scholarly journals miR-190a-5p Partially Represses the Abnormal Electrical Activity of SCN3B in Cardiac Arrhythmias by Downregulation of IL-2

2022 ◽  
Vol 8 ◽  
Author(s):  
Qianqian Li ◽  
Ziguan Zhang ◽  
Shanshan Chen ◽  
Zhengrong Huang ◽  
Mengru Wang ◽  
...  
Keyword(s):  
Cardiac Arrhythmias ◽  
Protective Factor ◽  
Sodium Current ◽  
Interleukin 2 ◽  
Pathological Process ◽  
Cardiac Conduction ◽  
Luciferase Reporter ◽  
Cardiac Tissues ◽  
Whole Cell Patch Clamp ◽  
Gated Channel

Cardiac arrhythmias (CAs) are generally caused by disruption of the cardiac conduction system; interleukin-2 (IL-2) is a key player in the pathological process of CAs. This study aimed to investigate the molecular mechanism underlying the regulation of IL-2 and the sodium channel current of sodium voltage-gated channel beta subunit 3 (SCN3B) by miR-190a-5p in the progression of CAs. ELISA results suggested the concentration of peripheral blood serum IL-2 in patients with atrial fibrillation (AF) to be increased compared to that in normal controls; fluorescence in situ hybridization indicated that the expression of IL-2 in the cardiac tissues of patients with AF to be upregulated and that miR-190a-5p to be downregulated. Luciferase reporter assay, quantitative real-time-PCR, and whole-cell patch-clamp experiments confirmed the downregulation of IL-2 by miR-190a-5p and influence of the latter on the sodium current of SCN3B. Overall, miR-190a-5p suppressed the increase in SCN3B sodium current caused by endogenous IL-2, whereas miR-190a-5p inhibitor significantly reversed this effect. IL-2 was demonstrated to be directly regulated by miR-190a-5p. We, therefore, concluded that the miR-190a-5p/IL-2/SCN3B pathway could be involved in the pathogenesis of CAs and miR-190a-5p might acts as a potential protective factor in pathogenesis of CAs.

Abstract 17193: Inhibiting Late Sodium Current Attenuates Isoproterenol-Induced Transient Inward Current and Delayed Afterdepolarizations in Ventricular Myocytes

Circulation ◽  
2015 ◽  
Vol 132 (suppl_3) ◽  
Author(s):  
Yejia Song ◽  
Nesrine El-Bizri ◽  
Sridharan Rajamani ◽  
Luiz Belardinelli
Keyword(s):  
Ventricular Myocytes ◽  
Sodium Current ◽  
Selective Inhibition ◽  
Adrenergic Stimulation ◽  
Patch Clamp Technique ◽  
Cardiac Tissues ◽  
Inward Current ◽  
Transient Inward Current ◽  
Whole Cell Patch Clamp ◽  
Series Of Experiments

Introduction: The β-adrenergic agonist isoproterenol (ISO) is known to induce the arrhythmogenic transient inward current (I Ti ) and delayed afterdepolarization (DAD) via a stimulation of L-type Ca 2+ current. Recent studies found that ISO-induced DADs in cardiac tissues are inhibited by GS967, a selective blocker of the late Na + current (I NaL ). Thus, we hypothesize that I NaL contributes to the actions of ISO, and selective inhibition of this current will reduce ISO-induced I Ti and DADs. Methods: Transmembrane currents and action potentials of rabbit and guinea pig (GP) ventricular myocytes were recorded using the whole-cell patch-clamp technique. ISO (0.1 μM), GS967 (1 μM) and the Na + channel blocker tetrodotoxin (TTX, 3 μM) were used in the experiments. Results: In rabbit myocytes, application of ISO caused an increase in the amplitude of I NaL from -0.10±0.03 to -0.32±0.04 pA/pF (n = 17, p < 0.05). The ISO-stimulated I NaL was inhibited by GS967 and TTX. In one series of experiments, ISO increased the I NaL from -0.14±0.04 to -0.35±0.06 pA/pF, and GS967 applied in the presence of ISO reduced the current to -0.14±0.03 pA/pF (n = 9, p < 0.05). In another series of experiments, the amplitude of I NaL was increased by ISO from -0.17±0.08 to -0.41±0.09 pA/pF, and was decreased to -0.09±0.08 pA/pF when TTX was applied with ISO (n = 5, p < 0.05). Application of ISO also induced I Ti and DADs. GS967 applied in the presence of ISO inhibited the amplitude of I Ti by 52±6%, from -1.79±0.30 to -0.87±0.16 pA/pF (n = 8, p < 0.05). Consistent with the inhibition of I Ti , GS967 suppressed the amplitude of ISO-induced DADs by 56±12%, from 6.54±1.59 to 3.22±1.27 mV (n = 5, p < 0.05). Similarly, in GP myocytes ISO-induced I Ti and DADs were decreased by GS967 from -1.14±0.21 to -0.73±0.16 pA/pF (n = 7, p < 0.05) and from 7.16±0.59 to 4.67±0.24 mV (n = 5, p < 0.05), respectively. Conclusions: An increased I NaL is likely to contribute to the proarrhythmic effects of ISO in cardiac myocytes. GS967 significantly attenuated ISO-induced I NaL , I Ti and DADs, suggesting that inhibiting this current could be an effective strategy to antagonize the arrhythmogenic actions of β-adrenergic stimulation.

