rectifier K(+) currents (I(Kr) and I(Ks)) are present in guinea pig and human ventricle, the guinea pig appears as an interesting model to examine the contribution of these currents in sex-related difference of cardiac repolarization. Accordingly, we compared ventricular repolarization in adult male and female guinea pigs using electrophysiological protocols together with Western blots analysis and perfused heart preparation. Our results indicate that there was no sex-related difference in the expression levels of the different K(+) channels studied (ERG, KvLQT1, minK and Kir2.1), nor in the density of the K(+) currents (I(Kr), I(Ks) and I(K1)) encoded by these channels. Action potential durations and QTc intervals were also similar between males and females. In addition, we compared QTc intervals using Langendorff-perfused whole hearts in the presence of I(Kr) and/or I(Ks) blockers. The I(Kr) blocker (5 microM E-4031) prolonged QTc intervals to a similar extent in male (24+/-2%) and female (29+/-3%, p=0.1) hearts. Similarly, the degree of QTc prolongation induced by 0.1 microM HMR1556 (I(Ks) blocker) was similar in both sexes (males: 15+/-2% and females 18+/-2%; p=0.2). In addition to their QT prolonging effects, the I(Kr) and I(Ks) blockers significantly reduced heart rate in both male and female guinea pigs. These studies clearly demonstrate that adult guinea pigs do not display sex differences in ventricular repolarization.     

Sarcoplasmic reticulum K(+) channels from human and sheep atrial cells display a specific electro-pharmacological profile

It has recently been proposed that the Ca(2+) uptake by the SR is inhibited by blocking Cl(-) and/or K(+) movements across this intracellular membrane. We have characterised the functional and pharmacological profile of the SR K(+) channel derived from human and sheep atrial cells. Mammalian atrial SR preparations were subjected to [(3)H]-ryanodine binding assays, SDS-PAGE analysis and channel protein reconstitution into planar lipid bilayers. Assessment of [(3)H]-ryanodine binding on the SR Ca(2+) release channel revealed that it was inhibited by both Ruthenium Red and Mg(2+) with IC(50) values of 4.11 microM and 9.12 m M, respectively. In crude populations as well as in all SR-enriched fractions, activity of K(+) selective channels was recorded. This channel displayed a high conductance value of 193 and 185 pS for human and sheep preparations respectively. Gating and conducting behaviours of this channel were unaffected by the addition of up to 5m M 4-Aminopyridine (4-AP), 100 n M Iberiotoxin (IbTX), 10 microM E-4031 and 30 microM amiodarone. However, 100n M Dendrotoxin (gamma-DTX) largely increase the occurrence of the SR K(+) channel subconducting states without an effect on the main unitary conductance. These results demonstrate that the SR K(+) channel, present in all mammalian atrial SR membranes tested (as assessed by [(3)H]-ryanodine binding and its typical inhibition by ruthenium red and the magnesium), displays different properties than those classically described for cardiac sarcolemmal K(+) channels. Despite the fact that the biophysical properties of the SR K(+) channel are well known, its molecular identity remains to be ascertained.     

Comparison of K+ currents in cardiac Purkinje cells isolated from rabbit and dog

The repolarization reserve determines the ability of drugs to prolong the cardiac action potential duration. Differences in K(+) currents between rabbit and dog cardiac Purkinje cells were studied by recording the transient outward K(+) current (I(to)) as well as the delayed rectifier K(+) currents (I(Ks) and I(Kr)) during repolarization. Purkinje fibers were dissected from dog and rabbit hearts and exposed to enzymatic digestion until isolated cells were obtained. Whole cell voltage clamp methods were used to measure K(+) currents in both cell types. Action potential (AP) recordings from Purkinje cells displayed a rapid phase 1 repolarization due to a prominent I(to) with densities of 13.3+/-2.3 and 9.6+/-0.6 pA/pF at +40 mV in dog and rabbit respectively. I(Ks) tail currents were significantly larger in dog Purkinje cells. I(Kr) tail current densities were comparable in Purkinje cell from both species. Rabbit ventricular and Purkinje cell AP waveforms were used for action potential clamp experiments in TSA201 cells expressing human ether a go-go related gene (HERG). HERG currents elicited by the ventricular waveform reached its maximum amplitude during phase 3 repolarization. In contrast, Purkinje cell AP waveform elicited markedly smaller HERG currents even though the action potential duration was longer. The observations suggest that the fast phase 1 and negative plateau of the Purkinje cell AP limits the contribution of I(Kr) to repolarization. These results provide evidence that rabbit Purkinje cells have a smaller repolarization reserve and provide a biophysical explanation for a previously observed higher sensitivity to QT prolonging drugs in rabbit than dog Purkinje fibers.     

