Abstract 356: Loss of Function Mutations of Sodium Channel Beta-1 and Beta-2 Subunits Associated with Atrial Fibrillation and ST-segment Elevation

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Hiroshi Watanabe ◽  
Dawood Darbar ◽  
Christiana R Ingram ◽  
Kim Jiramongkolchai ◽  
Sameer S Chopra ◽  
...  
Keyword(s):  
Atrial Fibrillation ◽  
Steady State ◽  
Sodium Channel ◽  
Sodium Current ◽  
Β Subunit ◽  
Loss Of Function ◽  
St Segment Elevation ◽  
Negative Shift ◽  
St Segment ◽  
Cardiac Sodium Channel

Background: We have recently reported mutations in the cardiac sodium channel gene SCN5A in 5.9% of patients with atrial fibrillation (AF). In this study, we tested the hypothesis that mutations in sodium channel β subunit genes SCN1B-4B contribute to AF susceptibility. Methods and results: All 4 βsubunit genes were resequenced in 376 patients with AF (118 patients with lone AF and 258 patients with AF and cardiovascular disease) and 188 ethnically-defined controls. We identified 2 non-synonymous variants in SCN1B (resulting in R85H, D153N) and 2 in SCN2B (R28Q, R28W) in patients with AF; these occur at residues highly conserved across mammals and were absent in controls. In 3 of 4 mutation carriers, there was saddle back type ST-segment elevation in the right precordial leads of electrocardiogram. Transcripts encoding both SCN1B and SCN2B were detected in human atrium and ventricle. To assess function in vitro , CHO cells were transfected with SCN5A without β subunit, SCN5A with wild-type (WT) β subunit, or SCN5A with mutant β subunit: all 4 mutants altered SCN5A current to a variable extent compared to WT β subunits. WT β1 increased SCN5A currents by 75%, and induced a negative shift in steady-state activation (−10.2 mV) and inactivation (−6.7 mV), compared to SCN5A alone. D153N β1 caused partial loss of function, with increased SCN5A current but to a smaller extent (24%) than WT β1, and a negative shift in steady-state activation (−12.1 mV) and inactivation (−8.1 mV) similar to WT. R85H β1 produced a pure loss of function, with currents no different from SCN5A alone. WT β2 did not change SCN5A current amplitude, while R28Q β2 and R28W β2 decreased current by 36% and 30%, respectively; and positively shifted steady-state activation by +7.4 mV and +5.1 mV, respectively, compared to WT. Conclusion: Loss of function mutations in sodium channel β subunits were identified in patients with AF, and were associated with a distinctive ECG phenotype. These findings further support the hypothesis that decreased sodium current enhances AF susceptibility.

Effects of Halothane and Isoflurane on Fast and Slow Inactivation of Human Heart hH1a Sodium Channels

1999 ◽  
Vol 90 (6) ◽  
pp. 1671-1683. ◽  
Author(s):  
Anna Stadnicka ◽  
Wai-Meng Kwok ◽  
Hali A. Hartmann ◽  
Zeljko J. Bosnjak
Keyword(s):  
Steady State ◽  
Heterologous Expression ◽  
Sodium Channel ◽  
Sodium Channels ◽  
Human Heart ◽  
Sodium Current ◽  
Volatile Anesthetic ◽  
Slow Inactivation ◽  
Fast Inactivation ◽  
Cardiac Sodium Channel

