Nav1.5-dependent persistent Na+ influx activates CaMKII in rat ventricular myocytes and N1325S mice

2011 ◽  
Vol 301 (3) ◽  
pp. C577-C586 ◽  
Author(s):  
Lina Yao ◽  
Peidong Fan ◽  
Zhan Jiang ◽  
Serge Viatchenko-Karpinski ◽  
Yuzhi Wu ◽  
...  
Keyword(s):  
Ventricular Myocytes ◽  
Neonatal Rat ◽  
Polymorphic Ventricular Tachycardia ◽  
Inhibitory Peptide ◽  
Rat Ventricular Myocytes ◽  
Protein Protein Interaction ◽  
Intracellular Ca ◽  
Dependent Protein ◽  
Neonatal Rat Ventricular Myocytes ◽  
Camkii Activation

Late Na+ current ( INaL) and Ca2+/calmodulin-dependent protein kinase II (CaMKII) are both increased in the diseased heart. Recently, CaMKII was found to phosphorylate the Na+ channel 1.5 (Nav1.5), resulting in enhanced INaL. Conversely, an increase of INaL would be expected to cause elevation of intracellular Ca2+ and activation of CaMKII. However, a relationship between enhancement of INaL and activation of CaMKII has yet to be demonstrated. We investigated whether Na+ influx via Nav1.5 leads to CaMKII activation and explored the functional significance of this pathway. In neonatal rat ventricular myocytes (NRVM), treatment with the INaL activators anemone toxin II (ATX-II) or veratridine increased CaMKII autophosphorylation and increased phosphorylation of CaMKII substrates phospholamban and ryanodine receptor 2. Knockdown of Nav1.5 (but not Nav1.1 or Nav1.2) prevented ATX-II-induced CaMKII phosphorylation, providing evidence for a specific role of Nav1.5 in CaMKII activation. In support of this view, CaMKII activity was also increased in hearts of transgenic mice overexpressing a gain-of-function Nav1.5 mutant (N1325S). The effects of both ATX-II and the N1325S mutation were reversed by either INaL inhibition (with ranolazine or tetrodotoxin) or CaMKII inhibition (with KN93 or autocamtide 2-related inhibitory peptide). Furthermore, ATX-II treatment also induced CaMKII-Nav1.5 coimmunoprecipitation. The same association between CaMKII and Nav1.5 was also found in N1325S mice, suggesting a direct protein-protein interaction. Pharmacological inhibitions of either CaMKII or INaL also prevented ATX-II-induced cell death in NRVM and reduced the incidence of polymorphic ventricular tachycardia induced by ATX-II in rat perfused hearts. Taken together, these results suggest that a Nav1.5-dependent increase in Na+ influx leads to activation of CaMKII, which in turn phosphorylates Nav1.5, further promoting Na+ influx. Pharmacological inhibition of either CaMKII or Nav1.5 can ameliorate cardiac dysfunction caused by excessive Na+ influx.

CARP Plays a Key Role in Cellular Growth Under Hypertrophic Stimuli in Neonatal Rat Ventricular Myocytes

2013 ◽  
Vol 9 ◽  
Author(s):  
Tara A Shrout
Keyword(s):  
Gene Expression ◽  
Ventricular Myocytes ◽  
Cellular Growth ◽  
Neonatal Rat ◽  
Transcriptional Level ◽  
Dual Nature ◽  
Hypertrophic Growth ◽  
Rat Ventricular Myocytes ◽  
Neonatal Rat Ventricular Myocytes ◽  
Over Time

Cardiac hypertrophy is a growth process that occurs in response to stress stimuli or injury, and leads to the induction of several pathways to alter gene expression. Under hypertrophic stimuli, sarcomeric structure is disrupted, both as a consequence of gene expression and local changes in sarcomeric proteins. Cardiac-restricted ankyrin repeat protein (CARP) is one such protein that function both in cardiac sarcomeres and at the transcriptional level. We postulate that due to this dual nature, CARP plays a key role in maintaining the cardiac sarcomere. GATA4 is another protein detected in cardiomyocytes as important in hypertrophy, as it is activated by hypertrophic stimuli, and directly binds to DNA to alter gene expression. Results of GATA4 activation over time were inconclusive; however, the role of CARP in mediating hypertrophic growth in cardiomyocytes was clearly demonstrated. In this study, Neonatal Rat Ventricular Myocytes were used as a model to detect changes over time in CARP and GATA4 under hypertrophic stimulation by phenylephrine and high serum media. Results were detected by analysis of immunoblotting. The specific role that CARP plays in mediating cellular growth under hypertrophic stimuli was studied through immunofluorescence, which demonstrated that cardiomyocyte growth with hypertrophic stimulation was significantly blunted when NRVMs were co-treated with CARP siRNA. These data suggest that CARP plays an important role in the hypertrophic response in cardiomyocytes.