The ultrarapid and the transient outward K(+) current in human atrial fibrillation. Their possible role in postoperative atrial fibrillation

Atrial fibrillation (AF) causes distinct changes in atrial conduction, characterized as electrical remodeling. Experimental data on the possible significance of alterations of specific K(+)outward currents in this process are still limited in human AF. The ultra-rapid delayed rectifier current (I(Kur)) has not been studied in AF with respect to its sensitivity to 4-Aminopyridine (4-AP). To clarify the role of (1) the 4-AP sensitive I(Kur)current, compared to recordings without using 4-AP (I(Kur*)), and (2) the transient outward current (I(to)) in changes of atrial repolarization associated with AF, whole cell voltage-clamp recordings were obtained from atrial myocytes of patients undergoing elective cardiac surgery, with and without a history of atrial fibrillation (AF/non-AF). Further, a possible relation between experimental data and postoperative AF was studied. In AF patients, I(Kur*)was reduced by 40% [5.00+/-0.32 pA/pF (non-AF) and 2.91+/-0. 45 pA/pF (AF) at +50 mV, P<0.0001, n=22/11], I(Kur)by 55% [3.81+/-0. 30 pA/pF (non-AF) and 1.71+/-0.20 pA/pF (AF) at +50 mV, P<0.0001, n=22/11]. The mean amplitude of I(Kur)was significantly smaller than I(Kur*). Consistently, I(to)was reduced by 44% [11.57+/-0.77 pA/pF (non-AF) and 6.51+/-1.31 pA/pF (AF), P<0.01, n=25/11]. In 48% of non-AF patients, postoperative AF was detected. The corresponding voltage-clamp recordings showed a trend to reduced I(Kur*)and I(Kur)currents, although it did not reach statistical significance. The consistent reduction of all three K(+)currents investigated due to the presence of AF indicates an important association of abnormalities in cellular repolarization with the onset and the self-sustaining nature of human AF.     

Four and a half LIM protein 1: a partner for KCNA5 in human atrium

AIMS: Protein-protein interactions are critical for the normal membrane trafficking, localization, and function of voltage-gated ion channels. In human heart, the Shaker-related voltage-gated K(+) channel KCNA5 alpha-subunit forms the major basis of an atrial-specific, ultra-rapid delayed rectifier K(+) current, I(Kur). We sought to identify proteins that interact with KCNA5 in human atrium and investigate their role in the I(Kur) complex. METHODS AND RESULTS: Using a glutathione-S-transferase (GST)-KCNA5 C-terminal fusion protein and mass spectrometry-based methods, the scaffolding protein four and a half LIM (for Lin-11, Isl-1, and Mec3) protein 1 (FHL1) was identified as a potential protein partner for KCNA5. Immunoprecipitation experiments confirmed a physical interaction of FHL1 with the K(+) channel complex in human atrium, as well as in Chinese hamster ovary (CHO) cells transfected with both KCNA5 and FHL1. In cotransfected cells, confocal microscopy demonstrated areas of colocalization after immunolabelling both proteins. To investigate the functional effects of this interaction, K(+) currents were recorded in CHO cells transfected with KCNA5 in the absence and presence of FHL1 coexpression. With coexpression of FHL1, K(+) current density was markedly increased, compared with cells expressing KCNA5 alone. This effect was associated with a shift in the voltage dependence of K(+) channel activation to more positive potentials, consistent with findings of I(Kur) in atrial myocytes. FHL1 also increased the extent and speed of K(+) current slow inactivation, with additional effects on the voltage dependence and recovery of this process. CONCLUSION: These results support a role of FHL1 as a key molecular component in the I(Kur) complex in human atrium, where it likely regulates functional expression of KCNA5.     