Background Cloning and heterologous expression of ion channels allow biophysical and molecular studies of the mechanisms of volatile anesthetic interactions with human heart sodium channels. Volatile anesthetics may influence the development of arrhythmias arising from cardiac sodium channel dysfunction. For that reason, understanding the mechanisms of interactions between these anesthetics and cardiac sodium channels is important. This study evaluated the mechanisms of volatile anesthetic actions on the cloned human cardiac sodium channel (hH1a) alpha subunit. Methods Inward sodium currents were recorded from human embryonic kidney (HEK293) cells stably expressing hH1a channels. The effects of halothane and isoflurane on current and channel properties were evaluated using the whole cell voltage-clamp technique. Results Halothane at 0.47 and 1.1 mM and isoflurane at 0.54 and 1.13 mM suppressed the sodium current in a dose- and voltage-dependent manner. Steady state activation was not affected, but current decay was accelerated. The voltage dependence of steady state fast and slow inactivations was shifted toward more hyperpolarized potentials. The slope factor of slow but not fast inactivation curves was reduced significantly. Halothane increased the time constant of recovery from fast inactivation. The recovery from slow inactivation was not affected significantly by either anesthetic. Conclusions In a heterologous expression system, halothane and isoflurane interact with the hH1a channels and suppress the sodium current. The mechanisms involve acceleration of the transition from the open to the inactivated state, stabilization of the fast and slow inactivated states, and prolongation of the inactivated state by delayed recovery from the fast inactivated to the resting state.

Investigations of the Navβ1b sodium channel subunit in human ventricle; functional characterization of the H162P Brugada syndrome mutant

2014 ◽  
Vol 306 (8) ◽  
pp. H1204-H1212 ◽  
Author(s):  
Lei Yuan ◽  
Jussi T. Koivumäki ◽  
Bo Liang ◽  
Lasse G. Lorentzen ◽  
Chuyi Tang ◽  
...  
Keyword(s):  
Steady State ◽  
Action Potential ◽  
Sodium Channel ◽  
Brugada Syndrome ◽  
Functional Characterization ◽  
Inherited Disease ◽  
Loss Of Function ◽  
Peak Current Density ◽  
Cardiac Sodium Channel ◽  
The Impact

Brugada syndrome (BrS) is a rare inherited disease that can give rise to ventricular arrhythmia and ultimately sudden cardiac death. Numerous loss-of-function mutations in the cardiac sodium channel Nav1.5 have been associated with BrS. However, few mutations in the auxiliary Navβ1–4 subunits have been linked to this disease. Here we investigated differences in expression and function between Navβ1 and Navβ1b and whether the H162P/Navβ1b mutation found in a BrS patient is likely to be the underlying cause of disease. The impact of Navβ subunits was investigated by patch-clamp electrophysiology, and the obtained in vitro values were used for subsequent in silico modeling. We found that Navβ1b transcripts were expressed at higher levels than Navβ1 transcripts in the human heart. Navβ1 and Navβ1b coexpressed with Nav1.5 induced a negative shift on steady state of activation and inactivation compared with Nav1.5 alone. Furthermore, Navβ1b was found to increase the current level when coexpressed with Nav1.5, Navβ1b/H162P mutated subunit peak current density was reduced by 48% (−645 ± 151 vs. −334 ± 71 pA/pF), V1/2 steady-state inactivation shifted by −6.7 mV (−70.3 ± 1.5 vs. −77.0 ± 2.8 mV), and time-dependent recovery from inactivation slowed by >50% compared with coexpression with Navβ1b wild type. Computer simulations revealed that these electrophysiological changes resulted in a reduction in both action potential amplitude and maximum upstroke velocity. The experimental data thereby indicate that Navβ1b/H162P results in reduced sodium channel activity functionally affecting the ventricular action potential. This result is an important replication to support the notion that BrS can be linked to the function of Navβ1b and is associated with loss-of-function of the cardiac sodium channel.

scholarly journals Calmodulin binds to the N-terminal domain of the cardiac sodium channel Nav1.5

2020 ◽  
Author(s):  
Zizun Wang ◽  
Sarah H. Vermij ◽  
Valentin Sottas ◽  
Anna Shestak ◽  
Daniela Ross-Kaschitza ◽  
...  
Keyword(s):  
Sodium Channel ◽  
Sodium Current ◽  
Dominant Negative ◽  
Dominant Negative Effect ◽  
Loss Of Function ◽  
Whole Cell ◽  
Calmodulin Binding ◽  
Neonatal Cardiomyocytes ◽  
Terminal Domain ◽  
Cardiac Sodium Channel

