scholarly journals Inhibition of minor intron splicing reduces Na+ and Ca2+ channel expression and function in cardiomyocytes

2021 ◽  
Author(s):  
Pablo Montañés-Agudo ◽  
Simona Casini ◽  
Simona Aufiero ◽  
Auriane C. Ernault ◽  
Ingeborg van der Made ◽  
...  
Keyword(s):  
Ventricular Myocytes ◽  
Intron Retention ◽  
Genetic Diseases ◽  
Gene Families ◽  
Calcium Currents ◽  
Neonatal Rat ◽  
Intron Splicing ◽  
Voltage Gated ◽  
Channel Expression ◽  
And Function

Eukaryotic genomes contain a tiny subset of ‘minor class’ introns with unique sequence elements that require their own splicing machinery. These minor introns are present in certain gene families with specific functions, such as voltage-gated sodium and voltage-gated calcium channels. Removal of minor introns by the minor spliceosome has been proposed as a post-transcriptional regulatory layer, which remains unexplored in the heart. Here, we investigate whether the minor spliceosome regulates electrophysiological properties of cardiomyocytes by knocking-down the essential minor spliceosome component U6atac in neonatal rat ventricular myocytes. Loss of U6atac led to robust minor intron retention within Scn5a and Cacna1c, resulting in reduced protein levels of Nav1.5 and Cav1.2. Functional consequences were studied through path-clamp analysis, and revealed reduced sodium and L-type calcium currents after loss of U6atac. In conclusion, minor intron splicing modulates voltage-dependent ion channel expression and function in cardiomyocytes. This may be of particular relevance in situations in which minor splicing activity changes, such as in genetic diseases affecting minor spliceosome components, or in acquired diseases in which minor spliceosome components are dysregulated, such as heart failure.

scholarly journals MicroRNA-204 Is Necessary for Aldosterone-Stimulated T-Type Calcium Channel Expression in Cardiomyocytes

2018 ◽  
Vol 19 (10) ◽  
pp. 2941 ◽  
Author(s):  
Riko Koyama ◽  
Tiphaine Mannic ◽  
Jumpei Ito ◽  
Laurence Amar ◽  
Maria-Christina Zennaro ◽  
...  
Keyword(s):  
Calcium Channels ◽  
Molecular Mechanisms ◽  
Calcium Currents ◽  
Neonatal Rat ◽  
Messenger Rnas ◽  
Screening Assay ◽  
Rat Cardiomyocytes ◽  
Ventricular Cardiomyocytes ◽  
Channel Expression ◽  
Beating Frequency

Activation of the mineralocorticoid receptor (MR) in the heart is considered to be a cardiovascular risk factor. MR activation leads to heart hypertrophy and arrhythmia. In ventricular cardiomyocytes, aldosterone induces a profound remodeling of ion channel expression, in particular, an increase in the expression and activity of T-type voltage-gated calcium channels (T-channels). The molecular mechanisms immediately downstream from MR activation, which lead to the increased expression of T-channels and, consecutively, to an acceleration of spontaneous cell contractions in vitro, remain poorly investigated. Here, we investigated the putative role of a specific microRNA in linking MR activation to the regulation of T-channel expression and cardiomyocyte beating frequency. A screening assay identified microRNA 204 (miR-204) as one of the major upregulated microRNAs after aldosterone stimulation of isolated neonatal rat cardiomyocytes. Aldosterone significantly increased the level of miR-204, an effect blocked by the MR antagonist spironolactone. When miR-204 was overexpressed in isolated cardiomyocytes, their spontaneous beating frequency was significantly increased after 24 h, like upon aldosterone stimulation, and messenger RNAs coding T-channels (CaV3.1 and CaV3.2) were increased. Concomitantly, T-type calcium currents were significantly increased upon miR-204 overexpression. Specifically repressing the expression of miR-204 abolished the aldosterone-induced increase of CaV3.1 and CaV3.2 mRNAs, as well as T-type calcium currents. Finally, aldosterone and miR-204 overexpression were found to reduce REST-NRSF, a known transcriptional repressor of CaV3.2 T-type calcium channels. Our study thus strongly suggests that miR-204 expression stimulated by aldosterone promotes the expression of T-channels in isolated rat ventricular cardiomyocytes, and therefore, increases the frequency of the cell spontaneous contractions, presumably through the inhibition of REST-NRSF protein.

