Cardiac Voltage-gated Sodium Channel Research Articles

A Missense Mutation in the Sodium Channel β2 Subunit RevealsSCN2Bas a New Candidate Gene for Brugada Syndrome

Brugada Syndrome (BrS) is a familial disease associated with sudden cardiac death. A 20%-25% of BrS patients carry genetic defects that cause loss-of-function of the voltage-gated cardiac sodium channel. Thus, 70%-75% of patients remain without a genetic diagnosis. In this work, we identified a novel missense mutation (p.Asp211Gly) in the sodium β2 subunit encoded by SCN2B, in a woman diagnosed with BrS. We studied the sodium current (INa ) from cells coexpressing Nav 1.5 and wild-type (β2WT) or mutant (β2D211G) β2 subunits. Our electrophysiological analysis showed a 39.4% reduction in INa density when Nav 1.5 was coexpressed with the β2D211G. Single channel analysis showed that the mutation did not affect the Nav 1.5 unitary channel conductance. Instead, protein membrane detection experiments suggested that β2D211G decreases Nav 1.5 cell surface expression. The effect of the mutant β2 subunit on the INa strongly suggests that SCN2B is a new candidate gene associated with BrS.

Expression of the sialyltransferase, ST3Gal4, impacts cardiac voltage-gated sodium channel activity, refractory period and ventricular conduction

The sequential glycosylation process typically ends with sialic acid residues added through trans-Golgi sialyltransferase activity. Individuals afflicted with congenital disorders of glycosylation often have reduced glycoprotein sialylation and present with multi-system symptoms including hypotonia, seizures, arrhythmia and cardiomyopathy. Cardiac voltage-gated Na+ channel (Nav) activity can be influenced by sialic acids likely contributing to an external surface potential causing channels to gate at less depolarized voltages. Here, a possible pathophysiological role for reduced sialylation is investigated by questioning the impact of gene deletion of the uniformly expressed beta-galactoside alpha-2,3-sialyltransferase 4 (ST3Gal4) on cardiac Nav activity, cellular refractory period and ventricular conduction. Whole-cell patch-clamp experiments showed that ventricular Nav from ST3Gal4 deficient mice (ST3Gal4−/−) gated at more depolarized potentials, inactivated more slowly and recovered from fast inactivation more rapidly than WT controls. Current-clamp recordings indicated a 20% increase in time to action potential peak and a 30ms decrease in ST3Gal4−/− myocyte refractory period, concurrent with increased Nav recovery rate. Nav expression, distribution and maximal Na+ current levels were unaffected by ST3Gal4 expression, indicating that reduced sialylation does not impact Nav surface expression and distribution. However, enzymatic desialylation suggested that ST3Gal4−/− ventricular Nav are less sialylated. Consistent with the shortened myocyte refractory period, epicardial conduction experiments using optical mapping techniques demonstrated a 27% reduction in minimum ventricular refractory period and increased susceptibility to arrhythmias in ST3Gal4−/− ventricles. Thus, deletion of a single sialyltransferase significantly impacts ventricular myocyte electrical signaling. These studies offer insight into diseases of glycosylation that are often associated with pathological changes in excitability and highlight the importance of glycosylation in cardiac physiology.

Stereospecific Synthesis of m-Hydroxymexiletine Enantiomers

m-Hydroxymexiletine (MHM) is a metabolite of mexiletine, a well known class IB anti-arrhythmic drug, which presents almost twice the activity of the parent compound on cardiac voltage-gated sodium channels. Given the different activity of mexiletine enantiomers on sodium currents (being the R-isomer the eutomer), it is conceivable that (R)- and(S)-MHM could differ in pharmacodynamic and pharmacokinetic properties, too. Herein we report the efficient synthesis of MHM enantiomers that could represent useful tools for further investigations on stereospecific requirements of the voltage-gated sodium channel binding site. MHM enantiomers and all the homochiral intermediates were fully characterized. The ee values for (R)- and (S)-MHM were >99%, as assessed by capillary electrophoresis using β-cyclodextrin sulfated sodium salt as a chiral selector.

