CaM kinase augments cardiac L-type Ca2+ current: a cellular mechanism for long Q-T arrhythmias

1999 ◽  
Vol 276 (6) ◽  
pp. H2168-H2178 ◽  
Author(s):  
Yuejin Wu ◽  
Leigh B. MacMillan ◽  
R. Blair McNeill ◽  
Roger J. Colbran ◽  
Mark E. Anderson
Keyword(s):  
Ventricular Myocytes ◽  
Ryanodine Receptors ◽  
Cam Kinase ◽  
Early Afterdepolarizations ◽  
Ultrastructural Studies ◽  
Intracellular Ca ◽  
Dependent Protein ◽  
T Tubules ◽  
Rabbit Ventricular Myocytes

Early afterdepolarizations (EAD) caused by L-type Ca2+ current ( I Ca,L) are thought to initiate long Q-T arrhythmias, but the role of intracellular Ca2+ in these arrhythmias is controversial. Rabbit ventricular myocytes were stimulated with a prolonged EAD-containing action potential-clamp waveform to investigate the role of Ca2+/calmodulin-dependent protein kinase II (CaM kinase) in I Ca,L during repolarization. I Ca,L was initially augmented, and augmentation was dependent on Ca2+ from the sarcoplasmic reticulum because the augmentation was prevented by ryanodine or thapsigargin. I Ca,Laugmentation was also dependent on CaM kinase, because it was prevented by dialysis with the inhibitor peptide AC3-I and reconstituted by exogenous constitutively active CaM kinase when Ba2+ was substituted for bath Ca2+. Ultrastructural studies confirmed that endogenous CaM kinase, L-type Ca2+ channels, and ryanodine receptors colocalized near T tubules. EAD induction was significantly reduced in current-clamped cells dialyzed with AC3-I (4/15) compared with cells dialyzed with an inactive control peptide (11/15, P = 0.013). These findings support the hypothesis that EADs are facilitated by CaM kinase.

scholarly journals Revisiting the ionic mechanisms of early afterdepolarizations in cardiomyocytes: predominant by Ca waves or Ca currents?

2012 ◽  
Vol 302 (8) ◽  
pp. H1636-H1644 ◽  
Author(s):  
Zhenghang Zhao ◽  
Hairuo Wen ◽  
Nadezhda Fefelova ◽  
Charelle Allen ◽  
Akemichi Baba ◽  
...  
Keyword(s):  
Ventricular Myocytes ◽  
Ryanodine Receptors ◽  
Convincing Evidence ◽  
Rate Dependency ◽  
Early Afterdepolarizations ◽  
Relative Contribution ◽  
Intracellular Ca ◽  
Ionic Mechanisms ◽  
Rabbit Ventricular Myocytes ◽  
Bayk 8644

Early afterdepolarizations (EADs) have been implicated in severe cardiac arrhythmias and sudden cardiac deaths. However, the mechanism(s) for EAD genesis, especially regarding the relative contribution of Ca2+ wave (CaW) vs. L-type Ca current ( ICa,L), still remains controversial. In the present study, we simultaneously recorded action potentials (APs) and intracellular Ca2+ images in isolated rabbit ventricular myocytes and systematically compared the properties of EADs in the following two pharmacological models: 1) hydrogen peroxide (H2O2; 200 μM); and 2) isoproterenol (100 nM) and BayK 8644 (50 nM) (Iso + BayK). We assessed the rate dependency of EADs, the temporal relationship between EADs and corresponding CaWs, the distribution of EADs over voltage, and the effects of blockers of ICa,L, Na/Ca exchangers, and ryanodine receptors. The most convincing evidence came from the AP-clamp experiment, in which the cell membrane clamp was switched from current clamp to voltage clamp using a normal AP waveform without EAD; CaWs disappeared in the H2O2 model, but persisted in the Iso + BayK model. We postulate that, although CaWs and reactivation of ICa,L may act synergistically in either case, reactivation of ICa,L plays a predominant role in EAD genesis under oxidative stress (H2O2 model), while spontaneous CaWs are a predominant cause for EADs under Ca2+ overload condition (Iso + BayK model).

