Potassium channels encoded by human ether‐à‐go‐go‐related gene (hERG) mediate the cardiac rapid delayed rectifier K+ current (IKr), which participates in ventricular repolarization and has a protective role against unwanted premature stimuli late in repolarization and early in diastole. Ionic current carried by hERG channels (IhERG) is known to exhibit a paradoxical dependence on external potassium concentration ([K+]e), but effects of acute [K+]e changes on the response of IhERG to premature stimulation have not been characterized. Whole‐cell patch‐clamp measurements of hERG current were made at 37°C from hERG channels expressed in HEK293 cells. Under conventional voltage‐clamp, both wild‐type (WT) and S624A pore‐mutant IhERG during depolarization to +20 mV and subsequent repolarization to −40 mV were decreased when superfusate [K+]e was decreased from 4 to 1 mmol/L. When [K+]e was increased from 4 to 10 mmol/L, pulse current was increased and tail IhERG was decreased. Increasing [K+]e produced a +10 mV shift in voltage‐dependent inactivation of WT IhERG and slowed inactivation time course, while lowering [K+]e from 4 to 1 mmol/L produced little change in inactivation voltage dependence, but accelerated inactivation time course. Under action potential (AP) voltage‐clamp, lowering [K+]e reduced the amplitude of IhERG during the AP and suppressed the maximal IhERG response to premature stimuli. Raising [K+]e increased IhERG early during the AP and augmented the IhERG response to premature stimuli. Our results are suggestive that during hypokalemia not only is the contribution of IKr to ventricular repolarization reduced but its ability to protect against unwanted premature stimuli also becomes impaired.
hERG potassium channels are important for ventricular repolarization and for protecting the ventricles of the heart from unwanted premature stimuli. This study shows that, in addition to reducing the contribution of hERG channel current to ventricular repolarization, hypokalemia impairs the protective response of hERG to premature stimulation.
Human ether‐à‐go‐go‐related gene; hyperkalemia; hypokalemia; long QT; potassium; potassium channels; QT interval
human Ether-à-go-go-Related Gene (hERG) encodes the pore-forming subunit of cardiac rapid delayed rectifier K+ current (IKr) channels, which play important roles in ventricular repolarization, in protecting the myocardium from unwanted premature stimuli, and in drug-induced Long QT Syndrome (LQTS). KCNE1, a small transmembrane protein, can coassemble with hERG. However, it is not known how KCNE1 variants influence the channel's response to premature stimuli or if they influence the sensitivity of hERG to pharmacological inhibition. Accordingly, whole-cell patch-clamp measurements of hERG current (IhERG) were made at 37°C from hERG channels coexpressed with either wild-type (WT) KCNE1 or with one of three KCNE1 variants (A8V, D76N, and D85N). Under both conventional voltage clamp and ventricular action potential (AP) clamp, the amplitude of IhERG was smaller for A8V, D76N, and D85N KCNE1 + hERG than for WT KCNE1 + hERG. Using paired AP commands, with the second AP waveform applied at varying time intervals following the first to mimic premature ventricular excitation, the response of IhERG carried by each KCNE1 variant was reduced compared to that with WT KCNE1 + hERG. The IhERG blocking potency of the antiarrhythmic drug quinidine was similar between WT KCNE1 and the three KCNE1 variants. However, the IhERG inhibitory potency of the antibiotic clarithromycin and of the prokinetic drug cisapride was altered by KCNE1 variants. These results demonstrate that naturally occurring KCNE1 variants can reduce the response of hERG channels to premature excitation and also alter the sensitivity of hERG channels to inhibition by some drugs linked to acquired LQTS.
Cardiac; cisapride; clarithromycin; hERG; KCNE1; Long QT; potassium channels; QT interval; quinidine
Rapid atrial arrhythmias such as atrial fibrillation (AF) predispose to ventricular arrhythmias, sudden cardiac death and stroke. Identifying the origin of atrial ectopic activity from the electrocardiogram (ECG) can help to diagnose the early onset of AF in a cost-effective manner. The complex and rapid atrial electrical activity during AF makes it difficult to obtain detailed information on atrial activation using the standard 12-lead ECG alone. Compared to conventional 12-lead ECG, more detailed ECG lead configurations may provide further information about spatio-temporal dynamics of the body surface potential (BSP) during atrial excitation. We apply a recently developed 3D human atrial model to simulate electrical activity during normal sinus rhythm and ectopic pacing. The atrial model is placed into a newly developed torso model which considers the presence of the lungs, liver and spinal cord. A boundary element method is used to compute the BSP resulting from atrial excitation. Elements of the torso mesh corresponding to the locations of the placement of the electrodes in the standard 12-lead and a more detailed 64-lead ECG configuration were selected. The ectopic focal activity was simulated at various origins across all the different regions of the atria. Simulated BSP maps during normal atrial excitation (i.e. sinoatrial node excitation) were compared to those observed experimentally (obtained from the 64-lead ECG system), showing a strong agreement between the evolution in time of the simulated and experimental data in the P-wave morphology of the ECG and dipole evolution. An algorithm to obtain the location of the stimulus from a 64-lead ECG system was developed. The algorithm presented had a success rate of 93%, meaning that it correctly identified the origin of atrial focus in 75/80 simulations, and involved a general approach relevant to any multi-lead ECG system. This represents a significant improvement over previously developed algorithms.