Abstract 15280: Comparisons of Action Potentials and Ionic Currents Between Neonatal Rat Ventricular Myocytes and Adult Zebrafish Ventricular Myocytes

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Adonis Z Wu ◽  
Shien-Fong Lin ◽  
Sheng-Nan Wu
Keyword(s):  
Electrical Properties ◽  
Ventricular Myocytes ◽  
Ionic Currents ◽  
Firing Frequency ◽  
Neonatal Rat ◽  
Adult Zebrafish ◽  
Patch Clamp Technique ◽  
Action Current ◽  
Rat Ventricular Myocytes ◽  
Neonatal Rat Ventricular Myocytes

Introduction: Zebrafish heart is established as a model to investigate cardiac electrical abnormalities. However, electrical properties of adult zebrafish cardiomyocytes are not sufficiently characterized. Hypothesis: In this study, by comparing the electrical properties between neonatal rat ventricular myocytes (NRVMs) and adult zebrafish ventricular myocytes (AZVMs), we intended to characterize the action potential (AP), action current (AC) and the properties of Na + current ( I Na ) in AZVMs. Methods: We used patch-clamp technique to characterize the electrical properties, including AP, AC and I Na , in cultured NRVMs and freshly isolated AZVMs. Results: NVRMs showed larger AP amplitude (119±6 vs. 79±4mV, p<.05) but shorter AP duration (APD 90 , 136±11 vs. 213±19 ms, p<.05) than those of AZVMs. The AP duration exhibited marked frequency-dependent alterations in AZVMs. Under the slow pacing rate, early after-depolarizations (EAD) emerged under slow pacing rate with 0.05 Hz. In cell-attached voltage-clamp recordings made from AZVMs, ACs could be elicited by +10 mV steps. As the depolarization step increased to +70 mV, the latency for appearance of ACs was progressively reduced from >123 ms to 9.8 ms. The presence of spontaneous ACs was monitored in spontaneously beating NRVMs and AZVMs. The AC amplitude in NRVMs was larger compared to that in AZVMs (17.3±2.1 vs. 11.6±1.1 pA, p<.05), although firing frequency of AC in NRVMs is higher than in AZVMs (1.13±0.09 vs. 0.38±0.03 Hz, p<.05). The lowering effect of ranolazine, a I Na antagonist, on firing frequency was significantly larger in NRVMs (1.13±0.09 to 0.31±0.02 Hz, p<.05) than in AZVMs (0.38±0.03 to 0.27±0.02 Hz). There was a hyperpolarizing shift of peak I Na in AZVM compared to NRVM. Conclusions: Our results demonstrated major differences in the cellular electrical behavior between AZVMs and NRVMs.

scholarly journals Chronotropic Response of Cultured Neonatal Rat Ventricular Myocytes to Short-Term Fluid Shear

2006 ◽  
Vol 46 (2) ◽  
pp. 113-122 ◽  
Author(s):  
Ilka Lorenzen-Schmidt ◽  
Geert W. Schmid-Schönbein ◽  
Wayne R. Giles ◽  
Andrew D. McCulloch ◽  
Shu Chien ◽  
...  
Keyword(s):  
Ventricular Myocytes ◽  
Neonatal Rat ◽  
Fluid Shear ◽  
Short Term ◽  
Chronotropic Response ◽  
Rat Ventricular Myocytes ◽  
Neonatal Rat Ventricular Myocytes

scholarly journals Hexokinase–mitochondrial interactions regulate glucose metabolism differentially in adult and neonatal cardiac myocytes

2013 ◽  
Vol 142 (4) ◽  
pp. 425-436 ◽  
Author(s):  
Guillaume Calmettes ◽  
Scott A. John ◽  
James N. Weiss ◽  
Bernard Ribalet
Keyword(s):  
Cardiac Tissue ◽  
Ventricular Myocytes ◽  
Glycogen Synthesis ◽  
Neonatal Rat ◽  
Metabolic Profiles ◽  
Rat Ventricular Myocytes ◽  
Glycolytic Activity ◽  
Extracellular Glucose ◽  
Functional Consequences ◽  
Neonatal Rat Ventricular Myocytes

In mammalian tumor cell lines, localization of hexokinase (HK) isoforms to the cytoplasm or mitochondria has been shown to control their anabolic (glycogen synthesis) and catabolic (glycolysis) activities. In this study, we examined whether HK isoform differences could explain the markedly different metabolic profiles between normal adult and neonatal cardiac tissue. We used a set of novel genetically encoded optical imaging tools to track, in real-time in isolated adult (ARVM) and neonatal (NRVM) rat ventricular myocytes, the subcellular distributions of HKI and HKII, and the functional consequences on glucose utilization. We show that HKII, the predominant isoform in ARVM, dynamically translocates from mitochondria and cytoplasm in response to removal of extracellular glucose or addition of iodoacetate (IAA). In contrast, HKI, the predominant isoform in NRVM, is only bound to mitochondria and is not displaced by the above interventions. In ARVM, overexpression of HKI, but not HKII, increased glycolytic activity. In neonatal rat ventricular myocytes (NVRM), knockdown of HKI, but not HKII, decreased glycolytic activity. In conclusion, differential interactions of HKI and HKII with mitochondria underlie the different metabolic profiles of ARVM and NRVM, accounting for the markedly increased glycolytic activity of NRVM.