scholarly journals Zfhx3 decreases expression of cardiac Nav1.5 channels and inhibits sodium current

2021 ◽  
Vol 42 (Supplement_1) ◽  
Author(s):  
R Caballero ◽  
A Camara-Checa ◽  
M Rubio-Alarcon ◽  
T Crespo-Garcia ◽  
M Dago ◽  
...  
Keyword(s):  
Sodium Current ◽  
Association Studies ◽  
Luciferase Reporter ◽  
National Budget ◽  
Funding Sources ◽  
Whole Cell Patch Clamp ◽  
Reporter Assays ◽  
Cardiac Excitability ◽  
Public Grant ◽  
In Cells

Abstract Background Zfhx3 (zinc finger homeobox 3) is a transcription factor (TF) encoded by the ZFHX3 gene. GWAS and gene-based association studies showed that ZFHX3 is one of the major atrial fibrillation (AF) susceptibility-conferring genes. The sodium current (INa), carried by Nav1.5 channels encoded by SCN5A, is responsible for atrial and ventricular action potential depolarization and determines cardiac excitability. Zfhx3 interacts with other TFs, such as Tbx5 and Pitx2c that increase and decrease INa, respectively. However, the effects of Zfhx3 on cardiac INa are currently unknown. Purpose We aimed to determine the effects of Zfhx3 on the INa on HL-1 cardiomyocytes. Methods cDNAs encoding human Zfhx3 together or not with Pitx2c or Tbx5 were transfected in HL-1 cells. Endogenous Zfhx3 expression in HL-1 cells was silenced by means of siRNAs. INa was recorded at room temperature using the whole-cell patch-clamp and luciferase reporter assays, qPCR and Western-blot (WB) analyses were also conducted. Results Expression analysis of RNA-seq data from human ventricular (n=432) samples (GTEx) demonstrated that Zfhx3 mRNA can be detected in the adult working myocardium. Transfection of Zfhx3 strongly reduced peak INa density (from −75.0±6.6 to −30.9±2.9 pA/pF; n≥26, P&lt;0.001). In contrast, Zfhx3 silencing augmented INa density compared to cells transfected with scrambled siRNA (from −65.9±8.9 to −104.6±10.8 pA/pF; n≥8, P&lt;0.05). Neither Zfhx3 expression nor silencing modified time and voltage dependence of activation and inactivation or the reactivation kinetics. Zfhx3 significantly reduced transcriptional activity of human SCN5A, PITX2 and TBX5 minimal promoters and, consequently, the mRNA and protein expression levels of Nav1.5, Pitx2c, and Tbx5 were diminished (n≥6, P&lt;0.05). In cells transfected with Zfhx3 together with Pitx2c, but not with Tbx5, INa density was significantly smaller than in cells expressing WT Zfhx3 alone (n≥15, P&lt;0.05). Further WB experiments demonstrated that Zfhx3 increased the expression of Nedd4–2 ubiquitin-protein ligase, which ubiquitinates Nav1.5 channels and favors their proteasomal degradation. Conclusions Zfhx3 inhibits INa as a result of a direct repressor effect on the SCN5A promoter, the modulation of Tbx5-increasing and Pitx2-decreasing effects on the INa, and the enhancement of Nav1.5 channel degradation. We propose a novel and complex mechanism that regulates the expression of sodium channels and the density of the INa, which are critical for the control of cardiac excitability. FUNDunding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Ministerio de Economía y CompetitividadComunidad Autόnoma de Madrid