Ectopic activity is associated with multiple cardiac disorders and has been implicated in the initiation of self-sustaining re-entrant excitation. Identifying the presence and origin of ectopic activity may be vital in improving diagnosis and treatment of disorders such as atrial fibrillation, and has been the subject of multiple studies. The electrical activity of the heart can be non-invasively monitored through the electrocardiogram. However, the standard 12-lead electrocardiogram may not provide sufficient information to resolve the focus of ectopic activity satisfactorily and accurately; more detailed multi-lead electrocardiograms may provide more information to be able to produce an algorithm to locate the origin of ectopic activity. Using a 3D computational atria-torso model developed in our laboratory, we simulated the electrical activity of the atria under normal and different ectopic conditions. The model was first validated by comparison to experimental data, and then used to develop an algorithm to identify the location of atrial ectopic focus using a 64-lead electrocardiogram. The algorithm developed was able to identify the origin of atrial ectopic activity in 75/80 simulations, which is a significant improvement compared to previously developed algorithms. Furthermore, the study suggests that multi-lead electrocardiograms provide significant benefits over the standard 12-lead configuration.
Noradrenaline plays an important role in the modulation of atrial electrophysiology. However, the identity of the modulated channels, their mechanisms of modulation, and their role in the action potential remain unclear. This study aimed to investigate the noradrenergic modulation of an atrial steady-state outward current (IKss).
Methods and results
Rat atrial myocyte whole-cell currents were recorded at 36°C. Noradrenaline potently inhibited IKss (IC50 = 0.90 nM, 42.1 ± 4.3% at 1 µM, n = 7) and potentiated the L-type Ca2+ current (ICaL, EC50 = 136 nM, 205 ± 40% at 1 µM, n = 6). Noradrenaline-sensitive IKss was weakly voltage-dependent, time-independent, and potentiated by the arachidonic acid analogue, 5,8,11,14-eicosatetraynoic acid (EYTA; 10 µM), or by osmotically induced membrane stretch. Noise analysis revealed a unitary conductance of 8.4 ± 0.42 pS (n = 8). The biophysical/pharmacological properties of IKss indicate a TREK-like K+ channel. The effect of noradrenaline on IKss was abolished by combined β1-/β2-adrenoceptor antagonism (1 µM propranolol or 10 µM β1-selective atenolol and 100 nM β2-selective ICI-118,551 in combination), but not by β1- or β2-antagonist alone. The action of noradrenaline could be mimicked by β2-agonists (zinterol and fenoterol) in the presence of β1-antagonist. The action of noradrenaline on IKss, but not on ICaL, was abolished by pertussis toxin (PTX) treatment. The action of noradrenaline on ICaL was mediated by β1-adrenoceptors via a PTX-insensitive pathway. Noradrenaline prolonged APD30 by 52 ± 19% (n = 5; P < 0.05), and this effect was abolished by combined β1-/β2-antagonism, but not by atenolol alone.
Noradrenaline inhibits a rat atrial TREK-like K+ channel current via a PTX-sensitive mechanism involving co-operativity of β1-/β2-adrenoceptors that contributes to atrial APD prolongation.
Background K+ current; Beta-adrenoceptor; K2P channel; Steady-state outward current; Osmotic stretch; TREK-1; Arachidonic acid
Chronic atrial fibrillation (AF) is associated with structural and electrical remodelling in the atria, which are associated with a high recurrence of AF. Through biophysically detailed computer modelling, this study investigated mechanisms by which AF-induced electrical remodelling promotes and perpetuates AF. A family of Courtemanche–Ramirez–Nattel variant models of human atrial cell action potentials (APs), taking into account of intrinsic atrial electrophysiological properties, was modified to incorporate various experimental data sets on AF-induced changes of major ionic channel currents (ICaL, IKur, Ito, IK1, IKs, INaCa) and on intracellular Ca2+ handling. The single cell models for control and AF-remodelled conditions were incorporated into multicellular three-dimensional (3D) atrial tissue models. Effects of the AF-induced electrical remodelling were quantified as the changes of AP profile, AP duration (APD) and its dispersion across the atria, and the vulnerability of atrial tissue to the initiation of re-entry. The dynamic behaviour of re-entrant excitation waves in the 3D models was characterised. In our simulations, AF-induced electrical remodelling abbreviated atrial APD non-uniformly across the atria; this resulted in relatively short APDs co-existing with marked regional differences in the APD at junctions of the crista terminalis/pectinate muscle, pulmonary veins/left atrium. As a result, the measured tissue vulnerability to re-entry initiation at these tissue junctions was increased. The AF-induced electrical remodelling also stabilized and accelerated re-entrant excitation waves, leading to rapid and sustained re-entry. Under the AF-remodelled condition, re-entrant scroll waves in the 3D model degenerated into persistent and erratic wavelets, leading to fibrillation. In conclusion, realistic 3D atrial tissue models indicate that AF-induced electrical remodelling produces regionally heterogeneous and shortened APD; these respectively facilitate initiation and maintenance of re-entrant excitation waves.