The functional role of cardiac T-tubules explored in a model of rat ventricular myocytes

2006 ◽  
Vol 364 (1842) ◽  
pp. 1187-1206 ◽  
Author(s):  
Michal Pásek ◽  
Jiři Šimurda ◽  
Georges Christé
Keyword(s):  
Ventricular Myocytes ◽  
Contractile Activity ◽  
Tubular Lumen ◽  
Current Clamp Mode ◽  
Intracellular Ca ◽  
Ionic Changes ◽  
Access Resistance ◽  
T Tubules ◽  
Tubular Membranes

The morphology of the cardiac transverse-axial tubular system (TATS) has been known for decades, but its function has received little attention. To explore the possible role of this system in the physiological modulation of electrical and contractile activity, we have developed a mathematical model of rat ventricular cardiomyocytes in which the TATS is described as a single compartment. The geometrical characteristics of the TATS, the biophysical characteristics of ion transporters and their distribution between surface and tubular membranes were based on available experimental data. Biophysically realistic values of mean access resistance to the tubular lumen and time constants for ion exchange with the bulk extracellular solution were included. The fraction of membrane in the TATS was set to 56%. The action potentials initiated in current-clamp mode are accompanied by transient K + accumulation and transient Ca 2+ depletion in the TATS lumen. The amplitude of these changes relative to external ion concentrations was studied at steady-state stimulation frequencies of 1–5 Hz. Ca 2+ depletion increased from 7 to 13.1% with stimulation frequency, while K + accumulation decreased from 4.1 to 2.7%. These ionic changes (particularly Ca 2+ depletion) implicated significant decrease of intracellular Ca 2+ load at frequencies natural for rat heart.

Intracellular Ca2+ regulates responsiveness of cardiac L-type Ca2+ current to protein kinase A: role of calmodulin

2004 ◽  
Vol 286 (1) ◽  
pp. H186-H194 ◽  
Author(s):  
Kenneth B. Walsh ◽  
Qi Cheng
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Ventricular Myocytes ◽  
External Solution ◽  
Inhibitory Peptide ◽  
Dependent Inhibition ◽  
Intracellular Ca ◽  
Dependent Protein ◽  
Kinase A

The goal of this study was to determine whether the protein kinase A (PKA) responsiveness of the cardiac L-type Ca2+ current ( ICa) is affected during transient increases in intracellular Ca2+ concentration. Ventricular myocytes were isolated from 3- to 4-day-old neonatal rats and cultured on aligned collagen thin gels. When measured in 1 or 2 mM Ca2+ external solution, the aligned myocytes displayed a large ICa that was weakly regulated (20% increase) during stimulation of PKA by 2 μM forskolin. In contrast, application of forskolin caused a 100% increase in ICa when the external Ca2+ concentration was reduced to 0.5 mM or replaced with Ba2+. This Ca2+-dependent inhibition was also observed when the cells were treated with 1 μM isoproterenol, 100 μM 3-isobutyl-1-methylxanthine, or 500 μM 8-bromo-cAMP. The responsiveness of ICa to PKA was restored during intracellular dialysis with a calmodulin (CaM) inhibitory peptide but not during treatment with inhibitors of protein kinase C, Ca2+/CaM-dependent protein kinase, or calcineurin. Adenoviral-mediated expression of a CaM molecule with mutations in all four Ca2+-binding sites also increased the PKA sensitivity of ICa. Finally, adult mouse ventricular myocytes displayed a greater response to forskolin and cAMP in external Ba2+. Thus Ca2+ entering the myocyte through the voltage-gated Ca2+ channel regulates the PKA responsiveness of ICa.

scholarly journals Sarcolemmal distribution of ICa and INCX and Ca2+ autoregulation in mouse ventricular myocytes

2017 ◽  
Vol 313 (1) ◽  
pp. H190-H199 ◽  
Author(s):  
Hanne C. Gadeberg ◽  
Cherrie H. T. Kong ◽  
Simon M. Bryant ◽  
Andrew F. James ◽  
Clive H. Orchard
Keyword(s):  
Steady State ◽  
Sarcoplasmic Reticulum ◽  
Cardiac Myocytes ◽  
Ventricular Myocytes ◽  
Surface Membrane ◽  
Ryanodine Receptors ◽  
Cell Voltage ◽  
Intact Cells ◽  
T Tubules