Abstract 17384: Cardiac-Specific Overexpression of Human Mutant Desmoplakin in Mice Disrupts Cardiac Voltage-Gated Sodium Channel Expression

Circulation ◽  
2018 ◽  
Vol 138 (Suppl_1) ◽  
Author(s):  
Subat Turdi ◽  
Jeffrey A Towbin
Keyword(s):  
Sodium Channel ◽  
Fatty Infiltration ◽  
Electrical Coupling ◽  
Cx43 Expression ◽  
Voltage Gated Sodium Channel ◽  
Voltage Gated ◽  
Intercalated Discs ◽  
Channel Expression ◽  
Life Threatening ◽  
And Function

Introduction: Arrhythmogenic cardiomyopathy (AC) is characterized by bi-ventricular dilation, fibro-fatty infiltration and life-threatening arrhythmias. Disruptions in cardiac voltage-gated sodium channel (Nav1.5) expression and function are known to cause arrhythmias. We have demonstrated that cardiac-specific overexpression of human mutant desmoplakin (DSP, Tg-R2834H) in mice leads to AC. However, whether mutant DSP expression in the heart affects the Nav1.5 distribution and function are unknown Hypothesis: Here, we tested whether Nav1.5 localization and expression are altered in the R2834H-Tg mouse hearts. Methods: Primary cardiomyocytes and frozen myocardial sections from non-transgenic (NTg), wild-type DSP (Tg-DSP) and Tg-R2834H mice were used for immunofluorescence studies to assess subcellular localization of DSP, desmin, Nav1.5, Cx43, plakoglobin and β-catenin. Western blot and qPCR were used for quantitative analysis. Results: Double staining of cardiomyocytes from NTg mice with DSP and Nav1.5 revealed that Nav1.5 was colocalized with DSP at the intercalated discs (IDs). In contrast, Tg-R2834H cardiomyocytes exhibited marked increase of mutant DSP expression at the IDs concomitant with a reduction in Nav1.5 immunoreactive signals. Tg-R2834H cardiomyocytes also revealed an aberration of DSP and desmin colocalizations at the IDs. There were not obvious differences in Cx43 expression between the genotypes, although the redistribution of Cx43 from the IDs to the sarcolemma was evident in Tg-R2834H cardiomyocytes. qPCR results correlated with reduced Nav1.5 mRNA expression in the Tg-R2834H mouse hearts. Conclusions: Defective DSP protein expression in the heart disrupts Nav1.5 localization and expression, implying an interaction between DSP and Nav1.5 to orchestrate normal mechanical and electrical coupling. Further electrophysiology studies to assess whole-cell Na + currents in these cardiomyocytes will provide insight into DSP and Nav1.5 interaction.

Abstract 1514: RXP-E: A Connexin43-Binding Peptide that Prevents Action Potential Propagation Block

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Rebecca Lewandowski ◽  
Kristina Procida ◽  
Ravi Vaidyanathan ◽  
José Jalife ◽  
Morten S Nielsen ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Action Potential ◽  
Rhode Island ◽  
Ventricular Myocytes ◽  
Heart Association ◽  
Calcium Currents ◽  
Neonatal Rat ◽  
List Type ◽  
Resistance Pathway

