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Effects of Sarcolemmal Background Ca²⁺ Entry and Sarcoplasmic Ca²⁺ Leak Currents on Electrophysiology and Ca²⁺ Transients in Human Ventricular Cardiomyocytes: A Computational Comparison.

ABSTRACT: The intricate regulation of the compartmental Ca²⁺ concentrations in cardiomyocytes is critical for electrophysiology, excitation-contraction coupling, and other signaling pathways. Research into the complex signaling pathways is motivated by cardiac pathologies including arrhythmia and maladaptive myocyte remodeling, which result from Ca²⁺ dysregulation. Of interest to this investigation are two types of Ca²⁺ currents in cardiomyocytes: 1) background Ca²⁺ entry, i.e., Ca²⁺ transport across the sarcolemma from the extracellular space into the cytosol, and 2) Ca²⁺ leak from the sarcoplasmic reticulum (SR) across the SR membrane into the cytosol. Candidates for the ion channels underlying background Ca²⁺ entry and SR Ca²⁺ leak channels include members of the mechano-modulated transient receptor potential (TRP) family. We used a mathematical model of a human ventricular myocyte to analyze the individual contributions of background Ca²⁺ entry and SR Ca²⁺ leak to the modulation of Ca²⁺ transients and SR Ca²⁺ load at rest and during action potentials. Background Ca²⁺ entry exhibited a positive relationship with both [Ca²⁺]_i and [Ca²⁺]_SR. Modulating SR Ca²⁺ leak had opposite effects of background Ca²⁺ entry. Effects of SR Ca²⁺ leak on Ca²⁺ were particularly pronounced at lower pacing frequency. In contrast to the pronounced effects of background and leak Ca²⁺ currents on Ca²⁺ concentrations, the effects on cellular electrophysiology were marginal. Our studies provide quantitative insights into the differential modulation of compartmental Ca²⁺ concentrations by the background and leak Ca²⁺ currents. Furthermore, our studies support the hypothesis that TRP channels play a role in strain-modulation of cardiac contractility. In summary, our investigations shed light on the physiological effects of the background and leak Ca²⁺ currents and their contribution to the development of disease caused by Ca²⁺ dysregulation.

SUBMITTER: Streiff ME

PROVIDER: S-EPMC9243544 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Ventricular arrhythmias can cause death in heart failure (HF). A trigger is the occurrence of Ca2+ waves which activate a Na+ -Ca2+ exchange (NCX) current, leading to delayed after-depolarisations and triggered action potentials. Waves arise when sarcoplasmic reticulum (SR) Ca2+ content reaches a threshold and are commonly induced experimentally by raising external Ca2+ , although the mechanism by which this causes waves is unclear and was the focus of this study. Intracellular Ca2+ was measured in voltage-clamped ventricular myocytes from both control sheep and those subjected to rapid pacing to produce HF. Threshold SR Ca2+ content was determined by applying caffeine (10 mM) following a wave and integrating wave and caffeine-induced NCX currents. Raising external Ca2+ induced waves in a greater proportion of HF cells than control. The associated increase of SR Ca2+ content was smaller in HF due to a lower threshold. Raising external Ca2+ had no effect on total influx via the L-type Ca2+ current, ICa-L , and increased efflux on NCX. Analysis of sarcolemmal fluxes revealed substantial background Ca2+ entry which sustains Ca2+ efflux during waves in the steady state. Wave frequency and background Ca2+ entry were decreased by Gd3+ or the TRPC6 inhibitor BI 749327. These agents also blocked Mn2+ entry. Inhibiting connexin hemi-channels, TRPC1/4/5, L-type channels or NCX had no effect on background entry. In conclusion, raising external Ca2+ induces waves via a background Ca2+ influx through TRPC6 channels. The greater propensity to waves in HF results from increased background entry and decreased threshold SR content. KEY POINTS: Heart failure is a pro-arrhythmic state and arrhythmias are a major cause of death. At the cellular level, Ca2+ waves resulting in delayed after-depolarisations are a key trigger of arrhythmias. Ca2+ waves arise when the sarcoplasmic reticulum (SR) becomes overloaded with Ca2+ . We investigate the mechanism by which raising external Ca2+ causes waves, and how this is modified in heart failure. We demonstrate that a novel sarcolemmal background Ca2+ influx via the TRPC6 channel is responsible for SR Ca2+ overload and Ca2+ waves. The increased propensity for Ca2+ waves in heart failure results from an increase of background influx, and a lower threshold SR content. The results of the present study highlight a novel mechanism by which Ca2+ waves may arise in heart failure, providing a basis for future work and novel therapeutic targets.