The balance of Ca2+ influx and efflux regulates the Ca2+ load of cardiac myocytes, a process known as autoregulation. Previous work has shown that Ca2+ influx, via L-type Ca2+ current ( ICa), and efflux, via the Na+/Ca2+ exchanger (NCX), occur predominantly at t-tubules; however, the role of t-tubules in autoregulation is unknown. Therefore, we investigated the sarcolemmal distribution of ICa and NCX current ( INCX), and autoregulation, in mouse ventricular myocytes using whole cell voltage-clamp and simultaneous Ca2+ measurements in intact and detubulated (DT) cells. In contrast to the rat, INCX was located predominantly at the surface membrane, and the hysteresis between INCX and Ca2+ observed in intact myocytes was preserved after detubulation. Immunostaining showed both NCX and ryanodine receptors (RyRs) at the t-tubules and surface membrane, consistent with colocalization of NCX and RyRs at both sites. Unlike INCX, ICa was found predominantly in the t-tubules. Recovery of the Ca2+ transient amplitude to steady state (autoregulation) after application of 200 µM or 10 mM caffeine was slower in DT cells than in intact cells. However, during application of 200 µM caffeine to increase sarcoplasmic reticulum (SR) Ca2+ release, DT and intact cells recovered at the same rate. It appears likely that this asymmetric response to changes in SR Ca2+ release is a consequence of the distribution of ICa, which is reduced in DT cells and is required to refill the SR after depletion, and NCX, which is little affected by detubulation, remaining available to remove Ca2+ when SR Ca2+ release is increased. NEW & NOTEWORTHY This study shows that in contrast to the rat, mouse ventricular Na+/Ca2+ exchange current density is lower in the t-tubules than in the surface sarcolemma and Ca2+ current is predominantly located in the t-tubules. As a consequence, the t-tubules play a role in recovery (autoregulation) from reduced, but not increased, sarcoplasmic reticulum Ca2+ release.

scholarly journals Stretch-activated channel activation promotes early afterdepolarizations in rat ventricular myocytes under oxidative stress

2009 ◽  
Vol 296 (5) ◽  
pp. H1227-H1235 ◽  
Author(s):  
Yanggan Wang ◽  
Ronald W. Joyner ◽  
Mary B. Wagner ◽  
Jun Cheng ◽  
Dongwu Lai ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Ventricular Myocytes ◽  
Ionic Current ◽  
Reversal Potential ◽  
Mechanical Stretch ◽  
Early Afterdepolarizations ◽  
Rat Ventricular Myocytes ◽  
Intracellular Ca ◽  
And Oxidative Stress

Mechanical stretch and oxidative stress have been shown to prolong action potential duration (APD) and produce early afterdepolarizations (EADs). Here, we developed a simulation model to study the role of stretch-activated channel (SAC) currents in triggering EADs in ventricular myocytes under oxidative stress. We adapted our coupling clamp circuit so that a model ionic current representing the actual SAC current was injected into ventricular myocytes and added as a real-time current. This current was calculated as ISAC = GSAC * ( Vm − ESAC), where GSAC is the stretch-activated conductance, Vm is the membrane potential, and ESAC is the reversal potential. In rat ventricular myocytes, application of GSAC did not produce sustained automaticity or EADs, although turn-on of GSAC did produce some transient automaticity at high levels of GSAC. Exposure of myocytes to 100 μM H2O2 induced significant APD prolongation and increase in intracellular Ca2+ load and transient, but no EAD or sustained automaticity was generated in the absence of GSAC. However, the combination of GSAC and H2O2 consistently produced EADs at lower levels of GSAC (2.6 ± 0.4 nS, n = 14, P < 0.05). Pacing myocytes at a faster rate further prolonged APD and promoted the development of EADs. SAC activation plays an important role in facilitating the development of EADs in ventricular myocytes under acute oxidative stress. This mechanism may contribute to the increased propensity to lethal ventricular arrhythmias seen in cardiomyopathies, where the myocardium stretch and oxidative stress generally coexist.