Abstract 17019: Two Distinct mechanisms by which Na+/Ca2+ dysregulation contributes to Arrhythmogenic Diastolic Ca2+ Release

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Przemyslaw Radwanski ◽  
Rengasayee Veeraraghavan ◽  
Björn Knollmann ◽  
Sándor Györke
Keyword(s):  
Ventricular Myocytes ◽  
Cellular Level ◽  
Polymorphic Ventricular Tachycardia ◽  
Cell Periphery ◽  
Individualized Approach ◽  
Triggered Arrhythmias ◽  
Local Contribution ◽  
Dependent Protein ◽  
Camkii Activation

Na + and Ca 2+ imbalance is associated with triggered arrhythmias resulting from diastolic Ca 2+ release (DCR) from sarcoplasmic reticulum (SR). Recent evidence suggests Na + channel blockade to be a promising therapy for pathologies, including catecholaminergic polymorphic ventricular tachycardia (CPVT). However, the specific mechanism(s) as to how Na + /Ca 2+ dysregulation contribute to arrhythmias is unknown. Confocal microscopy of ventricular myocytes isolated from CPVT mice lacking the cardiac calsequestrin was used to assess Ca 2+ handling response to isoproterenol (Iso) and various pharmacological interventions, while electrocardiograms were acquired during catecholamine challenge to assess the roles of various pools of Na + channels in CPVT. We identify two pools of Na + channels: one composed of cardiac-type Na + channels localized to cell periphery, and a ‘ local pool ’ comprised of neuronal Na + channels colocalizing with RyR2 in the T-tubules. Augmenting function of both Na + channel pools with ATX-II in the presence Iso resulted in SR Ca 2+ overload and activation of Ca 2+ /calmodulin-dependent protein kinase II (CaMKII), which precipitated DCR. These, in turn, translated into frequent arrhythmias in CPVT mice. Selectively augmenting function of ‘local pool’ neuronal Na + channels with β-Pompilidotoxin (β-PMTX) precipitated DCR on the cellular level causing frequent arrhythmias during catecholamine challenge in vivo . However, increasing local Na + fluxes reduced SR Ca 2+ load suggesting that local elevation in cytosolic Ca 2+ rather than global SR Ca 2+ overload underlies DCR and arrhythmias under such conditions. These data suggest two distinct mechanisms for Na + /Ca 2+ dysregulation-mediated arrhythmias. The first relies on SR Ca 2+ overload and CaMKII activation and the other on local contribution of Na + -Ca 2+ exchange to DCR. Consideration of these divergent mechanisms may enhance individualized approach to arrhythmia management.

scholarly journals Nitric oxide-induced cardioprotection in cultured rat ventricular myocytes

2000 ◽  
Vol 278 (4) ◽  
pp. H1211-H1217 ◽  
Author(s):  
Roby D. Rakhit ◽  
Richard J. Edwards ◽  
James W. Mockridge ◽  
Anwar R. Baydoun ◽  
Amanda W. Wyatt ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Ventricular Myocytes ◽  
Culture Conditions ◽  
Neonatal Rat ◽  
No Donor ◽  
Rat Ventricular Myocytes ◽  
Kinase C ◽  
Neonatal Rat Ventricular Myocytes ◽  
Oxide Synthase

The aim of this study was to investigate the role of nitric oxide (NO) in a cellular model of early preconditioning (PC) in cultured neonatal rat ventricular myocytes. Cardiomyocytes “preconditioned” with 90 min of stimulated ischemia (SI) followed by 30 min reoxygenation in normal culture conditions were protected against subsequent 6 h of SI. PC was blocked by N G-monomethyl-l-arginine monoacetate but not by dexamethasone pretreatment. Inducible nitric oxide synthase (NOS) protein expression was not detected during PC ischemia. Pretreatment (90 min) with the NO donor S-nitroso- N-acetyl-l,l-penicillamine (SNAP) mimicked PC, resulting in significant protection. SNAP-triggered protection was completely abolished by 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) but was unaffected by chelerythrine or the presence of glibenclamide and 5-hydroxydecanoate. With the use of RIA, SNAP treatment increased cGMP levels, which were blocked by ODQ. Hence, NO is implicated as a trigger in this model of early PC via activation of a constitutive NOS isoform. After exposure to SNAP, the mechanism of cardioprotection is cGMP dependent but independent of protein kinase C or ATP-sensitive K+ channels. This differs from the proposed mechanism of NO-induced cardioprotection in late PC.

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