Gap junctions (GJ’s) provide a low resistance pathway for cardiac electrical propagation. The role of GJ regulation in arrhythmias is unclear, partly due to the limited availability of pharmacological tools. Recently, we showed that a peptide called “RXP-E” binds to the carboxyl terminal of connexin43 (Cx43) and prevents chemically-induced uncoupling in Cx43-expressing N2a cells. Here, we used pull-down experiments to show that RXP-E binds to adult cardiac Cx43. Patch-clamp studies revealed that RXP-E prevented heptanol-induced and acidification-induced uncoupling in pairs of neonatal rat ventricular myocytes (NRVM’s). Separately, RXP-E was concatenated to a cytoplasmic transduction peptide for translocation into the cytoplasm (construct CTP-RXP-E). The effect of RXP-E on action potential (AP) propagation was assessed by high resolution optical mapping in monolayers of NRVMs, containing approximately 20% of randomly distributed myofibroblasts. Conduction velocity (CV) was 164 ± 8mm/sec (avg±SEM; n=12; pacing frequency 2Hz) in untreated cells, and 158±10mm/sec, (n=6) and 180±7mm/sec (n=10) in monolayers treated with CTP-RXPE or a scrambled version of the peptide (CTP-Scr), respectively (pNS). Exposure of either untreated, or CTP-Scr-treated monolayers to heptanol caused propagation block. However, when heptanol (2 mmol/L) was added to the superfusate of monolayers loaded with CTP-RXP-E, AP propagation was maintained, albeit at a slower velocity (87±5mm/sec;n=4; P<0.001). Similarly, intracellular acidification (pHi=6.2) caused a loss of AP propagation in control or CTP-Scr monolayers; however, propagation was maintained in CTP-RXP-E treated cells, though at a slower rate (CV=93 ± 28mm/sec; n=4). Consistent with these results, patch clamp experiments revealed that RXP-E did not prevent heptanol-induced block of sodium or calcium currents, nor did it alter the voltage dependence or amplitude of Kir2.1/Kir2.3 currents. RXP-E is the first synthetic molecule known to: bind cardiac Cx43; prevent heptanol and acidification-induced uncoupling of cardiac GJ’s and preserve AP propagation among cardiac myocytes. RXP-E can be used to characterize the role of GJ’s in the function of multicellular systems, including the heart. This research has received full or partial funding support from the American Heart Association, AHA Founders Affiliate (Connecticut, Maine, Massachusetts, New Hampshire, New Jersey, New York, Rhode Island, Vermont).

scholarly journals Voltage Gated Proton Channel Expression and Function in Breast Cancer Cells

2017 ◽  
Vol 112 (3) ◽  
pp. 333a
Author(s):  
Dan Bare ◽  
Vladimir V. Cherny ◽  
Abde M. Abukhdeir ◽  
Thomas E. DeCoursey ◽  
Deri Morgan
Keyword(s):  
Breast Cancer ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Proton Channel ◽  
Voltage Gated ◽  
Channel Expression ◽  
And Function

scholarly journals Voltage-gated sodium channel expression and function in injured and uninjured sensory ganglia neurons

2012 ◽  
Vol 13 (4) ◽  
pp. S41
Author(s):  
B. Moyer ◽  
R. Yin ◽  
D. Liu ◽  
M. Chhoa ◽  
C. Li ◽  
...  
Keyword(s):  
Sodium Channel ◽  
Sensory Ganglia ◽  
Voltage Gated Sodium Channel ◽  
Voltage Gated ◽  
Channel Expression ◽  
And Function

scholarly journals Caveolin-3 KO disrupts t-tubule structure and decreases t-tubular ICa density in mouse ventricular myocytes

2018 ◽  
Vol 315 (5) ◽  
pp. H1101-H1111 ◽  
Author(s):  
Simon M. Bryant ◽  
Cherrie H. T. Kong ◽  
Judy J. Watson ◽  
Hanne C. Gadeberg ◽  
David M. Roth ◽  
...  
Keyword(s):  
Cardiac Hypertrophy ◽  
Ventricular Myocytes ◽  
Wild Type ◽  
Tubule Formation ◽  
Membrane Area ◽  
Cellular Hypertrophy ◽  
Channel Expression ◽  
T Tubules ◽  
And Function

Caveolin-3 (Cav-3) is a protein that has been implicated in t-tubule formation and function in cardiac ventricular myocytes. In cardiac hypertrophy and failure, Cav-3 expression decreases, t-tubule structure is disrupted, and excitation-contraction coupling is impaired. However, the extent to which the decrease in Cav-3 expression underlies these changes is unclear. We therefore investigated the structure and function of myocytes isolated from the hearts of Cav-3 knockout (KO) mice. These mice showed cardiac dilatation and decreased ejection fraction in vivo compared with wild-type control mice. Isolated KO myocytes showed cellular hypertrophy, altered t-tubule structure, and decreased L-type Ca2+ channel current ( ICa) density. This decrease in density occurred predominantly in the t-tubules, with no change in total ICa, and was therefore a consequence of the increase in membrane area. Cav-3 KO had no effect on L-type Ca2+ channel expression, and C3SD peptide, which mimics the scaffolding domain of Cav-3, had no effect on ICa in KO myocytes. However, inhibition of PKA using H-89 decreased ICa at the surface and t-tubule membranes in both KO and wild-type myocytes. Cav-3 KO had no significant effect on Na+/Ca2+ exchanger current or Ca2+ release. These data suggest that Cav-3 KO causes cellular hypertrophy, thereby decreasing t-tubular ICa density. NEW & NOTEWORTHY Caveolin-3 (Cav-3) is a protein that inhibits hypertrophic pathways, has been implicated in the formation and function of cardiac t-tubules, and shows decreased expression in heart failure. This study demonstrates that Cav-3 knockout mice show cardiac dysfunction in vivo, while isolated ventricular myocytes show cellular hypertrophy, changes in t-tubule structure, and decreased t-tubular L-type Ca2+ current density, suggesting that decreased Cav-3 expression contributes to these changes in cardiac hypertrophy and failure.