The antianginal drug ranolazine, which combines inhibitory actions on rapid and sustained sodium currents with inhibition of the hERG/IKr potassium channel, shows promise as an antiarrhythmic agent. This study investigated the structural basis of hERG block by ranolazine, with lidocaine used as a low potency, structurally similar comparator. Recordings of hERG current (IhERG) were made from cell lines expressing wild-type (WT) or mutant hERG channels. Docking simulations were performed using homology models built on MthK and KvAP templates. In conventional voltage clamp, ranolazine inhibited IhERG with an IC50 of 8.03 μM; peak IhERG during ventricular action potential clamp was inhibited ~ 62% at 10 μM. The IC50 values for ranolazine inhibition of the S620T inactivation deficient and N588K attenuated inactivation mutants were respectively ~ 73-fold and ~ 15-fold that for WT IhERG. Mutations near the bottom of the selectivity filter (V625A, S624A, T623A) exhibited IC50s between ~ 8 and 19-fold that for WT IhERG, whilst the Y652A and F656A S6 mutations had IC50s ~ 22-fold and 53-fold WT controls. Low potency lidocaine was comparatively insensitive to both pore helix and S6 mutations, but was sensitive to direction of K+ flux and particularly to loss of inactivation, with an IC50 for S620T-hERG ~ 49-fold that for WT IhERG. Docking simulations indicated that the larger size of ranolazine gives it potential for a greater range of interactions with hERG pore side chains compared to lidocaine, in particular enabling interaction of its two aromatic groups with side chains of both Y652 and F656. The N588K mutation is responsible for the SQT1 variant of short QT syndrome and our data suggest that ranolazine is unlikely to be effective against IKr/hERG in SQT1 patients.
•hERG K+ channels regulate cardiac action potential repolarization.•The molecular basis of hERG block by ranolazine and structurally related lidocaine was studied.•S6 Y652A and F656A mutations affected greatly ranolazine but not lidocaine binding.•T623 and S624 residues may directly interact with ranolazine but not lidocaine.•N588K and S620T attenuated inactivation mutants had reduced sensitivity to both drugs.
Antiarrhythmic; Docking; hERG; Lidocaine; QT interval; Ranolazine
Acidosis affects the mechanical and electrical activity of mammalian hearts but comparatively little is known about its effects on the function of the atrio-ventricular node (AVN). In this study, the electrical activity of the epicardial surface of the left ventricle of isolated Langendorff-perfused rabbit hearts was examined using optical methods. Perfusion with hypercapnic Tyrode's solution (20% CO2, pH 6.7) increased the time of earliest activation (Tact) from 100.5 ± 7.9 to 166.1 ± 7.2 ms (n = 8) at a pacing cycle length (PCL) of 300 ms (37°C). Tact increased at shorter PCL, and the hypercapnic solution prolonged Tact further: at 150 ms PCL, Tact was prolonged from 131.0 ± 5.2 to 174.9 ± 16.3 ms. 2:1 AVN block was common at shorter cycle lengths. Atrial and ventricular conduction times were not significantly affected by the hypercapnic solution suggesting that the increased delay originated in the AVN. Isolated right atrial preparations were superfused with Tyrode's solutions at pH 7.4 (control), 6.8 and 6.3. Low pH prolonged the atrial-Hisian (AH) interval, the AVN effective and functional refractory periods and Wenckebach cycle length significantly. Complete AVN block occurred in 6 out of 9 preparations. Optical imaging of conduction at the AV junction revealed increased conduction delay in the region of the AVN, with less marked effects in atrial and ventricular tissue. Thus acidosis can dramatically prolong the AVN delay, and in combination with short cycle lengths, this can cause partial or complete AVN block and is therefore implicated in the development of brady-arrhythmias in conditions of local or systemic acidosis.
atrio-ventricular node; optical mapping; acidosis; right atrium; atrio-ventricular block; bradycardia
Recently, both the manufacturer of quetiapine and the US Food and Drug Administration warned healthcare providers and patients about quetiapine-induced QTc interval prolongation and torsade de pointes (TdP) when using this drug within the approved labeling.