Proton Sensors in the Pore Domain of the Cardiac Voltage-gated Sodium Channel

Protons impart isoform-specific modulation of inactivation in neuronal, skeletal muscle, and cardiac voltage-gated sodium (Na(V)) channels. Although the structural basis of proton block in Na(V) channels has been well described, the amino acid residues responsible for the changes in Na(V) kinetics during extracellular acidosis are as yet unknown. We expressed wild-type (WT) and two pore mutant constructs (H880Q and C373F) of the human cardiac Na(V) channel, Na(V)1.5, in Xenopus oocytes. C373F and H880Q both attenuated proton block, abolished proton modulation of use-dependent inactivation, and altered pH modulation of the steady-state and kinetic parameters of slow inactivation. Additionally, C373F significantly reduced the maximum probability of use-dependent inactivation and slow inactivation, relative to WT. H880Q also significantly reduced the maximum probability of slow inactivation and shifted the voltage dependence of activation and fast inactivation to more positive potentials, relative to WT. These data suggest that Cys-373 and His-880 in Na(V)1.5 are proton sensors for use-dependent and slow inactivation and have implications in isoform-specific modulation of Na(V) channels.

Proteasome inhibitor (MG132) rescues Nav1.5 protein content and the cardiac sodium current in dystrophin-deficient mdx5cv mice

The cardiac voltage-gated sodium channel, Nav1.5, plays a central role in cardiac excitability and impulse propagation and associates with the dystrophin multiprotein complex at the lateral membrane of cardiomyocytes. It was previously shown that Nav1.5 protein content and the sodium current (lNa) were both decreased in cardiomyocytes of dystrophin-deficient mdx5cv mice. In this study, wild-type and mdx5cv mice were treated for 7 days with the proteasome inhibitor MG132 (10 μg/Kg/24 h) using implanted osmotic mini pumps. MG132 rescued both the total amount of Nav1.5 protein and lNa but, unlike in previous studies, de novo expression of dystrophin was not observed in skeletal or cardiac muscle. This study suggests that the reduced expression of Nav1.5 in dystrophin-deficient cells is dependent on proteasomal degradation.

Sudden arrhythmic death: from basic science to clinical practice

EDITORIAL article Front. Physiol., 25 November 2013Sec. Cardiac Electrophysiology Volume 4 - 2013 | https://doi.org/10.3389/fphys.2013.00339

The genetic component of Brugada syndrome

Brugada syndrome (BrS) is a clinical entity first described in 1992. BrS is characterized by ST-segment elevations in the right precordial leads and susceptibility to ventricular arrhythmias and sudden cardiac death. It affects young subjects, predominantly males, with structurally normal hearts. The prevalence varies with ethnicity ranging from 1:2,000 to 1:100,000 in different parts of the world. Today, hundreds of variants in 17 genes have been associated with BrS of which mutations in SCN5A, coding for the cardiac voltage-gated sodium channel, accounts for the vast majority. Despite this, approximately 70% of BrS cases cannot be explained genetically with the current knowledge. Moreover, the monogenic role of some of the variants previously described as being associated with BrS has been questioned by their occurrence in about 4% (1:23) of the general population as found in NHLBI GO Exome Sequencing Project (ESP) currently including approximately 6500 individuals. If we add the variants described in the five newest identified genes associated with BrS, they appear at an even higher prevalence in the ESP (1:21). The current standard treatment of BrS is an implantable cardioverter-defibrillator (ICD). The risk stratification and indications for ICD treatment are based on the ECG and on the clinical and family history. In this review we discuss the genetic basis of BrS.

An Improved Synthesis of m-Hydroxymexiletine, a Potent Mexiletine Metabolite

m-Hydroxymexiletine (MHM), a minor metabolite of the class IB anti-arrhythmic drug mexiletine, is about two fold more potent than the parent compound on human cardiac voltage-gated sodium channels (hNav1.5), and equipotent to mexiletine on human skeletal-muscle voltage-gated sodium channels (hNav1.4). Herein, an alternative and simplified synthesis of this promising compound has been accomplished. This route, as well as being more efficient, has the advantage, over the first, to avoid the use of oxidizing agents, such as the meta-chloroperoxybenzoic acid.