scholarly journals Dual role of junctin in the regulation of ryanodine receptors and calcium release in cardiac ventricular myocytes

2011 ◽  
Vol 589 (24) ◽  
pp. 6063-6080 ◽  
Author(s):  
Beth A. Altschafl ◽  
Demetrios A. Arvanitis ◽  
Oscar Fuentes ◽  
Qunying Yuan ◽  
Evangelia G. Kranias ◽  
...  
Keyword(s):  
Calcium Release ◽  
Ventricular Myocytes ◽  
Ryanodine Receptors ◽  
Dual Role ◽  
Cardiac Ventricular Myocytes

Abstract 3503: Hydrogen Peroxide-Induced Afterdepolarizations are Dependent on Calmodulin Kinase II Signaling

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Lai-Hua Xie ◽  
Fuhua Chen ◽  
James N Weiss
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Ventricular Myocytes ◽  
Heart Association ◽  
Early Afterdepolarizations ◽  
Calmodulin Kinase ◽  
Repolarization Reserve ◽  
Delayed Afterdepolarizations ◽  
Rabbit Ventricular Myocytes ◽  
Camkii Activation

Background: In the heart, hydrogen peroxide (H 2 O 2 ) has been shown to cause early afterdepolarizations (EADs) and triggered activity by impairing Na current (I Na ) inactivation. Since H 2 O 2 has been recently shown to activate Ca 2+ /calmodulin kinase II (CaMKII), and since CaMKII activation has also been reported to impair I Na inactivation and predispose to EADs, we hypothesized that CaMKII activation by H 2 O 2 may be an important factor in the genesis of EADs induced by oxidative stress. Methods and Results: Patch-clamped Fluo-4 AM-loaded rabbit ventricular myocytes were exposed to H 2 O 2 (0.1–1mM), which induced spontaneous EADs after 5–15 min. Both the I Na blocker tetrodoxtin (TTX, 10 μM) and the I Ca,L blocker nifedipine shortened AP duration (APD) and suppressed EADs. H 2 O 2 increased both peak and steady-state I Ca,L under square-pulse voltage clamp, and enhanced I Ca,L to a greater extent during the AP plateau than during the AP upstroke under AP clamp conditions. In addition, by prolonging the AP plateau and increasing Ca influx via maintained I Ca,L , H 2 O 2 -induced EADs frequently caused DADs delayed afterdepolarizations (DADs) due to spontaneous SR Ca release waves after repolarization. KN-93(1 μM), a CaMKII inhibitor, prevented H 2 O 2 -induced EADs (n=4), whereas the inactive analogue KN-92 did not (n=5). Conclusion: These findings indicate that H 2 O 2 -induced EADs depend on both impaired I Na inactivation to reduce repolarization reserve and enhanced I Ca,L to reverse repolarization. Intact CaMKII signaling is necessary for EAD generation in this setting, presumably via its actions on I Na and I Ca,L , although direct redox effects on other ion channels/transporters may also be important. Our observations support a link between increased oxidative stress, CaMKII activation and afterdepolarizations as triggers of lethal ventricular arrhythmias in diseased heart. This research has received full or partial funding support from the American Heart Association, AHA National Center.

Abstract 18111: Flecainide Exerts its Antiarrhythmic Action in CPVT Through Blockade of Neuronal Na+ channel-mediated Arrhythmogenic Diastolic Ca2+ Release

Circulation ◽  
2015 ◽  
Vol 132 (suppl_3) ◽  
Author(s):  
Przemyslaw Radwanski ◽  
Hsiang-Ting Ho ◽  
Björn Knollmann ◽  
Andriy Belevych ◽  
Sándor Györke
Keyword(s):  
Ventricular Myocytes ◽  
Ryanodine Receptors ◽  
Cellular Level ◽  
Antiarrhythmic Effect ◽  
Na Channel ◽  
Antiarrhythmic Action ◽  
Pharmacological Interventions ◽  
Cellular Excitability

