REGULATION OF VOLTAGE-GATED SODIUM CURRENT BY WNT SIGNALLING IN HEALTHY AND BRUGADA SYNDROME HUMAN CARDIOMYOCYTES

Esaxerenone (ESAX; CS-3150, Minnebro®) is known to be a newly non-steroidal mineralocorticoid receptor (MR) antagonist. However, its modulatory actions on different types of ionic currents in electrically excitable cells remain largely unanswered. The present investigations were undertaken to explore the possible perturbations of ESAX on the transient, late and persistent components of voltage-gated Na+ current (INa) identified from pituitary GH3 or MMQ cells. GH3-cell exposure to ESAX depressed the transient and late components of INa with varying potencies. The IC50 value of ESAX required for its differential reduction in peak or late INa in GH3 cells was estimated to be 13.2 or 3.2 μM, respectively. The steady-state activation curve of peak INa remained unchanged during exposure to ESAX; however, recovery of peak INa block was prolonged in the presence 3 μM ESAX. In continued presence of aldosterone (10 μM), further addition of 3 μM ESAX remained effective at inhibiting INa. ESAX (3 μM) potently reversed Tef-induced augmentation of INa. By using isosceles-triangular ramp pulse with varying durations, the amplitude of persistent INa measured at high or low threshold was enhanced by the presence of tefluthrin (Tef), in combination with the appearance of the figure-of-eight hysteretic loop; moreover, hysteretic strength of the current was attenuated by subsequent addition of ESAX. Likewise, in MMQ lactotrophs, the addition of ESAX also effectively decreased the peak amplitude of INa along with the increased current inactivation rate. Taken together, the present results provide a noticeable yet unidentified finding disclosing that, apart from its antagonistic effect on MR receptor, ESAX may directly and concertedly modify the amplitude, gating properties and hysteresis of INa in electrically excitable cells.

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

European Heart Journal ◽

10.1093/eurheartj/ehz748.0356 ◽

2019 ◽

Vol 40 (Supplement_1) ◽

Author(s):

A Zaytseva ◽

A V Karpushev ◽

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

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Nongenomic Thyroxine Regulation of Voltage-gated Sodium Current During Embryonic Development

Immunology Endocrine & Metabolic Agents - Medicinal Chemistry ◽

10.2174/187152211796642800 ◽

2011 ◽

Vol 11 (3) ◽

pp. 157-164 ◽

Cited By ~ 3

Author(s):

Marc A. Yonkers ◽

Angeles B. Ribera

Keyword(s):

Embryonic Development ◽

Sodium Current ◽

Voltage Gated

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Protease-Activated Receptor-1 Mediates Thrombin-Induced Persistent Sodium Current in Human Cardiomyocytes

Molecular Pharmacology ◽

10.1124/mol.107.043182 ◽

2008 ◽

Vol 73 (6) ◽

pp. 1622-1631 ◽

Cited By ~ 17

Author(s):

Caroline Pinet ◽

Vincent Algalarrondo ◽

Sylvie Sablayrolles ◽

Bruno Le Grand ◽

Christophe Pignier ◽

...

Keyword(s):

Sodium Current ◽

Persistent Sodium Current ◽

Human Cardiomyocytes ◽

Protease Activated Receptor 1

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Sodium channel kinetic changes that produce Brugada syndrome or progressive cardiac conduction system disease

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.01025.2005 ◽

2007 ◽

Vol 292 (1) ◽

pp. H399-H407 ◽

Cited By ~ 33

Author(s):

Zhu-Shan Zhang ◽

Joseph Tranquillo ◽

Valentina Neplioueva ◽

Nenad Bursac ◽

Augustus O. Grant

Keyword(s):