ABSTRACTThe cardiac voltage-gated sodium channel Nav1.5 conducts the rapid inward sodium current crucial for cardiomyocyte excitability. Loss-of-function mutations in its gene SCN5A are linked to cardiac arrhythmias such as Brugada Syndrome (BrS). Several BrS-associated mutations in the Nav1.5 N-terminal domain exert a dominant-negative effect (DNE) on wild-type channel function, for which mechanisms remain poorly understood. We aim to contribute to the understanding of BrS pathophysiology by characterizing three mutations in the Nav1.5 N-terminal domain (NTD): Y87C–here newly identified–, R104W and R121W. In addition, we hypothesize that the calcium sensor protein calmodulin is a new NTD binding partner.Recordings of whole-cell sodium currents in TsA-201 cells expressing WT and variant Nav1.5 showed that Y87C and R104W but not R121W exert a DNE on WT channels. Biotinylation assays revealed reduction in fully glycosylated Nav1.5 at the cell surface and in whole-cell lysates. Localization of Nav1.5 WT channel with the ER however did not change in the presence of variants, shown by transfected and stained rat neonatal cardiomyocytes. We next demonstrated that calmodulin binds Nav1.5 N-terminus using in silico modeling, SPOTS, pull-down and proximity ligation assays. This binding is impaired in the R121W variant and in a Nav1.5 construct missing residues 80-105, a predicted calmodulin binding site.In conclusion, we present the first evidence that calmodulin binds to the Nav1.5 NTD, which seems to be a determinant for the DNE.

Abstract 5675: Brugada-Type ST Segment Elevation after Sodium Channel Blockade is Caused by Right Ventricular Excitation Failure in Discontinuous Myocardium

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Mark G Hoogendijk ◽  
Mark Potse ◽  
Andre C Linnenbank ◽  
Arie O Verkerk ◽  
Hester M den Ruijter ◽  
...  
Keyword(s):  
Right Ventricular ◽  
Sodium Channel ◽  
Isolated Heart ◽  
Fatty Infiltration ◽  
Channel Blockade ◽  
St Segment Elevation ◽  
St Segment ◽  
Cardiac Sodium Channel ◽  
The Right ◽  
Unipolar Electrograms

Introduction: Brugada-type ST segment elevation has been associated with reduced sodium channel function and structural heart disease and is a risk marker for sudden cardiac death. Depolarizing and repolarizing abnormalities in the right ventricular wall have been described in patients. The mechanism of ST segment elevation however, is still debated. We determined depolarization and repolarization characteristics in an isolated heart of a young patient carrying a sodium channel mutation previously described in a family with Brugada syndrome. Methods and Results: A 15-year old female patient with a loss-of-function mutation (G752R) in the gene encoding the cardiac sodium channel underwent cardiac transplantation for end-stage heart failure in dilated cardiomyopathy. The explanted, perfused heart was submerged in perfusion fluid. We simultaneously recorded an “Einthoven” configuration pseudo-ECG and 194 unipolar electrograms from the endo- and epicardium of both ventricles. At baseline, a single shortly coupled premature stimulus resulted in more fractionated electrograms in the right than left ventricle (38 vs 6%, p < 0.001, Z-test). After sodium channel blockade by ajmaline, ST segment elevation of 0.3 mV was observed in pseudo-aVR which coincided with the virtual disappearance of the QRS complex in unipolar electrograms at the basal epicardium of the right ventricle followed by monophasic ST segment elevation. The local origin of this phenomenon was demonstrated by Laplacian electrograms. Neither early repolarization nor late activation correlated with the ST-change. Marked epicardial fatty infiltration was present at sites of local ST segment elevation, but not elsewhere. Simulations in a bidomain whole-heart computer model showed that either sodium current reduction or random replacement of 50% of the right ventricular epicardium by fat alone does not cause ST segment elevation. In combination however, part of the epicardial myocardium was not excited throughout the cardiac cycle by current-to-load mismatch resulting in Brugada-type ST segment elevation. Conclusions: Brugada-type ST segment elevation after sodium channel blockade is caused by excitation failure in discontinuous myocardium at the right ventricular epicardium.

scholarly journals Phospho-ablation of cardiac sodium channel Nav1.5 mitigates susceptibility to atrial fibrillation and improves glucose homeostasis under conditions of diet-induced obesity