We reviewed the case-report literature and found 12 case reports of QTc interval prolongation in the setting of quetiapine administration. There were no cases of quetiapine-induced TdP or sudden cardiac death (SCD) among patients using quetiapine appropriately and free of additional risk factors for QTc interval prolongation and TdP. Among the 12 case reports risk factors included female sex (nine cases), coadministration of a drug associated with QTc interval prolongation (eight cases), hypokalemia or hypomagnesemia (six cases) quetiapine overdose (five cases), cardiac problems (four cases), and coadministration of cytochrome P450 3A4 inhibitors (two cases). There were four cases of TdP. As drug-induced TdP is a rare event, prospective studies to evaluate the risk factors associated with QTc prolongation and TdP are difficult to design, would be very costly, and would require very large samples to capture TdP rather than its surrogate markers. Furthermore, conventional statistical methods may not apply to studies of TdP, which is rare and an ‘outlier’ manifestation of QTc prolongation. We urge drug manufacturers and regulatory agencies to periodically publish full case reports of psychotropic drug-induced QTc interval prolongation, TdP, and SCD so that clinicians and investigators may better understand the clinical implications of prescribing such drugs as quetiapine.
case reports; drug-induced QTc interval prolongation; quetiapine; risk factors; torsade de pointes
A 71-year-old woman collapsed while working as a grocery store cashier. CPR was performed and an AED revealed torsades de pointes (TdP). She was subsequently defibrillated resulting in restoration of sinus rhythm with a QTc interval of 544 msec. Further evaluation revealed a diagnosis of Takotsubo Cardiomyopathy (TCM) contributing to the development of a multifactorial acquired long QT syndrome (LQTS). The case highlights the role of TCM as a cause of LQTS in the setting of multiple risk factors including old age, female gender, hypokalemia, and treatment with QT prolonging medications. It also highlights the multifactorial nature of acquired LQTS and lends support to growing evidence of an association with TCM.
structurally and therapeutically diverse drugs interact with
the human heart K+ channel hERG by binding within the K+ permeation pathway of the open channel, leading to drug-induced
‘long QT syndrome’. Drug binding to hERG is often stabilized
by inactivation gating. In the absence of a crystal structure, hERG
pore homology models have been used to characterize drug interactions.
Here we assess potentially inactivated states of the bacterial K+ channel, KcsA, as templates for inactivated state hERG pore
models in the context of drug binding using computational docking.
Although Flexidock and GOLD docking produced low energy score poses
in the models tested, each method selected a MthK K+ channel-based
model over models based on the putative inactivated state KcsA structures
for each of the 9 drugs tested. The variety of docking poses found
indicates that an optimal arrangement for drug binding of aromatic
side chains in the hERG pore can be achieved in several different
configurations. This plasticity of the drug “binding site”
is likely to be a feature of the hERG inactivated state. The results
demonstrate that experimental data on specific drug interactions can
be used as structural constraints to assess and refine hERG homology
Mechanisms underlying the genesis of re-entrant substrate for the most common cardiac arrhythmia, atrial fibrillation (AF), are not well understood. In this study, we develop a multi-scale three-dimensional computational model that integrates cellular electrophysiology of the left atrium (LA) and pulmonary veins (PVs) with the respective tissue geometry and fibre orientation. The latter is reconstructed in unique detail from high-resolution (approx. 70 μm) contrast micro-computed tomography data. The model is used to explore the mechanisms of re-entry initiation and sustenance in the PV region, regarded as the primary source of high-frequency electrical activity in AF. Simulations of the three-dimensional model demonstrate that an initial break-down of normal electrical excitation wave-fronts can be caused by the electrical heterogeneity between the PVs and LA. High tissue anisotropy is then responsible for the slow conduction and generation of a re-entrant circuit near the PVs. Evidence of such circuits has been seen clinically in AF patients. Our computational study suggests that primarily the combination of electrical heterogeneity and conduction anisotropy between the PVs and LA tissues leads to the generation of a high-frequency (approx. 10 Hz) re-entrant source near the PV sleeves, thus providing new insights into the arrhythmogenic mechanisms of excitation waves underlying AF.
integrative modelling; cardiac arrhythmias; re-entrant waves
Erythromycin is a macrolide antibiotic that is widely used for various infections of the upper respiratory tract, skin, and soft tissue. Similar to other macrolides (clarithromycin, azithromycin), erythromycin has been linked to QTc interval prolongation and torsade de pointes (TdP) arrhythmia. We sought to identify factors that link to erythromycin-induced/associated QTc interval prolongation and TdP.
Methods and Results:
In a critical evaluation of case reports, we found 29 cases: 22 women and 7 men (age range 18–95 years). With both oral and intravenous erythromycin administration, there was no significant relationship between dose and QTc interval duration in these cases. Notably, all patients had severe illness. Other risk factors included female sex, older age, presence of heart disease, concomitant administration of either other QTc prolonging drugs or agents that were substrates for or inhibitors of CYP3A4. Most patients had at least two risk factors.