Acute effects of repetitive depolarization on sodium current in chick myocytes

1991 ◽  
Vol 260 (6) ◽  
pp. H1810-H1818
Author(s):  
M. R. Gold ◽  
G. R. Strichartz
Keyword(s):  
Time Course ◽  
Sodium Current ◽  
Complete Recovery ◽  
Acute Effects ◽  
Na Current ◽  
Patch Clamp Technique ◽  
Membrane Hyperpolarization ◽  
Atrial Cells ◽  
Whole Cell Patch Clamp ◽  
Recovery From Inactivation

Acute effects of repetitive depolarization on the inward Na+ current (INa) of cultured embryonic chick atrial cells were studied using the whole cell patch-clamp technique. Stimulation rates of 1 Hz or greater produced a progressive decrement of peak INa. With depolarizations to 0 mV of 150-ms duration, applied at 2 Hz from a holding potential of -100 mV, the steady-state decrement was approximately 20%. The magnitude of this effect increased with stimulation frequency and with test potential depolarization and decreased with membrane hyperpolarization. Analysis of INa kinetics revealed that reactivation was sufficiently slow to preclude complete recovery from inactivation with interpulse intervals less than 1,000 ms. Moreover, reactivation accelerated markedly with membrane hyperpolarization, in parallel with the response to repetitive stimulation. The multiexponential time course of recovery of peak INa from repetitive depolarization was similar to that observed after single stimuli; however, there was a shift toward a greater proportion of current recovering with the slower of two time constants. It is concluded that incomplete recovery from inactivation is responsible for the decrement in INa observed with short interpulse intervals.

scholarly journals Irbesartan prevents sodium channel remodeling in a canine model of atrial fibrillation

2018 ◽  
Vol 19 (1) ◽  
pp. 147032031875526 ◽  
Author(s):  
Xuewen Wang ◽  
Guangping Li
Keyword(s):  
Atrial Fibrillation ◽  
Sodium Current ◽  
Renin Angiotensin System ◽  
Canine Model ◽  
Atrial Pacing ◽  
Chain Reaction ◽  
Patch Clamp Technique ◽  
Angiotensin System ◽  
Whole Cell Patch Clamp ◽  
Polymerase Chain

Introduction: Activation of the renin-angiotensin system (RAS) plays an important role in atrial electrical remodeling (AER). The purpose of the present study was to evaluate the effects of irbesartan on cardiac sodium current (INa) in a canine model of atrial fibrillation. Materials and methods: Eighteen dogs were randomized into sham, pacing or pacing+irbesartan groups ( n = 6 in each group). The dogs in the pacing and irbesartan group were paced at 500 bpm for two weeks. Irbesartan (60 mg·kg−1·d−1) was administered orally in the pacing+irbesartan groups. INa was recorded using the whole-cell patch clamp technique from canine atrial myocytes. The expressions of cardiac Na+ channels (Nav1.5) mRNA were semi-quantified by reverse transcription-polymerase chain reaction. Results: Our results showed that INa density and Nav1.5 mRNA expression in the pacing group decreased significantly ( p < 0.05 vs. sham). However, rapid atrial pacing had no effects on the half-activation voltage (V1/2act) and half-inactivation voltage (V1/2inact) of INa ( p > 0.05 vs. sham). Irbesartan significantly increased INa densities and gene expression and hyperpolarized V1/2act without concomitant changes in V1/2inact. Conclusions: Irbesartan significantly increased INa densities, which contributed to improving intra-atrial conduction and prevented the induction and promotion of AF in atrial pacing dogs.

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

2007 ◽  
Vol 292 (1) ◽  
pp. H399-H407 ◽  
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.

Is there a second external lidocaine binding site on mammalian cardiac cells?

1989 ◽  
Vol 257 (1) ◽  
pp. H79-H84 ◽  
Author(s):  
L. A. Alpert ◽  
H. A. Fozzard ◽  
D. A. Hanck ◽  
J. C. Makielski
Keyword(s):  
Binding Site ◽  
Cardiac Arrhythmias ◽  
Local Anesthetics ◽  
Sodium Current ◽  
Cardiac Cells ◽  
Na Channel ◽  
Functional Difference ◽  
Na Channels ◽  
Internal Perfusion ◽  
Cardiac Purkinje Cells

Lidocaine and its permanently charged analogue QX-314 block sodium current (INa) in nerve, and by this mechanism, lidocaine produces local anesthesia. When administered clinically, lidocaine prevents cardiac arrhythmias. Nerve and skeletal muscle are much more sensitive to local anesthetics when the drugs are applied inside the cell, indicating that the binding site for local anesthetics is located on the inside of those Na channels. Using a large suction pipette for voltage clamp and internal perfusion of single cardiac Purkinje cells, we demonstrate that a charged lidocaine analogue blocks INa not only when applied from the inside but also from the outside, unlike noncardiac tissue. This functional difference in heart predicts that a second local anesthetic binding site exists outside or near the outside of cardiac Na channels and emphasizes that the cardiac Na channel is different from that in nerve.