2021 ◽  
Author(s):  
Revati S. Dewal ◽  
Amara Greer-Short ◽  
Cemantha Lane ◽  
Shinsuke Nirengi ◽  
Pedro Acosta Manzano ◽  
...  
Keyword(s):  
Atrial Fibrillation ◽  
Sodium Channel ◽  
Food Consumption ◽  
Glucose Homeostasis ◽  
Sodium Current ◽  
Whole Body ◽  
Late Sodium Current ◽  
Diet Induced Obesity ◽  
Cardiac Sodium Channel ◽  
Whole Body Metabolism

Abstract Background Atrial fibrillation (AF) is the most common sustained arrhythmia, with growing evidence identifying obesity as an important risk factor for the development of AF. Although defective atrial myocyte excitability due to stress-induced remodeling of ion channels is commonly observed in the setting of AF, little is known about the mechanistic link between obesity and AF. Recent studies have identified increased cardiac late sodium current (INa,L) downstream of calmodulin-dependent kinase II (CaMKII) activation as an important driver of AF susceptibility. Methods Here, we investigated a possible role for CaMKII-dependent INa,L in obesity-induced AF using wild-type (WT) and whole-body knock-in mice that ablates phosphorylation of the Nav1.5 sodium channel and prevents augmentation of the late sodium current (S571A; SA mice). Results A high-fat diet (HFD) increased susceptibility to arrhythmias in WT mice, while SA mice were protected from this effect. Unexpectedly, SA mice had improved glucose homeostasis and decreased body weight compared to WT mice. However, SA mice also had reduced food consumption compared to WT mice. Controlling for food consumption through pair feeding of WT and SA mice abrogated differences in weight gain and AF inducibility, but not atrial fibrosis, premature atrial contractions or metabolic capacity. Conclusions These data demonstrate a novel role for CaMKII-dependent regulation of Nav1.5 in mediating susceptibility to arrhythmias and whole-body metabolism under conditions of diet-induced obesity.

P1597Two novel mutations in SCN5A gene cause enhanced inactivation and lead to Brugada syndrome

2019 ◽  
Vol 40 (Supplement_1) ◽  
Author(s):  
A Zaytseva ◽  
A V Karpushev ◽  
A V Karpushev ◽  
Y Fomicheva ◽  
Y Fomicheva ◽  
...  
Keyword(s):  
Steady State ◽  
Sodium Channel ◽  
Brugada Syndrome ◽  
Slow Inactivation ◽  
Fast Inactivation ◽  
Novel Mutations ◽  
Patch Clamp Technique ◽  
Inactivation Curve ◽  
Voltage Gated ◽  
Cardiac Sodium Channel

Abstract Background Mutations in gene SCN5A, encoding cardiac potential-dependent sodium channel Nav1.5, are associated with various arrhythmogenic disorders among which the Brugada syndrome (BrS) and the Long QT syndrome (LQT) are the best characterized. BrS1 is associated with sodium channel dysfunction, which can be reflected by decreased current, impaired activation and enhanced inactivation. We found two novel mutations in our patients with BrS and explored their effect on fast and slow inactivation of cardiac sodium channel. Purpose The aim of this study was to investigate the effect of BrS (Y739D, L1582P) mutations on different inactivation processes in in vitro model. Methods Y739D and L1582P substitutions were introduced in SCN5A cDNA using site-directed mutagenesis. Sodium currents were recorded at room temperature in transfected HEK293-T cells using patch-clamp technique with holding potential −100 mV. In order to access the fast steady-state inactivation curve we used double-pulse protocol with 10 ms prepulses. To analyze voltage-dependence of slow inactivation we used two-pulse protocol with 10s prepulse, 20ms test pulse and 25ms interpulse at −100mV to allow recovery from fast inactivation. Electrophysiological measurements are presented as mean ±SEM. Results Y739D mutation affects highly conserved tyrosine 739 among voltage-gated sodium and calcium channels in the segment IIS2. Mutation L1582P located in the loop IVS4-S5, and leucine in this position is not conserved among voltage-gated channels superfamily. We have shown that Y739D leads to significant changes in both fast and slow inactivation, whereas L1582P enhanced slow inactivation only. Steady-state fast inactivation for Y739D was shifted on 8.9 mV towards more negative potentials compare with that for WT, while L1582P did not enhanced fast inactivation (V1/2 WT: −62.8±1.7 mV; Y739D: −71.7±2.3 mV; L1582P: −58.7±1.4 mV). Slow inactivation was increased for both substitutions (INa (+20mV)/INa (−100mV) WT: 0.45±0.03; Y739D: 0,34±0.09: L1582P: 0.38±0.04). Steady-state fast inactivation Conclusions Both mutations, observed in patients with Brugada syndrome, influence on the slow inactivation process. Enhanced fast inactivation was shown only for Y739D mutant. The more dramatic alterations in sodium channel biophysical characteristics are likely linked with mutated residue conservativity. Acknowledgement/Funding RSF #17-15-01292