Abstract 19390: Sirtuin 1 Modulates Cardiac Sodium Channels in vivo

The rapid upstroke in phase 0 of the cardiac action potential is the result of the inward sodium current (I Na ) carried by the cardiac voltage-gated sodium channel Na v 1.5 (SCN5A). We recently reported that the overexpression of Silent Information Regulator 1 (Sirtuin 1, SIRT1), an NAD + dependent protein deacetylase, deacetylated SCN5A, increased its cell surface expression, and increased I Na in HEK-293 cells and rat neonatal cardiac myocytes. Overexpression of a dominant negative SIRT1 construct decreased SCN5A cell surface expression and I Na . The role of acetylation of SCN5A and its regulation by SIRT1 has not been shown in vivo, however. We engineered cardiac specific Sirt1 knockout (cSirt1KO) mice by breeding Sirt1 flox/flox with α-MHC-cre mice on a C57BL-6 background. cSirt1KO compared to littermate Sirt1 flox/flox mice showed much less expression of Sirt1 mRNA (12±6%, n=3) and protein in the heart but not in the kidney, and Scn5a immunoprecipitated from cardiac membrane fractions of the cSirt1KO mice showed an ∼2-fold increase in acetylation (Figure A). At 3 months of age, EKGs from anesthetized cSirt1KO compared to littermate Sirt1 flox/flox mice showed PR prolongation (43±3 ms, n=4, vs. 37±2 ms, n=5; p<0.02) and telemetry from ambulatory cSirt1KO mice showed high degree heart block not seen in the controls (Figure B). Thus, our results show that Scn5a is acetylated in vivo in the mouse, that the degree of acetylation is regulated by cardiac-specific expression of Sirt1, and that decreases in Sirt1 lead to conduction disease similar to those seen in Scn5a +/- mice. These findings raise the possibility that pharmacological manipulation of Sirt1 may provide a means to modulate cardiac I Na and arrhythmias.

The β1-Subunit of Nav1.5 Cardiac Sodium Channel Is Required for a Dominant Negative Effect through α-α Interaction

Brugada syndrome (BrS) is an inherited autosomal dominant cardiac channelopathy. Several mutations on the cardiac sodium channel Nav1.5 which are responsible for BrS lead to misfolded proteins that do not traffic properly to the plasma membrane. In order to mimic patient heterozygosity, a trafficking defective mutant, R1432G was co-expressed with Wild Type (WT) Nav1.5 channels in HEK293T cells. This mutant significantly decreased the membrane Na current density when it was co-transfected with the WT channel. This dominant negative effect did not result in altered biophysical properties of Nav1.5 channels. Luminometric experiments revealed that the expression of mutant proteins induced a significant reduction in membrane expression of WT channels. Interestingly, we have found that the auxiliary Na channel β1-subunit was essential for this dominant negative effect. Indeed, the absence of the β1-subunit prevented the decrease in WT sodium current density and surface proteins associated with the dominant negative effect. Co-immunoprecipitation experiments demonstrated a physical interaction between Na channel α-subunits. This interaction occurred only when the β1-subunit was present. Our findings reveal a new role for β1-subunits in cardiac voltage-gated sodium channels by promoting α-α subunit interaction which can lead to a dominant negative effect when one of the α-subunits shows a trafficking defective mutation.

Effects of fluorine-containing opener of ATP-sensitive potassium channels, pinacidil-derivative flocalin, on cardiac voltage-gated sodium and calcium channels

Fluorine-containing pinacidil-derivative flocalin is an effective adenosine triphosphate-sensitive potassium (K(ATP))-channel opener with pronounced vasodilatory, cardioprotective effects and low general toxicity. By activating cardiac K(ATP) channels, flocalin hyperpolarizes cardiac myocytes, decreases their excitability, reduces Ca(2+) entry, and inhibits Ca(2+)-dependent signalling processes. Since our previous studies indicated that the drug also influences the rate of rise and amplitude of the cardiomyocyte's action potential, here we have investigated its possible actions on depolarizing inward currents through voltage-gated sodium (VGSC) and L-type calcium (VGCC) channels. Experiments were conducted on cultured cardiac myocytes prepared from the whole hearts of neonatal rats and maintained in culture for 1-3days using whole-cell patch-clamp technique with no distinction of myocyte's type. Flocalin concentration dependently inhibited the Na(+) inward current through VGSCs with IC(50) = 17.4μM and a maximal extent of 0.54, slowed down its inactivation kinetics, and hyperpolarized steady-state inactivation by 5.6mV. The drug also inhibited calcium current through L-type VGCCs with IC(50) = 24.1μM and a maximal block of 0.38, without affecting its inactivation but producing 5.3-mV hyperpolarization shifting of steady-state activation. Inhibition of both depolarizing currents by flocalin in addition to its ability to open K(ATP) channels enhances the suppressive action of the drug on cardiac excitability and broadens its pharmacological effects. Since, according to our previous data, cardiac K(ATP)-channel opening by flocalin occurs with ЕC(50) = 8μM, the possibility of partial blockade of VGSC and L-type VGCCs should be considered when determining the therapeutic concentrations of the compound during its use as a cardioprotector.