Effects of ambasilide, quinidine, flecainide and verapamil on ultra-rapid delayed rectifier potassium currents in canine atrial myocytes

OBJECTIVE: A dog atrial ultra-rapid delayed rectifier current (I(Kur. d)) is involved in canine atrial repolarization and shares similarities with the human atrial ultra-rapid delayed rectifier (I(Kur)). Almost no information is available about the actions of antiarrhythmic drugs on I(Kur.d). This study evaluated effects of ambasilide, quinidine, flecainide and verapamil on I(Kur.d) in isolated canine atrial myocytes. METHODS: Standard whole-cell patch clamp techniques were used to study the effects of multiple concentrations of each drug. RESULTS: All drugs produced reversible concentration-, voltage- and time-dependent I(Kur.d) inhibition. Significant effects of quinidine, flecainide and ambasilide were noted at atrial-effective antiarrhythmic concentrations in the dog. Upon the onset of a depolarizing pulse, block developed exponentially in relation to time, with the blocking rate-constant increasing with drug concentration, consistent with open-channel blockade and permitting the calculation of forward and reverse rate-constants. For all drugs, the 50% blocking concentration (EC(50)) showed significant voltage-dependence, decreasing at more positive potentials. The magnitude of voltage-dependent block was directly related to the degree of drug-induced shift in the voltage dependence of activation (r=0.97), pointing to open-channel block as a mechanism for voltage-dependent action. An additional component of voltage-dependence suggested that blocking sites were subjected to 17-21% of the transmembrane voltage field. CONCLUSIONS: Ambasilide, quinidine, flecainide and verapamil inhibit I(Kur.d), with preferential action on the open state. I(Kur.d) inhibition may play a role in antiarrhythmic effects in canine atrial arrhythmia models. Comparisons between the effects of these drugs on I(Kur.d) and previously studied effects on I(Kur) suggest potential opportunities for investigating the molecular structural determinants of drug-blocking action on atrial-specific ultrarapid delayed rectifiers.     

Structural determinants and biophysical properties of HERG and KCNQ1 channel gating

The delayed rectifier K(+) currents, I(Kr) and I(Ks,) play a critical role in modulating the plateau phase of the cardiac action potential. HERG encodes the alpha-subunit of channels underlying I(Kr), while I(Ks) is composed of subunits encoded by KCNQ1 and KCNE1. Mutations in any of these genes cause the long QT syndrome, a disorder of myocellular repolarization that predisposes affected individuals to life-threatening arrhythmias. Elucidation of the molecular basis of these currents has led to significant advancements in our understanding of fundamental properties of channel function. This review summarizes the current state of knowledge regarding the structural determinants and biophysical properties of HERG and KCNQ1 channels.     

Atrial effects of the novel K(+)-channel-blocker AVE0118 in anesthetized pigs

OBJECTIVES: AVE0118 is a novel blocker of the K(+) channels K(v)1.5 and K(v)4.3 which are the molecular basis for the human cardiac ultrarapid delayed rectifier potassium current (I(Kur)) and the transient outward current (I(to)). The objective of this study was to investigate the effect of AVE0118 on atrial refractoriness (ERP), left atrial vulnerability (LAV) and on left atrial monophasic action potentials (MAP) in pentobarbital anesthetized pigs in comparison to the selective I(Kr) blocker dofetilide in order to assess the therapeutic potential of the novel K(+) channel blocker for atrial fibrillation. METHODS: Atrial ERP was determined with the S1-S2-stimulus method in the free walls of left and right atrium at 240, 300 and 400 ms basic cycle length (BCL). The inducibility of mostly nonsustained atrial tachyarrhythmias by the premature S2 extrastimulus, which is very high in the left pig atrium and referred to as LAV, was evaluated before and after drugs. Left atrial epicardial MAP was recorded to study the influence of the potassium channel blockers on the time course of repolarization. Left ventricular epicardial MAP, ERP and QT interval were measured to investigate a possible effect of AVE0118 on ventricular repolarization. RESULTS: ERPs determined at 240, 300 and 400 ms BCL were significantly shorter in the left vs. right atrium (99+/-3, 106+/-4 and 113+/-3 ms vs. 133+/-4 ms, 142+/-4 and 149+/-5, respectively; p<0.001; n=21). AVE0118 administered i.v. dose-dependently prolonged the atrial ERP independent from rate and inhibited LAV (100% at 0.5 and 1 mg/kg) while having no effect at all on the corrected QT (QTc) interval. At 1 mg/kg (n=5) AVE0118 prolonged left vs. right atrial ERP by 49.6+/-4.1 ms vs. 37.7+/-9.7 ms (means+/-SEM of changes at 240, 300, and 400 ms BCL), respectively, corresponding to a relative increase of 53.2+/-6.2% vs. 27.6+/-6.8% (p<0.05 for percent increase of left vs. right atrial ERP). In a separate group of pigs (n=5) AVE0118 had no effect on left ventricular ERP at 333, 400 and 500 ms BCL and no effect on MAP duration and QT at 600 ms BCL. After 1 mg/kg of AVE0118 the atrial MAP was significantly prolonged already at 10% repolarization (P<0.05; n=7) reaching the maximum at 40% repolarization. In contrast to AVE0118 the effect of dofetilide (10 microg/kg) on atrial MAP started to become significant only at 60% repolarization (n=6) with a maximum increase at 90%. Dofetilide, which prolonged the QTc interval by 16.9% (P<0.001), had a significantly stronger effect on right (34.7+/-5 ms) vs. left atrial ERP (23.5+/-7 ms) at 300 ms BCL, respectively, but did not significantly inhibit LAV (14%; n=6). CONCLUSION: The novel K(+) channel blocker AVE0118 prolonged atrial ERP and showed strong atrial antiarrhythmic efficacy with no apparent effect on ventricular repolarization in pigs in vivo.     