On the basis of case report evaluation, we believe that major risk factors for erythromycin-associated TdP are female sex, heart disease and old age, particularly against a background of severe illness. Coadministration of erythromycin with other drugs that inhibit or are metabolized by CYP3A4 or with QTc prolonging drugs should be avoided in this setting.
drug-induced QTc interval prolongation; erythromycin; risk factors; torsade de pointes
Exudative AMD (wet AMD) is treated by monthly injection into the eye of anti-VEGF proteins. VEGF is alternatively spliced to produce numerous isoforms that differ in angiogenic activity. Serine-rich protein kinase-1 (SRPK1) has been identified as a regulator of pro-angiogenic VEGF splicing by phosphorylating serine-rich splicing factor-1 (SRSF1), which binds to VEGF pre-mRNA. We tested the hypothesis that topical (eye drop) SRPK1-selective inhibitors could be generated that reduce pro-angiogenic isoforms, and prevent choroidal neovascularization in vivo.
Novel inhibitors were tested for SRPK inhibition in vitro, pro-angiogenic VEGF production in RPE cells by PCR and ELISA, and for inhibition of choroidal neovascularisation in mice and rats.
A novel disubstituted furan inhibitor was selective for the SRPK family of kinases and reduced expression of pro-angiogenic but not antiangiogenic VEGF isoforms. This inhibitor and previously identified SRPK inhibitors significantly reduced choroidal neovascularisation in vivo. Topical administration of SRPK inhibitors dose-dependently blocked CNV with an EC50 of 9 μM.
These results indicate that novel SRPK1 selective inhibitors could be a potentially novel topical (eye drop) therapeutic for wet AMD.
Exudative AMD results from excess angiogenic VEGF production. Switching VEGF splicing to the antiangiogenic isoforms by inhibiting serine-rich protein kinase-1 (SRPK1) can be achieved by topical application of novel SRPK1 inhibitors. This could result in novel treatments that do not require injection in wet AMD.
VEGF; splicing; AMD
Atrial fibrillation (AF), the commonest cardiac arrhythmia, has been strongly linked with arrhythmogenic sources near the pulmonary veins (PVs), but underlying mechanisms are not fully understood. We aim to study the generation and sustenance of wave sources in a model of the PV tissue.
Methods and results
A previously developed biophysically detailed three-dimensional canine atrial model is applied. Effects of AF-induced electrical remodelling are introduced based on published experimental data, as changes of ion channel currents (ICaL, IK1, Ito, and IKur), the action potential (AP) and cell-to-cell coupling levels. Pharmacological effects are introduced by blocking specific ion channel currents. A combination of electrical heterogeneity (AP tissue gradients of 5–12 ms) and anisotropy (conduction velocities of 0.75–1.25 and 0.21–0.31 m/s along and transverse to atrial fibres) can results in the generation of wave breaks in the PV region. However, a long wavelength (171 mm) prevents the wave breaks from developing into re-entry. Electrical remodelling leads to decreases in the AP duration, conduction velocity and wavelength (to 49 mm), such that re-entry becomes sustained. Pharmacological effects on the tissue heterogeneity and vulnerability (to wave breaks and re-entry) are quantified to show that drugs that increase the wavelength and stop re-entry (IK1 and IKur blockers) can also increase the heterogeneity (AP gradients of 26–27 ms) and the likelihood of wave breaks.
Biophysical modelling reveals large conduction block areas near the PVs, which are due to discontinuous fibre arrangement enhanced by electrical heterogeneity. Vulnerability to re-entry in such areas can be modulated by pharmacological interventions.
Atrial arrhythmias; Pulmonary veins; Computational modelling; Re-entrant waves; Drug effects
Over the past year, three articles have appeared in the New England Journal of Medicine describing conflicting findings about azithromycin and cardiac safety, particular azithromycin-induced QTc interval prolongation and torsade de pointes. The FDA wants healthcare providers to consider azithromycin-induced fatal cardiac arrhythmias for patients already at risk for cardiac death and other potentially arrhythmogenic cardiovascular conditions. In a systematic review of case reports we sought to determine factors that link to azithromycin-induced/associated QTc interval prolongation and torsade de pointes. We found 12 cases: seven female and five male. Of the nine adults with reported azithromycin doses, concurrent QTc interval measurement, and without congenital long QT syndrome, we found no significant relationship between dose and QTc interval duration. Additional risk factors were female sex, older age, heart disease, QTc interval prolonging drugs and metabolic inhibitors, hypokalemia, and bradycardia. All 12 subjects had at least two additional risk factors. Elderly women with heart disease appear to be at particularly risk for drug-related QTc interval prolongation and torsade de pointes.
azithromycin; cardiovascular death; drug-induced QTc interval prolongation; risk factors; torsade de pointes
In the light of the recent United States Food and Drug Administration (FDA) warning to clinicians on using previously approved doses of citalopram because of the purported higher risk of torsade de pointes (TdP), we pursued the broader question: are selective serotonin reuptake inhibitor (SSRI) antidepressant agents as a group unsafe because they might induce QTc interval prolongation and TdP?