Project description:IntroductionHuman induced pluripotent stem cells (hiPSCs) harboring cardiac myosin heavy chain 6 promoter can differentiate into functional cardiomyocytes called "iCell cardiomyocytes" under blasticidin treatment condition. While iCell cardiomyocytes are expected to be used for predicting cardiotoxicity of drugs, their responses to antiarrhythmic agents remain to be elucidated. We first examined electrophysiological properties of iCell cardiomyocytes and mRNA levels of ion channels and Ca handling proteins, and then evaluated effects of class I antiarrhythmic agents on their Na+ and Ca2+ currents.MethodsiCell cardiomyocytes were cultured for 8-14 days (38-44 days after inducing their differentiation), according to the manufacturer's protocol. We determined their action potentials (APs) and sarcolemmal ionic currents using whole-cell patch clamp techniques, and also mRNA levels of ion channels and Ca handling proteins by RT-PCR. Effects of three class I antiarrhythmic agents, pirmenol, pilsicainide and mexiletine, on Na+ channel current (INa) and L-type Ca2+ channel current (ICaL) were evaluated by the whole-cell patch clamp.ResultsiCell cardiomyocytes revealed sinoatrial node-type (18%), atrial-type (18%) and ventricular-type (64%) spontaneous APs. The maximum peak amplitudes of INa, ICaL, and rapidly-activating delayed-rectifier K+ channel current were -62.7 ± 13.7, -8.1 ± 0.7, and 3.0 ± 1.0 pA/pF, respectively. The hyperpolarization-activated cation channel and inward-rectifier K+ channel currents were observed, whereas the T-type Ca2+ channel or slowly-activating delayed-rectifier K+ channel current was not detectable. mRNAs of Nav1.5, Cav1.2, Kir2.1, HCN4, KvLQT1, hERG and SERCA2 were detected, while that of HCN1, minK or MiRP was not. The class Ia antiarrhythmic agent pirmenol and class Ic agent pilsicainide blocked INa in a concentration-dependent manner with IC50 of 0.87 ± 0.37 and 0.88 ± 0.16 ?M, respectively; the class Ib agent mexiletine revealed weak INa block with a higher IC50 of 30.0 ± 3.0 ?M. Pirmenol, pilsicainide and mexiletine blocked ICaL with IC50 of 2.00 ± 0.39, 7.7 ± 2.5 and 5.0 ± 0.1 ?M, respectively.ConclusionsIn iCell cardiomyocytes, INa was blocked by the class Ia and Ic antiarrhythmic agents and ICaL was blocked by all the class I agents within the ranges of clinical concentrations, suggesting their cardiotoxicity.

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Effects of Sarcolemmal Background Ca²⁺ Entry and Sarcoplasmic Ca²⁺ Leak Currents on Electrophysiology and Ca²⁺ Transients in Human Ventricular Cardiomyocytes: A Computational Comparison.

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Effects of Sarcolemmal Background Ca2+ Entry and Sarcoplasmic Ca2+ Leak Currents on Electrophysiology and Ca2+ Transients in Human Ventricular Cardiomyocytes: A Computational Comparison.

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OmicsDI is part of the ELIXIR infrastructure

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Effects of Sarcolemmal Background Ca²⁺ Entry and Sarcoplasmic Ca²⁺ Leak Currents on Electrophysiology and Ca²⁺ Transients in Human Ventricular Cardiomyocytes: A Computational Comparison.