scholarly journals Calcium and Potassium Channels in Experimental Subarachnoid Hemorrhage and Transient Global Ischemia

2012 ◽  
Vol 2012 ◽  
pp. 1-8 ◽  
Author(s):  
Marcel A. Kamp ◽  
Maxine Dibué ◽  
Toni Schneider ◽  
Hans-Jakob Steiger ◽  
Daniel Hänggi
Keyword(s):  
Subarachnoid Hemorrhage ◽  
Animal Models ◽  
Ion Channel ◽  
Potassium Channel ◽  
Global Ischemia ◽  
Voltage Gated ◽  
Delayed Cerebral Vasospasm ◽  
Channel Expression ◽  
Transient Global Ischemia ◽  
And Function

Healthy cerebrovascular myocytes express members of several different ion channel families which regulate resting membrane potential, vascular diameter, and vascular tone and are involved in cerebral autoregulation. In animal models, in response to subarachnoid blood, a dynamic transition of ion channel expression and function is initiated, with acute and long-term effects differing from each other. Initial hypoperfusion after exposure of cerebral vessels to oxyhemoglobin correlates with a suppression of voltage-gated potassium channel activity, whereas delayed cerebral vasospasm involves changes in other potassium channel and voltage-gated calcium channels expression and function. Furthermore, expression patterns and function of ion channels appear to differ between main and small peripheral vessels, which may be key in understanding mechanisms behind subarachnoid hemorrhage-induced vasospasm. Here, changes in calcium and potassium channel expression and function in animal models of subarachnoid hemorrhage and transient global ischemia are systematically reviewed and their clinical significance discussed.

Electrophysiological properties of neonatal rat ventricular myocytes with α1-adrenergic-induced hypertrophy

1998 ◽  
Vol 275 (2) ◽  
pp. H577-H590 ◽  
Author(s):  
John P. Gaughan ◽  
Colleen A. Hefner ◽  
Steven R. Houser
Keyword(s):  
Current Density ◽  
Ventricular Myocytes ◽  
Neonatal Rat ◽  
Electrophysiological Properties ◽  
Rat Ventricular Myocytes ◽  
Kinase C ◽  
Adrenergic Antagonist ◽  
Channel Expression ◽  
Protein Kinase C Inhibitor ◽  
Neonatal Rat Ventricular Myocytes

The electrophysiology of neonatal rat ventricular myocytes with and without hypertrophy has not been characterized. The α1-adrenergic agonist phenylephrine induced hypertrophy in neonatal rat ventricular myocytes. After 48 h of exposure to 20 μM phenylephrine, cell surface area of hypertrophied myocytes was 44% larger than control. Action potential duration was significantly longer in hypertrophy than in control. There was an increase in L-type Ca2+ current in control after 48 h in culture, but current density was significantly less in hypertrophy (−4.7 ± 0.8 hypertrophy vs. −10.7 ± 1.2 control pA/pF, n = 22, P < 0.05). T-type Ca2+ current density was not different. The α-adrenergic antagonist prazosin blocked the hypertrophy and the chronic effect of phenylephrine on L-type Ca2+ current. Transient outward K+ current density was decreased 70% in hypertrophy and was blocked with 4-aminopyridine. No change in Na+ current density was observed. Staurosporine, a protein kinase C inhibitor, eliminated the hypertrophy and the effect on L-type Ca2+current. These studies showed that phenylephrine-induced hypertrophy occurred via the α1-adrenergic pathway and caused electrophysiological changes and effects on ion channel expression.

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