We reviewed the literature and found only 15 case reports (6 of fluoxetine, 1 of sertraline and 8 of citalopram) of SSRI-associated QTc interval prolongation linking to TdP.
A total of 13 cases contained sufficient information for analysis. In the setting of TdP, QTc interval prolongation does not clearly relate to SSRI dose.
Applying conventional statistics as the FDA does may not be the best tool to study this phenomenon because SSRI-associated TdP is a very rare event and hence best understood as an ‘extreme outlier’. Despite the limitations inherent in case report material, case reports on drug-associated QTc interval prolongation and TdP provide valuable information that should be considered along with other sources of information for clinical guidance.
antidepressant drugs; QTc interval prolongation; SSRIs; torsade de pointes
Nav1.5 is the principal voltage-gated sodium channel expressed in heart, and is also expressed at lower abundance in embryonic dorsal root ganglia (DRG) with little or no expression reported postnatally. We report here the expression of Nav1.5 mRNA isoforms in adult mouse and rat DRG. The major isoform of mouse DRG is Nav1.5a, which encodes a protein with an IDII/III cytoplasmic loop reduced by 53 amino acids. Western blot analysis of adult mouse DRG membrane proteins confirmed the expression of Nav1.5 protein. The Na+ current produced by the Nav1.5a isoform has a voltage-dependent inactivation significantly shifted to more negative potentials (by ~5 mV) compared to the full-length Nav1.5 when expressed in the DRG neuroblastoma cell line ND7/23. These results imply that the alternatively spliced exon 18 of Nav1.5 plays a role in channel inactivation and that Nav1.5a is likely to make a significant contribution to adult DRG neuronal function.
We reviewed the literature and found 31 adult cases and 1 newborn case of methadone-associated QTc interval prolongation and/or torsade de pointes (TdP). Parametric statistics may not be useful in studying this issue because methadone-associated TdP is a very rare event and, hence, “an extreme outlier” consistent with scalable randomness. We may have to rely upon narrative medicine in the form of case reports with all its limitations and hazards to provide our best understanding. We report risk factors for methadone-associated QTc interval prolongation and TdP based on review of published case reports. We believe both drug manufacturers and the FDA would better serve our patients and inform clinicians if they more readily reported drug-induced outliers such as methadone-associated TdP using a case report format.
Drug-induced QTc interval prolongation; methadone; risk factors; torsade de pointes
The manufacturers of clarithromycin sought a drug similar in efficacy to erythromycin but with a superior side-effect profile. They generally achieved this outcome, but postmarketing findings identified a series of reports linking clarithromycin to QTc interval prolongation and torsades de pointes (TdP) ultimately leading to a Black Box Warning. We sought to clarify risk factors associated with TdP among case reports of patients receiving clarithromycin linked to QTc interval prolongation and TdP.
Methods and results:
In a detailed literature search, we found 15 women, five men, and one boy meeting our search criteria. Among the 17 adults with reported clarithromycin dose and concurrent QTc interval measurement, we found no statistically significant relationship between clarithromycin dose and QTc interval duration. This did not change for the adults who developed TdP. Among adults, major risk factors were female sex (15), old age (11) and heart disease (17). A total of eight adult subjects had all three major risk factors and 14 of the 20 adults had at least two major risk factors. All adult subjects had at least two risk factors besides clarithromycin. A total of four of the 20 adults received cisapride and three received disopyramide. Three adults were considered to suffer from some aspect of the congenital long QT syndrome.
We believe that the risk factor description for this drug should be refined to emphasize the major risk factors of (1) female sex, (2) old age and (3) heart disease.
Clarithromycin; drug-induced QTc interval prolongation; risk factors; torsades de pointes
Introduction: Genetic forms of the Short QT Syndrome (SQTS) arise due to cardiac ion channel mutations leading to accelerated ventricular repolarization, arrhythmias and sudden cardiac death. Results from experimental and simulation studies suggest that changes to refractoriness and tissue vulnerability produce a substrate favorable to re-entry. Potential electromechanical consequences of the SQTS are less well-understood. The aim of this study was to utilize electromechanically coupled human ventricle models to explore electromechanical consequences of the SQTS.