The human Nav1.5 F1486 deletion associated with long QT syndrome leads to impaired sodium channel inactivation and reduced lidocaine sensitivity

The deletion of phenylalanine 1486 (F1486del) in the human cardiac voltage-gated sodium channel (hNav1.5) is associated with fatal long QT (LQT) syndrome. In this study we determined how F1486del impairs the functional properties of hNav1.5 and alters action potential firing in heterologous expression systems (human embryonic kidney (HEK) 293 cells) and their native cardiomyocyte background. Cells expressing hNav1.5-F1486del exhibited a loss-of-function alteration, reflected by an 80% reduction of peak current density, and several gain-of-function alterations, including reduced channel inactivation, enlarged window current, substantial augmentation of persistent late sodium current and an increase in ramp current. We also observed substantial action potential duration (APD) prolongation and prominent early afterdepolarizations (EADs) in neonatal cardiomyocytes expressing the F1486del channels, as well as in computer simulations of myocyte activity. In addition, lidocaine sensitivity was dramatically reduced, which probably contributed to the poor therapeutic outcome observed in the patient carrying the hNav1.5-F1486del mutation. Therefore, despite the significant reduction in peak current density, the F1486del mutation also leads to substantial gain-of-function alterations that are sufficient to cause APD prolongation and EADs, the predominant characteristic of LQTs. These data demonstrate that hNav1.5 mutations can have complex functional consequences and highlight the importance of identifying the specific molecular defect when evaluating potential treatments for individuals with prolonged QT intervals.

An Improved Synthesis of m-Hydroxymexiletine, a Potent Mexiletine Metabolite

m-Hydroxymexiletine (MHM), a minor metabolite of the class IB anti-arrhythmic drug mexiletine, is about two fold more potent than the parent compound on human cardiac voltage-gated sodium channels (hNav1.5), and equipotent to mexiletine on human skeletal-muscle voltage-gated sodium channels (hNav1.4). Herein, an alternative and simplified synthesis of this promising compound has been accomplished. This route, as well as being more efficient, has the advantage, over the first, to avoid the use of oxidizing agents, such as the meta-chloroperoxybenzoic acid.

Cardiac Sodium Channel Nav1.5 Mutations and Cardiac Arrhythmia

As a major cardiac voltage-gated sodium channel isoform in the heart, the Nav1.5 channel is essential for cardiac action potential initiation and subsequent propagation throughout the heart. Mutations of Nav1.5 have been linked to a variety of cardiac diseases such as long QT syndrome (LQTs), Brugada syndrome, cardiac conduction defect, atrial fibrillation, and dilated cardiomyopathy. The mutagenesis approach and heterologous expression systems are most frequently used to study the function of this channel. This review focuses primarily on recent findings of Nav1.5 mutations associated with type 3 long QT syndrome (LQT3) in particular. Understanding the functional changes of the Nav1.5 mutation may offer critical insight into the mechanism of long QT3 syndrome. In addition, this review provides the updated information on the current progress of using various experimental model systems to study primarily the long QT3 syndrome.

Ranolazine Effects on NaV1.2 and Modulation by pH

Ranolazine is an anti-anginal drug previously shown to block persistent currents of the cardiac voltage gated sodium channel, NaV1.5. The effects of ranolazine, however, have not yet been described in all sodium channel isoforms. We studied the effects of ranolazine on the neuronal sodium channel isoform, NaV1.2, and its modulation by extracellular protons. Ionic currents were measured from Chinese Hamster Ovary (CHO) cells expressing the α-subunit of NaV1.2, using whole cell patch clamp techniques. Voltage protocols were run with extracellular solutions of pH 7.4 and pH 6.0 before and after the addition of ranolazine. The addition of 100 μM ranolazine at pH 7.4 led to a significant decrease in late sodium current, faster rate of open state fast inactivation, and slower recovery from inactivation. Similar trends were seen in preliminary experiments at 10 μM and 30 μM ranolazine. In addition, we observed a tonic block of peak current and an increase in total use-dependent block. Many of these effects were different at low pH. Low pH led to a significant depolarizing shift of the conductance curve, and slowed the onset of fast inactivation. Adding ranolazine at pH 6.0 significantly decreased the rate of fast inactivation recovery, and increased the total use-dependent block and rate of open state inactivation. Although directions of the effects were unchanged, the magnitude of these effects was significantly different between the two pH values. Our results suggest that ranolazine stabilizes fast inactivation at both pH 7.4, and pH 6.0. It is possible that ranolazine and protons may compete for a common binding site or indirectly interact leading to decreased ranolazine efficacy at low pH. (Supported by a research grant from Gilead Sciences, Inc. and an NSERC Discovery Grant to PCR, and an NSERC URSA to CHP.)