Ghrelin reduces voltage-gated potassium currents in GH3 cells via cyclic GMP pathways

Ghrelin is an endogeneous growth hormone secretagogue (GHS) causing release of GH from pituitary somatotropes through the GHS receptor. Secretion of GH is linked directly to intracellular free Ca²⁺ concentration ([Ca²⁺]_i), which is determined by Ca²⁺ influx and release from intracellular Ca²⁺ storage sites. Ca²⁺ influx is via voltage-gated Ca²⁺ channels, which are activated by cell depolarization. Membrane potential is mainly determined by transmembrane K⁺ channels. The present study investigates the in vitro effect of ghrelin on membrane voltage-gated K⁺ channels in the GH₃ rat somatotrope cell line. Nystatin-perforated patch clamp recording was used to record K⁺ currents under voltage-clamp conditions. In the presence of Co²⁺ (1 mM, Ca²⁺ channel blocker) and tetrodotoxin (1 μM, Na⁺ channel blocker) in the bath solution, two types of voltage-gated K⁺ currents were characterized on the basis of their biophysical kinetics and pharmacological properties. We observed that transient K⁺ current (I _A) represented a significant proportion of total K⁺ currents in some cells, whereas delayed rectifier K⁺ current (I _K) existed in all cells. The application of ghrelin (10 nM) reversibly and significantly decreased the amplitude of both I _A and I _K currents to 48% and 64% of control, respectively. Application of apamin (1 μM, SK channel blocker) or charybdotoxin (1 μM, BK channel blocker) did not alter the K⁺ current or the response to ghrelin. The ghrelin-induced reduction in K⁺ currents was not affected by PKC and PKA inhibitors. KT5823, a specific PKG inhibitor, totally abolished the K⁺ current response to ghrelin. These results suggest that ghrelininduced reduction of voltage-gated K⁺ currents in GH₃ cells is mediated through a PKG-dependent pathway. A decrease in voltage-gated K⁺ currents may increase the frequency, duration, and amplitude of action potentials and contribute to GH secretion from somatotropes.     

IKs, a slow and intriguing cardiac K+ channel and its associated long QT diseases

Shaping of cardiac action potentials depends on a finely tuned orchestra of ion channels. Among them, K(+) channels probably form the most diverse family. They are responsible for inwardly rectifying (I(K1), I(KAch), I(KATP)), transient (I(to)), and sustained outward rectifying (I(Kur), I(Kr), I(Ks)) K(+) currents. The properties of these cardiac K(+) channels have recently been extensively reviewed. This article focuses on recent progress made toward understanding the molecular structure of the particular channel responsible for the slow outward K(+) current I(Ks) and its implication in the delayed ventricular repolarization that characterizes the congenital long QT syndrome.     

Pharmacological modulation of the hyperpolarization-activated current (I f) in human atrial myocytes: focus on G protein-coupled receptors