Methods and Results: The Rice et al. mechanical model was coupled to the ten Tusscher et al. ventricular cell model. Previously validated K+ channel formulations for SQT variants 1 and 3 were incorporated. Functional effects of the SQTS mutations on [Ca2+]i transients, sarcomere length shortening and contractile force at the single cell level were evaluated with and without the consideration of stretch-activated channel current (Isac). Without Isac, at a stimulation frequency of 1Hz, the SQTS mutations produced dramatic reductions in the amplitude of [Ca2+]i transients, sarcomere length shortening and contractile force. When Isac was incorporated, there was a considerable attenuation of the effects of SQTS-associated action potential shortening on Ca2+ transients, sarcomere shortening and contractile force. Single cell models were then incorporated into 3D human ventricular tissue models. The timing of maximum deformation was delayed in the SQTS setting compared to control.
Conclusion: The incorporation of Isac appears to be an important consideration in modeling functional effects of SQT 1 and 3 mutations on cardiac electro-mechanical coupling. Whilst there is little evidence of profoundly impaired cardiac contractile function in SQTS patients, our 3D simulations correlate qualitatively with reported evidence for dissociation between ventricular repolarization and the end of mechanical systole.
short QT syndrome; stretch-activated channel; mechanical contraction; 3D model; human ventricles
► We report the block of the β-adrenoceptor-activated cardiac CFTR Cl− current by Ni2+. ► Extracellular Ni2+ inhibits the current activated by β1-adrenoceptors in a concentration-dependent manner. ► The action of Ni2+ is insensitive to β2-blockade. ► Ni2+ does not affect the β-adrenoceptor-activated current from the intracellular side. ► The data are consistent with an action of Ni2+ at the β1-adrenoceptor from the external side.
Cardiac ventricular myocytes exhibit a protein kinase A-dependent Cl− current (ICl.PKA) mediated by the cystic fibrosis transmembrane conductance regulator (CFTR). There is conflicting evidence regarding the ability of the divalent cation nickel (Ni2+), which has been used widely in vitro in the study of other cardiac ionic conductances, to inhibit ICl.PKA. Here the action of Ni2+ on ICl.PKA activated by β-adrenergic stimulation has been elucidated. Whole-cell patch-clamp recordings were made from rabbit isolated ventricular myocytes. Externally applied Ni2+ blocked ICl.PKA activated by 1 μM isoprenaline with a log IC50 (M) of −4.107 ± 0.075 (IC50 = 78.1 μM) at +100 mV and −4.322 ± 0.107 (IC50 = 47.6 μM) at −100 mV. Thus, the block of ICl.PKA by Ni2+ was not strongly voltage dependent. Ni2+ applied internally via the patch-pipette was ineffective at inhibiting isoprenaline-activated ICl,PKA, but in the same experiments the current was suppressed by external Ni2+ application, indicative of an external site of Ni2+ action. In the presence of 1 μM atenolol isoprenaline was ineffective at activating ICl.PKA, but in the presence of the β2-adrenoceptor inhibitor ICI 118,551 isoprenaline still activated Ni2+-sensitive ICl.PKA. Collectively, these data demonstrate that Ni2+ ions produce marked inhibition of β1-adrenoceptor activated ventricular ICl.PKA at submillimolar [Ni2+]: an action that is likely to involve an interaction between Ni2+ and β1-adrenoceptors. The concentration-dependence for ICl.PKA inhibition seen here indicates the potential for confounding effects on ICl,PKA to occur even at comparatively low Ni2+ concentrations, when Ni2+ is used to study other cardiac ionic currents under conditions of β-adrenergic agonism.
Rabbit cardiomyocytes; PKA-dependent Cl− current; CFTR; CFTR-inhibitor; Nickel; Ni2+
The familial Short QT Syndrome (SQTS) is associated with an increased risk of cardiac arrhythmia and sudden death. Gain-of-function mutations in the hERG K+ channel protein have been linked to variant 1 of the SQTS. A hERG channel pore (T618I) mutation has recently been identified in families with heritable SQTS. This study aimed to determine effects of the T618I-hERG mutation on (i) hERG current (IhERG) elicited by ventricular action potentials; (ii) the sensitivity of IhERG to inhibition by four clinically used antiarrhythmic drugs.
Electrophysiological recordings of IhERG were made at 37°C from HEK 293 cells expressing wild-type (WT) or T618I hERG. Whole-cell patch clamp recording was performed using both conventional voltage clamp and ventricular action potential (AP) clamp methods.
Under conventional voltage-clamp, WT IhERG peaked at 0-+10 mV, whilst for T618I IhERG maximal current was right-ward shifted to ∼ +40 mV. Voltage-dependent activation and inactivation of T618I IhERG were positively shifted (respectively by +15 and ∼ +25 mV) compared to WT IhERG. The IhERG ‘window’ was increased for T618I compared to WT hERG. Under ventricular AP clamp, maximal repolarising WT IhERG occurred at ∼ -30 mV, whilst for T618I hERG peak IhERG occurred earlier during AP repolarisation, at ∼ +5 mV. Under conventional voltage clamp, half-maximal inhibitory concentrations (IC50) for inhibition of IhERG tails by quinidine, disopyramide, D-sotalol and flecainide for T618I hERG ranged between 1.4 and 3.2 fold that for WT hERG. Under action potential voltage clamp, T618I IC50s ranged from 1.2 to 2.0 fold the corresponding IC50 values for WT hERG.