Abstract 15925: Localization of Nav1.5 at the Intercellular Junctions Resolved by Diffraction-Unlimited Microscopy in Three Dimensions and by Correlative Light-Electron Microscopy

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Alejandra Leo-Macias ◽  
Esperanza Agullo-Pascual ◽  
Eli Rothenberg ◽  
Mario Delmar
Keyword(s):  
Electron Microscopy ◽  
Sodium Channel ◽  
Ventricular Myocytes ◽  
Sodium Current ◽  
Intercellular Junctions ◽  
Three Dimensions ◽  
Macromolecular Complex ◽  
Adult Heart ◽  
Mechanical Coupling ◽  
Cardiac Sodium Channel

Sodium current amplitude, kinetics and regulation depend on the properties of the pore-forming protein (mostly NaV1.5 in adult heart) and on the specific molecular partners with which the channel protein associates. The composition of the voltage-gated sodium channel macromolecular complex is location-specific; yet, the exact position of NaV1.5 in the subcellular landscape of the intercalated disc (ID), remains unclear. We implemented diffraction unlimited microscopy (direct stochastic optical reconstruction microscopy, or “dSTORM”) to localize the pore-forming subunit of the cardiac sodium channel NaV1.5 with a resolution of 20nm on the XY plane. In isolated adult ventricular myocytes, NaV1.5 was found in distinct semi-circular clusters. When the entire population of clusters within a 500 nm window from the ID was considered (more than 350 individual clusters analyzed), 75% of them localized to N-cadherin rich sites. NaV1.5-distal clusters were found at an average 313±15 nm from the cell end. Introducing an astigmatic lens in the light path allowed us to solve cluster location in three dimensions, at resolutions of 20 nm in XY and 40 nm in the z plane. Three-dimensional images confirmed the preferential localization at or near N-cadherin plaques, and further suggested that NaV1.5 arrives to the membrane via N-cadherin-anchored paths, most likely microtubules. In additional experiments, we developed a novel approach to correlate the image of NaV1.5 clusters by dSTORM with the cellular ultrastructure as resolved by electron microscopy on the same sample. This “correlative light-electron microscopy” method confirmed the preference of NaV1.5 clusters at sites of mechanical coupling. Overall, we provide the first ultrastructural description of NaV1.5 at the cardiac ID and its relation with the major electron-dense domains of the adult heart. Our data support a model by which microtubule-mediated delivery of NaV1.5 anchors at N-cadherin-rich sites, likely “mixed junctions” also containing desmosomal molecules (such as plakophilin-2; see Cerrone et al; Circulation 129:1092-1103, 2014) and connexin43. These findings have major implications to the understanding of sodium current disruption in diseases affecting the integrity of the ID.

scholarly journals Interatrial block as a predictor of atrial fibrillation in patients with ST-segment elevation myocardial infarction

2018 ◽  
Vol 41 (9) ◽  
pp. 1232-1237 ◽  
Author(s):  
Göksel Çinier ◽  
Ahmet İlker Tekkeşin ◽  
Duygu Genç ◽  
Ufuk Yıldız ◽  
Emrecan Parsova ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Atrial Fibrillation ◽  
St Segment Elevation ◽  
Segment Elevation Myocardial Infarction ◽  
St Segment ◽  
Interatrial Block
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