AIMS: In human atrial myocytes (HuAM) two beta-adrenergic receptors (beta-AR) and four splicing-variants of the serotonin 5-HT(4) receptor are present. Multiple coupling with G stimulatory (G(s)) and G inhibitory (G(i)) proteins has been proposed for both beta(2)-AR and 5-HT((4b)) subtypes, but no functional data exist in HuAM. Serotonin (5-HT) and catecholamines are able to trigger arrhythmias in human atrium, but the underlying cellular mechanisms are not completely understood. The pacemaker current (I(f)) is an inward Na(+)/K(+) current, constitutively present in HuAM and directly modulated by cAMP; I(f) could play a role in triggering human atrial arrhythmias. This study evaluated the different G protein coupling of beta(1)-AR, beta(2)-AR and 5-HT(4) receptors by assessing the modulation of I(f) by selective stimuli. METHODS: HuAM were isolated from right atrial appendages and utilized for patch-clamp recording. The coupling of receptor subtypes with G(i) proteins was tested by incubating HuAM in pertussis toxin (PTX). RESULTS: Beta(1)-AR stimulation (Isoprenaline [ISO] + ICI 118,551), and 5-HT caused a concentration-dependent significant shift of the half activation potential of I(f) activation curve (DeltaV(h)), P < 0.01. beta(2)-AR stimulation (ISO 1 microM + CGP 20712A) also significantly shifted V(h) (P < 0.0001), but with DeltaV(h)[beta(2)-AR] significantly smaller than the effect caused by 1 microM beta(1)-AR stimulation (P < 0.05). Pre-treatment of HuAM with PTX did not alter the effect of beta(1)-AR stimulation (both 0.1 and 1 microM) and 1 microM 5-HT on I(f), but significantly increased the effect in response to beta(2)-AR stimulation and 0.1 microM 5-HT (P < 0.05 for both), thus suggesting a G(i) protein coupling of these receptors. CONCLUSIONS: Our results provide the first functional evidence of the different G protein coupling of beta(1)-AR, beta(2)-AR and 5-HT(4) receptors in HuAM. Further they support the view that I(f) current might play an important role in triggering catecholamines and serotonin-induced atrial arrhythmias.     

Rapid stimulation causes electrical remodeling in cultured atrial myocytes

OBJECTIVE: Rapid stimulation causes electrical remodeling in the intact atrium, with shortening of action potential duration (APD), down-regulation of L-type Ca2+ currents (I(Ca,L)), and increased vulnerability to atrial fibrillation (AF). The essential elements required for this process are currently unknown. We tested the hypothesis that rapid stimulation of cardiomyocytes in vitro is sufficient to recapitulate the remodeling process, and that atrial cells subjected to rapid pacing in culture would display changes similar to those that occur in vivo. METHODS: Atrial (HL-1) cells were cultured in the presence of rapid field stimulation (300 beats per min) for 24 h. Action potentials and ionic currents were recorded from stimulated cells, as well as control cells cultured in parallel, using whole-cell voltage-clamp techniques. RESULTS: Rapid stimulation of atrial cells for 24 h significantly shortened APD. HL-1 cells displayed both I(Ca,L) blocked by nimodipine, and T-type Ca2+ currents (I(Ca,T)) sensitive to mibefradil. Rapid activation in culture caused down-regulation of I(Ca,L), while I(Ca,T) was similarly reduced. Multiple outward currents were present in response to a depolarizing voltage-clamp protocol, and rapid pacing resulted in up-regulation of the rapidly-activating delayed rectifier K+ current, I(Kr). CONCLUSIONS: Rapid stimulation of atrial cells in culture produces electrical remodeling, recapitulating principal phenotypic features of atrial tachycardia remodeling in vivo. Our results demonstrate that an important component of this process is cell autonomous, given that in vivo conditions are not required for the development of electrical remodeling.     

Ultra-rapid delayed rectifier channels: molecular basis and therapeutic implications

The ultrarapid delayed rectifier channels have attracted considerable interest as targets for 'atrial-selective' antiarrhythmic drugs because they contribute to atrial but not to ventricular repolarization. Thus, I(Kur) channel blockers are expected to prolong selectively the atrial effective refractory period without inducing proarrhythmic effects due to excessive ventricular action potential prolongation. Here we provide an overview of the properties of I(Kur) channels in expression systems and native cardiomyocytes. The ion conducting pore of the channel is formed by four Kv1.5 α-subunits, whereas the ancillary β-subunits Kvβ1.2, Kvβ1.3, and Kvβ2.1 control channel trafficking and plasma membrane integration as well as activation and inactivation kinetics. Investigation of I(Kur) channel blockers in cardiomyocytes is complicated (i) by substantial overlap of I(Kur) with other currents, notably the transient outward current I(to), (ii) by lack of drug selectivity, and (iii) by disease-induced regulation of I(Kur). Some new compounds developed as I(Kur) blockers are described and their efficacy in treatment of atrial fibrillation (AF) is discussed. Current evidence suggests that pure I(Kur) channel block may not be sufficient to suppress AF.     