The T618I mutation produces a more modest effect on repolarising IhERG than reported previously for the N588K-hERG variant 1 SQTS mutation. All drugs studied here appear substantially to retain their ability to inhibit IhERG in the setting of the SQTS-linked T618I mutation.
Introduction: β-adrenergic stimulation increases the heart rate by accelerating the electrical activity of the pacemaker of the heart, the sinoatrial node (SAN). Ionic mechanisms underlying the actions of β-adrenergic stimulation are not yet fully understood. Isoprenaline (ISO), a β-adrenoceptor agonist, shifts voltage-dependent If activation to more positive potentials resulting in an increase of If, which has been suggested to be the main mechanism underlying the effect of β-adrenergic stimulation. However, ISO has been found to increase the firing rate of rabbit SAN cells when If is blocked. ISO also increases ICaL, Ist, IKr, and IKs; and shifts the activation of IKr to more negative potentials and increases the rate of its deactivation. ISO has also been reported to increase the intracellular Ca2+ transient, which can contribute to chronotropy by modulating the “Ca2+ clock.” The aim of this study was to analyze the ionic mechanisms underlying the positive chronotropy of β-adrenergic stimulation using two distinct and well established computational models of the electrical activity of rabbit SAN cells. Methods and results: We modified the Boyett et al. (2001) and Kurata et al. (2008) models of electrical activity for the central and peripheral rabbit SAN cells by incorporating equations for the known dose-dependent actions of ISO on various ionic channel currents (ICaL, Ist, IKr, and IKs), kinetics of IKr and If, and the intracellular Ca2+ transient. These equations were constructed from experimental data. To investigate the ionic basis of the effects of ISO, we simulated the chronotropic effect of a range of ISO concentrations when ISO exerted all its actions or just a subset of them. Conclusion: In both the Boyett et al. and Kurata et al. SAN models, the chronotropic effect of ISO was found to result from an integrated action of ISO on ICaL, If, Ist, IKr, and IKs, among which an increase in the rate of deactivation of IKr plays a prominent role, though the effect of ISO on If and [Ca2+]i also plays a role.
sinoatrial node; isoprenaline; action potential
► ACh and ET-1 activate a K+ current in cardiac atrioventricular nodal cells. ► Tertiapin-Q sensitive IKACh activated via M2 receptors shows bi-exponential ‘fade’. ► ET-1 activates a similar current that also fades. ► The fade reflects desensitization rather than altered K+ ion driving force. ► Acetylcholine is able to cross-desensitize the AVN cell response to endothelin-1.
The atrioventricular node (AVN) is a vital component of the pacemaker-conduction system of the heart, co-ordinating conduction of electrical excitation from cardiac atria to ventricles and acting as a secondary pacemaker. The electrical behaviour of the AVN is modulated by vagal activity via activation of muscarinic potassium current, IKACh. However, it is not yet known if this response exhibits ‘fade’ or desensitization in the AVN, as established for the heart’s primary pacemaker – the sinoatrial node. In this study, acute activation of IKACh in rabbit single AVN cells was investigated using whole-cell patch clamp at 37 °C. 0.1–1 μM acetylcholine (ACh) rapidly activated a robust IKACh in AVN myocytes during a descending voltage-ramp protocol. This response was inhibited by tertiapin-Q (TQ; 300 nM) and by the M2 muscarinic ACh receptor antagonist AFDX-116 (1 μM). During sustained ACh exposure the elicited IKACh exhibited bi-exponential fade (τf of 2.0 s and τs 76.9 s at −120 mV; 1 μM ACh). 10 nM ET-1 elicited a current similar to IKACh, which faded with a mono-exponential time-course (τ of 52.6 s at −120 mV). When ET-1 was applied following ACh, the ET-1 activated response was greatly attenuated, demonstrating that ACh could desensitize the response to ET-1. For neither ACh nor ET-1 was the rate of current fade dependent upon the initial response magnitude, which is inconsistent with K+ flux mediated changes in electrochemical driving force as the underlying mechanism. Collectively, these findings demonstrate that TQ sensitive inwardly rectifying K+ current in cardiac AVN cells, elicited by M2 muscarinic receptor or ET-1 receptor activation, exhibits fade due to rapid desensitization.
Acetylcholine (ACh); Atrioventricular node; AV node; AVN; Endothelin-1 (ET-1); GIRK; IKACh; Inward rectifier; Muscarinic potassium current; Tertiapin-Q