A role for a background sodium current in spontaneous action potentials and secretion from rat lactotrophs

1996 ◽  
Vol 271 (6) ◽  
pp. C1927-C1934 ◽  
Author(s):  
S. Sankaranarayanan ◽  
S. M. Simasko
Keyword(s):  
Action Potentials ◽  
Sodium Current ◽  
Plaque Assay ◽  
Input Resistance ◽  
Cytosolic Calcium ◽  
Calcium Dependent ◽  
Whole Cell Patch Clamp ◽  
Spontaneous Action ◽  
Spontaneous Action Potentials ◽  
Spontaneous Secretion

We have used the perforated-patch variation of whole cell patch-clamp techniques, measurements of cytosolic calcium with use of fura 2, and secretion measurements with use of the reverse-hemolytic plaque assay to address the role of depolarizing background currents in maintaining spontaneous action potentials and spontaneous secretion from rat lactotrophs in primary culture. Replacement of bath sodium with tris(hydroxymethyl)aminomethane or N-methyl-D-glucamine caused a dramatic hyperpolarization of the cells, a cessation of spontaneous action potentials, and an increase in input resistance of cells. Tetrodotoxin had no effect on spontaneous action potentials, and removal of bath calcium stopped spiking but did not hyperpolarize the cells. The hyperpolarization in response to removal of bath sodium was associated with a decrease in cytosolic calcium levels. Finally, removal of bath sodium caused a decrease in spontaneous secretion of prolactin from lactotrophs. These data suggest that a background sodium current is essential to drive the membrane to threshold for firing spontaneous calcium-dependent action potentials in lactotrophs. This, in turn, results in elevated intracellular calcium, which supports spontaneous secretion of prolactin from these cells.

scholarly journals MiR-19b-3p Regulates MAPK1 Expression in Embryonic Fibroblasts from the Great Tit (Parus Major) Under Hypoxic Conditions

2018 ◽  
Vol 46 (2) ◽  
pp. 546-560 ◽  
Author(s):  
Xiumin Chen ◽  
Yanhua Qu ◽  
Yalin Cheng ◽  
Jing Wang ◽  
Xiaohua Lei ◽  
...  
Keyword(s):  
High Altitude ◽  
Target Genes ◽  
Parus Major ◽  
Computational Prediction ◽  
Mapk Signaling ◽  
Great Tit ◽  
Luciferase Reporter ◽  
Embryonic Fibroblasts ◽  
Cardiac Tissues ◽  
Hypoxic Conditions

Background/Aims: Genomic adaptations to high altitudes have been well studied in the last several years; however, the roles of microRNAs (miRNAs), which are essential modulators of a variety of genes and key cellular processes, have rarely been explored. Here, we explored the interactions between miRNAs and their target genes as an adaptation to high altitude in an avian species, the great tit (Parus major), which is widely distributed across the Eurasian continent at altitudes between 4500 m and sea level. Because the MAPK signaling pathway plays a crucial role in the hypoxia response in the great tit, we chose MAPK1 as a target candidate gene. Methods: We established a great tit embryonic fibroblast line and subsequently studied the relationship between miRNA-19b-3p and MAPK1 in normoxia and hypoxia groups. Meanwhile, the great tit embryonic fibroblasts (GEFs) were treated or transfected with miR-19b-3p mimics, inhibitors, or si-MAPK1, and their proliferation was subsequently assessed using the MTT assay. The expression of the miRNAs and MAPK1 was measured by real-time PCR and Western blotting. Results: We identified 14 miRNAs in the cardiac tissues of great tits that are related to hypoxia adaptation. MAPK1 binds only to miR-19b-3p of the 14 miRNAs predicted by both TargetScan and miRanda software. Specifically, we validated the computational prediction of miR-19b-3p binding to the 3’UTR of MAPK1 using a luciferase reporter assay. Our results show that miR-19b-3p promotes GEFs proliferation and up-regulates MAPK1 expression. Moreover, miR-19b-3p mimics and MAPK1 knockdown induce GEFs apoptosis and regulate the cell cycle under hypoxic conditions. Conclusions: Our study is the first to describe an important miRNA-mediated regulatory mechanism of high altitude adaptation in a non-model wild songbird and highlights the importance of studies on miRNA-mediated mechanisms of hypoxic adaptations in other animals.