Proton Modulation of Gating Currents in the Cardiac Voltage-Gated Sodium Channel, NaV1.5

Low pH reduces single channel conductance and destabilizes the fast-inactivated state of the cardiac voltage-gated sodium channel (NaV1.5) by increasing both window and persistent currents (Jones et al., 2011, Biophys. J. 101(7)). Since fast inactivation is tightly coupled to NaV channel voltage sensor activation, we hypothesized that alterations in the kinetics of voltage sensor movement may underlie the destabilization of the fast-inactivated state in NaV1.5. To test this, we expressed NaV1.5 channels in Xenopus oocytes and recorded gating currents using a cut-open voltage clamp with extracellular solution titrated to either pH 7.4 (control) or pH 6.0. At pH 6.0, compared to pH 7.4, the V1/2 of the Q(V) curve was significantly depolarized (from −57.8±4.3 mV to −40.8±5.1 mV). Additionally, the slow time constant of charge recovery was significantly reduced from 16.1±5.0 ms at pH 7.4 to 9.7±4.2 ms at pH 6.0. These data suggest a molecular basis for the increased persistent and window currents previously shown in NaV1.5 channels at reduced extracellular pH. Specifically, protons may electrostatically affect the rate of voltage sensor movement, either by directly binding to extracellular residues (e.g. H880) or indirectly by binding to carboxylates in the pore domain (Kahn et al., J Physiol. 2002, 543). (Supported by an NSERC Discovery Grant to PCR, a CFI Infrastructure grant to PCR and TC, and a CIHR Vanier Scholarship to DKJ.)

Perturbation of sodium channel structure by an inherited Long QT Syndrome mutation

The cardiac voltage-gated sodium channel (NaV1.5) underlies impulse conduction in the heart and its depolarization-induced inactivation is essential in control of the duration of the QT interval of the electrocardiogram (ECG). Perturbation of Nav1.5 inactivation by drugs or inherited mutation can underlie and trigger cardiac arrhythmias. The carboxy terminus plays an important role in channel inactivation, but complete structural information on its predicted structural domain is unknown. Here we measure interactions between the functionally critical distal C-T alpha helix (H6) and the proximal structured EF hand motif using transition metal ion FRET. We measure distances at three loci along H6 relative to an intrinsic tryptophan, demonstrating the proximal-distal interaction in a contiguous carboxy terminus polypeptide. Using these data together with the existing NaV1.5 carboxy terminus NMR structure, we construct a model of the predicted structured region of the carboxy terminus. An arrhythmia associated H6 mutant which impairs inactivation decreases FRET, indicating destabilization of the distal-proximal intramolecular interaction. These data provide a structural correlate to the pathological phenotype of the mutant channel.

What Can Genetic Studies of Left Ventricular Mass Tell Us?