Action Potential ◽

Sodium Channel ◽

Brugada Syndrome ◽

Sodium Current ◽

Conduction System ◽

Cardiac Conduction ◽

Cardiac Conduction System ◽

Wild Type ◽

System Disease ◽

Conduction System Disease

Some mutations of the sodium channel gene NaV1.5 are multifunctional, causing combinations of LQTS, Brugada syndrome and progressive cardiac conduction system disease (PCCD). The combination of Brugada syndrome and PCCD is uncommon, although they both result from a reduction in the sodium current. We hypothesize that slow conduction is sufficient to cause S-T segment elevation and undertook a combined experimental and theoretical study to determine whether conduction slowing alone can produce the Brugada phenotype. Deletion of lysine 1479 in one of two positively charged clusters in the III/IV inter-domain linker causes both syndromes. We have examined the functional effects of this mutation using heterologous expression of the wild-type and mutant sodium channel in HEK-293-EBNA cells. We show that ΔK1479 shifts the potential of half-activation, V1/2m, to more positive potentials ( V1/2m = −36.8 ± 0.8 and −24.5 ± 1.3 mV for the wild-type and ΔK1479 mutant respectively, n = 11, 10). The depolarizing shift increases the extent of depolarization required for activation. The potential of half-inactivation, V1/2h, is also shifted to more positive potentials ( V1/2h = −85 ± 1.1 and −79.4 ± 1.2 mV for wild-type and ΔK1479 mutant respectively), increasing the fraction of channels available for activation. These shifts are quantitatively the same as a mutation that produces PCCD only, G514C. We incorporated experimentally derived parameters into a model of the cardiac action potential and its propagation in a one dimensional cable (simulating endo-, mid-myocardial and epicardial regions). The simulations show that action potential and ECG changes consistent with Brugada syndrome may result from conduction slowing alone; marked repolarization heterogeneity is not required. The findings also suggest how Brugada syndrome and PCCD which both result from loss of sodium channel function are sometimes present alone and at other times in combination.

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Abstract 5317: Identification and Characterization of a Novel Susceptibility Gene, SCN3B, for Idiopathic Ventricular Fibrillation

Circulation ◽

10.1161/circ.118.suppl_18.s_526 ◽

2008 ◽

Vol 118 (suppl_18) ◽

Author(s):

Carmen R Valdivia ◽

Argelia Mereidos-Domingo ◽

Thimothy J Algiers ◽

Michael J Ackerman ◽

Jonathan C Makielski

Keyword(s):

Ventricular Fibrillation ◽

Sodium Channel ◽

Brugada Syndrome ◽

Mutational Analysis ◽

Sodium Current ◽

Susceptibility Gene ◽

Site Directed Mutagenesis ◽

Macromolecular Complex ◽

Patch Clamp Technique ◽

Idiopathic Ventricular Fibrillation

Background: Mutations in the Na V 1.5 sodium channel macromolecular complex have been identified in some cases classified as idiopathic ventricular fibrillation (IVF). IVF and Brugada syndrome (BrS) are partially overlapping syndromes. Here, we report a mutation in SCN3B- encoded sodium channel β3 subunit as a novel pathogenic mechanism for IVF. Methods: Comprehensive open reading frame mutational analysis of SCN5A, GPD1L, and the beta subunit genes ( SCN1–4B ) was performed using PCR, DHPLC, and direct DNA sequencing of DNA extracted from a 20-year-old patient diagnosed with IVF. The SCN3B mutation was made by site directed mutagenesis and co-transfected with SCN5A into HEK-293 cells for functional chraracterization using the patch clamp technique. Results: A novel missense mutation, V54G-SCN3B, was identified in a 20-year-old male following collapse and external defibrillation from VF. After recovery, there was no detectable electrocardiographic abnormality. Imaging studies demonstrated a structurally normal heart, and the patient was diagnosed with IVF. The mutation was absent in 800 reference alleles and involved a highly conserved residue in the extracellular domain of the beta 3 subunit. No other mutations were identified in the 5 other genes. HEK cells expressing SCN5A and either WT-, or V54G-SCN3B were studied 24 hours after transfection. Cells expressing V54G-SCN3B showed significant decrease in sodium current density of 60±20 pA/pF compared to 203±35 pA/pF in WT-SCN3B (n=14–19). In addition V54G-SCN3B significantly shifted the activation curve +5 mV without affecting inactivation. Conclusions: This study provides the first molecular and cellular evidence implicating SCN3B in IVF. Given the marked loss-of-function to the sodium channel by V54G-SCN3B and the overlap between IVF and BrS, it will be interesting to determine whether mutations in SCN3B explain some cases of genotype negative Brugada syndrome.

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