Background: Flecaininde is an effective antiarrhythmic in management of CPVT. Its antiarrhythmic action has been attributed to direct effect on RyR2 and reduced cellular excitability through the inhibition of cardiac-type Na + channels. Recently we demonstrated that neuronal Na + channels (nNa v s) colocalize with the ryanodine receptors (RyR2) Ca 2+ release channels on the sarcoplasmic reticulum. Here we explore a novel mechanism that may contribute to the antiarrhythmic effect of flecainide, mainly uncoupling of aberrant Na + /Ca 2+ signaling through nNa v inhibition. Methods: To study the effects of flecainide on Ca 2+ signaling we used a murine model of cardiac calsequestrin-associated CPVT. We performed confocal microscopy in intact isolated ventricular myocytes to assess Ca 2+ handling and recorded late Na + current (I Na ) during various pharmacological interventions. Surface electrocardiograms were performed during catecholamine challenge to monitor arrhythmic activity in vivo . Results: During catecholamine stimulation with isoproterenol (Iso; 100 nM) disruption of the cross-talk between nNa v s and RyR2 by nNa v blockade with 100nM tetrodotoxin (TTX) and riluzole (10μM) as well as flecainide (2.5μM) reduced Iso-promoted late I Na and DCR in isolated intact CPVT cardiomyocytes. To further examine the role of nNa v -mediated late I Na in genesis of DCR we augmented nNa v channel activity with β-Pompilidotoxin (β-PMTX, 40μM). Effects of β-PMTX in CPVT cardiomyocytes were reversed by nNa v blockade with TTX and riluzole as well as flecainide. This reduction in late I Na and DCR frequency with riluzole and flecainide in the presence of β-PMTX on cellular level translated to decreased ventricular arrhythmias in CPVT mice. Conclusion: These data suggest that disruption of nNa v -mediated late I Na can prevent arrhythmogenic DCR in CPVT. Importantly, the antiarrhythmic effects of flecainide can be attributed, at least in part, to its nNa v blocking properties.

Distinct Intracellular Calcium Profiles Following Influx Through N- Versus L-Type Calcium Channels: Role of Ca2+-Induced Ca2+ Release

2004 ◽  
Vol 92 (1) ◽  
pp. 135-143 ◽  
Author(s):  
Keith Tully ◽  
Steven N. Treistman
Keyword(s):  
Intracellular Calcium ◽  
Ryanodine Receptors ◽  
Decay Rates ◽  
Selective Activation ◽  
Intracellular Ca ◽  
Pheochromocytoma Pc12 ◽  
High Concentrations ◽  
Neuronal Functions ◽  
Temporal Profiles

Selective activation of neuronal functions by Ca2+ is determined by the kinetic profile of the intracellular calcium ([Ca2+]i) signal in addition to its amplitude. Concurrent electrophysiology and ratiometric calcium imaging were used to measure transmembrane Ca2+ current and the resulting rise and decay of [Ca2+]i in differentiated pheochromocytoma (PC12) cells. We show that equal amounts of Ca2+ entering through N-type and L-type voltage-gated Ca2+ channels result in significantly different [Ca2+]i temporal profiles. When the contribution of N-type channels was reduced by ω-conotoxin MVIIA treatment, a faster [Ca2+]i decay was observed. Conversely, when the contribution of L-type channels was reduced by nifedipine treatment, [Ca2+]i decay was slower. Potentiating L-type current with BayK8644, or inactivating N-type channels by shifting the holding potential to −40 mV, both resulted in a more rapid decay of [Ca2+]i. Channel-specific differences in [Ca2+]i decay rates were abolished by depleting intracellular Ca2+ stores with thapsigargin or by blocking ryanodine receptors with ryanodine, suggesting the involvement of Ca2+-induced Ca2+ release (CICR). Further support for involvement of CICR is provided by the demonstration that caffeine slowed [Ca2+]i decay while ryanodine at high concentrations increased the rate of [Ca2+]i decay. We conclude that Ca2+ entering through N-type channels is amplified by ryanodine receptor mediated CICR. Channel-specific activation of CICR provides a mechanism whereby the kinetics of intracellular Ca2+ leaves a fingerprint of the route of entry, potentially encoding the selective activation of a subset of Ca2+-sensitive processes within the neuron.

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