scholarly journals CircSERPINE2 protects against osteoarthritis by targeting miR-1271 and ETS-related gene

2019 ◽  
Vol 78 (6) ◽  
pp. 826-836 ◽  
Author(s):  
Shuying Shen ◽  
Yizheng Wu ◽  
Junxin Chen ◽  
Ziang Xie ◽  
Kangmao Huang ◽  
...  
Keyword(s):  
In Situ Hybridisation ◽  
Rabbit Model ◽  
Pathological Process ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Fluorescence In Situ Hybridisation ◽  
Related Gene ◽  
Downstream Target

ObjectivesCircular RNAs (circRNA) expression aberration has been identified in various human diseases. In this study, we investigated whether circRNAs could act as competing endogenous RNAs to regulate the pathological process of osteoarthritis (OA).MethodsCircRNA deep sequencing was performed to the expression of circRNAs between OA and control cartilage tissues. The regulatory and functional role of CircSERPINE2 upregulation was examined in OA and was validated in vitro and in vivo, downstream target of CircSERPINE2 was explored. RNA pull down, a luciferase reporter assay, biotin-coupled microRNA capture and fluorescence in situ hybridisation were used to evaluate the interaction between CircSERPINE2 and miR-1271-5 p, as well as the target mRNA, E26 transformation-specific-related gene (ERG). The role and mechanism of CircSERPINE2 in OA was also explored in rabbit models.ResultsThe decreased expression of CircSERPINE2 in the OA cartilage tissues was directly associated with excessive apoptosis and imbalance between anabolic and catabolic factors of extracellular matrix (ECM). Mechanistically, CircSERPINE2 acted as a sponge of miR-1271-5 p and functioned in human chondrocytes (HCs) through targeting miR-1271-5 p and ERG. Intra-articular injection of adeno-associated virus-CircSERPINE2-wt alleviated OA in the rabbit model.ConclusionsOur results reveal an important role for a novel circRNA-CircSERPINE2 in OA progression. CircSERPINE2 overexpression could alleviate HCs apoptosis and promote anabolism of ECM through miR-1271-ERG pathway. It provides a potentially effective therapeutic strategy for OA progression.

scholarly journals Mural propagation of descending vasa recta responses to mechanical stimulation

2013 ◽  
Vol 305 (3) ◽  
pp. F286-F294 ◽  
Author(s):  
Zhong Zhang ◽  
Kristie Payne ◽  
Chunhua Cao ◽  
Thomas L. Pallone
Keyword(s):  
Membrane Potential ◽  
Patch Clamp ◽  
Mechanical Stimulation ◽  
Channel Blockade ◽  
Patch Clamp Recording ◽  
Automated Method ◽  
Whole Cell Patch Clamp ◽  
Vasa Recta ◽  
Gated Channel ◽  
Phosphoinositide 3 Kinase

To investigate the responses of descending vasa recta (DVR) to deformation of the abluminal surface, we devised an automated method that controls duration and frequency of stimulation by utilizing a stream of buffer from a micropipette. During stimulation at one end of the vessel, fluorescent responses from fluo4 or bis[1,3-dibutylbarbituric acid-(5)] trimethineoxonol [DiBAC4(3)], indicating cytoplasmic calcium ([Ca2+]CYT) or membrane potential, respectively, were recorded from distant cells. Alternately, membrane potential was recorded from DVR pericytes by nystatin whole cell patch-clamp. Mechanical stimulation elicited reversible [Ca2+]CYT responses that increased with frequency. Individual pericyte responses along the vessel were initiated within a fraction of a second of one another. Those responses were inhibited by gap junction blockade with 18 β-glycyrrhetinic acid (100 μM) or phosphoinositide 3 kinase inhibition with 2-morpholin-4-yl-8-phenylchromen-4-one (50 μM). [Ca2+]CYT responses were blocked by removal of extracellular Ca2+ or L-type voltage-gated channel blockade with nifedipine (10 μM). At concentrations selective for the T-type channel blockade, mibefradil (100 nM) was ineffective. During mechanostimulation, pericytes rapidly depolarized, as documented with either DiBAC4(3) fluorescence or patch-clamp recording. Single stimuli yielded depolarizations of 22.5 ± 2.2 mV while repetitive stimuli at 0.1 Hz depolarized pericytes by 44.2 ± 4.0 mV. We conclude that DVR are mechanosensitive and that rapid transmission of signals along the vessel axis requires participation of gap junctions, L-type Ca2+ channels, and pericyte depolarization.

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