Gender-based differences in cardiac repolarization in mouse ventricle

The mouse heart has become a widely used model for genetic studies of heart diseases. Thus, understanding gender differences in mouse cardiac repolarization is crucial to the interpretation of such studies. The objective of this study was to evaluate whether there are gender differences in cardiac repolarization in mouse ventricle and to gain insights into the ionic and molecular mechanisms underlying these differences. Action potential durations (APDs) and K(+) currents in male and female ventricular myocytes were compared using a patch-clamp technique. APD(20), APD(50), and APD(90) were found to be significantly longer in females than males. Examination of the different K(+) currents revealed that a significantly lower current density exists in female ventricular myocytes compared with male myocytes for the ultrarapid delayed rectifier K(+) current, I(Kur) (at +30 mV, male, 33.2+/-2.9 pA/pF [n= 22]; female, 20.9+/-1.73 pA/pF [n= 19], P<0.001). Consistent with these findings were the results of the ribonuclease protection assay, Western blots, and confocal analysis that showed a significantly lower expression level of Kv1.5 (coding for I(Kur)) in female compared with male ventricle. The additional K(+) currents present in mouse ventricle exhibited no gender differences. In agreement with these electrophysiological data, no differences in the expression levels for the K(+) channels underlying these currents were detected between both sexes. This study demonstrates that adult mice exhibit gender differences in cardiac repolarization. The expression of Kv1.5 and of its corresponding K(+) current, I(Kur), is significantly lower in female mouse ventricle, and as a result, the APD is lengthened.     

Heterogeneity of sodium current in atrial vs epicardial ventricular myocytes of adult guinea pig hearts

The different sodium channel currents (I(Na)) were reported in myocardium, neuron, and skeletal muscles. To study whether I(Na) is homogeneous within the heart, we applied whole-cell voltage clamp technique to evaluate fast voltage-gated I(Na) in atrial and ventricular myocytes isolated from guinea pig heart. It was found that the density of inward I(Na) was 50% greater at -35 mV in atrial (-42.6+/-2.9 pA/pF) than in ventricular (-27.5+/-1.8 pA/pF, P<0.01) myocytes. The half activation and inactivation voltages (V(0.5)) of I(Na) in atrial myocytes were shifted 4.5+/-0.2 and 9.6+/-0.3 mV negative to those of ventricular myocytes. Time constants for I(Na) activation (tau(m)) and inactivation (tau(h)) were twice as rapid in atrial as in ventricular myocytes. The tau(m) and tau(h) were 0.34+/-0.03 and 1.36+/-0.07 ms for atrial myocytes, and 0.69+/-0.05 and 3.27+/-0.23 ms for ventricular myocytes, respectively. Recovery of I(Na) from inactivation was slower in atrial than in ventricular myocytes, whereas the development of resting state inactivation was more rapid in atrial (tau=67.5+/-4.3 ms) than in ventricular (152.8+/-7.5 ms, P<0.01) myocytes. The results reveal marked heterogeneity of I(Na) in the density and biophysical properties in atrial and ventricular myocytes, and the study suggests the potential possibility of tissue specific cardiac sodium channel isoforms.     

Sex and strain differences in adult mouse cardiac repolarization: importance of androgens

OBJECTIVE: Gender differences in mouse cardiac repolarization have been reported to be due to the stimulatory action of androgens on the ultrarapid delayed rectifier K(+) current (I(Kur)) and its underlying Kv1.5 channel. To confirm the regulation of ventricular repolarization by androgens, the present study compared two strains of mice (CD-1 and C57BL/6) that present different androgen levels. METHODS AND RESULTS: Measurement of testosterone levels in different strains of mice (CD-1, C57BL/6, C3H and FVB) revealed that male C57BL/6 mice had very low levels of testosterone, whereas males of the other strains displayed normal testosterone levels. Furthermore, whole-cell voltage clamp recordings in isolated ventricular myocytes showed that the current density of I(Kur) in male C57BL/6 mice was similar to that in female mice but smaller with respect to male CD-1 mice. Androgen replacement in male C57BL/6 mice as well as in castrated male CD-1 mice shortened ventricular repolarization, increased I(Kur) current density, and increased expression of Kv1.5 channels. CONCLUSION: Strain and gender differences observed in mouse cardiac repolarization can be explained by different androgen levels. As a consequence, androgens are major regulatory factors in cardiac repolarization and special attention should be paid to the hormonal status of the animal when studying hormonal regulation of cardiac repolarization.