Left ventricular hypertrophy (LVH), whether measured by electrocardiography, echocardiography, or at autopsy, is associated with increased risk of cardiovascular morbidity and mortality.1,2 Although LVH has been associated with several clinical characteristics such as age, sex, body mass index, and hypertension, the effects and their directionality have varied according to the criteria for LVH.3 Aggregation within families suggests a heritable component due to genetics,4 although a recent study of echocardiographic left ventricular mass in a large sample was unable to identify common genetic factors.5 Treatment of hypertensives with LVH can lead to its regression and with it reduction in risk of cardiovascular disease.6 Thus, it appears to be a modifiable risk factor, although LVH induced by exercise training is thought to be benign. Thus, LVH is a complex trait with diverse definitions, has multiple clinical and genetic contributors and is a modifiable risk factor in some but not all settings. Article see p 626 It is not surprising therefore that the (patho-) physiological mechanisms that underlie LVH remain incompletely characterized. Human genetics offers one opportunity to expose mechanisms not previously recognized to play a role in the development of LVH. Shah et al report in this issue of Circulation Cardiovascular Genetics a genetic screen to identify common genetic variants related to electrocardiographic correlates of left ventricular mass (ECG LVM): 2 reflecting simple voltage measures—Sokolow-Lyon voltage and 12-lead QRS voltage sum—and 2 incorporating QRS duration—Cornell product and QRS voltage product.7 The investigators genotyped gene-focused single-nucleotide polymorphisms (SNPs) found on the ITMAT-Broad-CARE (IBC) array, marketed by Illumina as the HumanCVD BeadChip 50K array, with ≈35 000 SNPs passing quality control. The array includes SNPs at 2100 candidate genes selected for being implicated in processes thought to be involved in cardiovascular disease as well as some …

Abstract 17305: Overexpression of Sodium Channel beta 4 Subunit is Responsible for Increase in Late Sodium Current in Mice with Mecp2 Deficiency

Introduction: The methyl-CpG-binding protein (MeCP2) is an epigenetic modifier of genetic expression and repression, and its dysfunction has been shown to cause Rett Syndrome, a progressive neurological disorder associated with sudden cardiac death. MeCP2 dysfunction has also been shown to be associated with QT interval in human patients, however the underlying mechanisms are unclear. We evaluated mice deficient in MeCP2 (KO) to determine whether there were differences in expression of genes commonly associated with long QT syndrome (LQTS). Hypothesis: MeCP2 dysfunction alters expression of plasma membrane ion channels or its interacting proteins, which are necessary for normal cardiomyocyte action potential. Methods and Results: qPCR was performed on wild type mice (WT) and KO heart lysates. Among 11 LQTS genes, SCN4B, one of beta subunits of the cardiac voltage gated sodium channel, was elevated 2.5 fold (n = 3, P = 0.04), while other 10 LQTS genes were not significantly altered. Western blot analysis on heart lysates showed increase of SCN4B protein in KO, consistent with immunostaining on isolated ventricular myocytes, which showed a qualitative increase of plasma membrane enhancement of SCN4B. Sodium current was recorded from a holding potential of -120 mV and activated to -20 mV. There was no change in peak sodium current, however, late sodium current was significantly increased in KO compared to WT (WT, -170.4 ± 29.2 pA, n = 5 vs. Rett, -590.0 ± 145.7 pA, n = 5; WT vs. Rett, p < 0.01). and Phenytoin, a late sodium current blocking agent, reduced late sodium current near normal levels (Rett + Phenytoin, -183.8 ± 40.6 pA, n = 5; Rett vs. Rett + Phenytoin, p < 0.05). Conclusion: Mice deficient in MeCP 2 have an increase in SCN4B expression, which may be associated with prolongation of QT interval. Phenytoin inhibits the late sodium current and may be an effective treatment for prevention of arrhythmias in patients with MeCP2 dysfunction.

Extracellular Proton Modulation of the Cardiac Voltage-Gated Sodium Channel, NaV1.5

Low pH depolarizes the voltage dependence of voltage-gated sodium (NaV) channel activation and fast inactivation. A complete description of NaV channel proton modulation, however, has not been reported. The majority of NaV channel proton modulation studies have been completed in intact tissue. Additionally, several NaV channel isoforms are expressed in cardiac tissue. Characterizing the proton modulation of the cardiac NaV channel, NaV1.5, will thus help define its contribution to ischemic arrhythmogenesis, where extracellular pH drops from pH 7.4 to as low as pH 6.0 within ∼10 min of its onset. We expressed the human variant of NaV1.5 with and without the modulating β1 subunit in Xenopus oocytes. Lowering extracellular pH from 7.4 to 6.0 affected a range of biophysical gating properties heretofore unreported. Specifically, acidic pH destabilized the fast-inactivated and slow-inactivated states, and elevated persistent INa. These data were incorporated into a ventricular action potential model that displayed a reduced maximum rate of depolarization as well as disparate increases in epicardial, mid-myocardial, and endocardial action potential durations, indicative of an increased heterogeneity of repolarization. Portions of these data were previously reported in abstract form.