Action Potentials In Cardiac Cells Research Articles

Uncertainty assessment of proarrhythmia predictions derived from multi-level in silico models

In silico methods can be used for an early assessment of arrhythmogenic properties of drug candidates. However, their use for decision-making is conditioned by the possibility to estimate the predictions’ uncertainty. This work describes our efforts to develop uncertainty quantification methods for the predictions produced by multi-level proarrhythmia models. In silico models used in this field usually start with experimental or predicted IC50 values that describe drug-induced ion channel blockade. Using such inputs, an electrophysiological model computes how the ion channel inhibition, exerted by a drug in a certain concentration, translates to an altered shape and duration of the action potential in cardiac cells, which can be represented as arrhythmogenic risk biomarkers such as the APD90. Using this framework, we identify the main sources of aleatory and epistemic uncertainties and propose a method based on probabilistic simulations that replaces single-point estimates predicted using multiple input values, including the IC50s and the electrophysiological parameters, by distributions of values. Two selected variability types associated with these inputs are then propagated through the multi-level model to estimate their impact on the uncertainty levels in the output, expressed by means of intervals. The proposed approach yields single predictions of arrhythmogenic risk biomarkers together with value intervals, providing a more comprehensive and realistic description of drug effects on a human population. The methodology was tested by predicting arrhythmogenic biomarkers on a series of twelve well-characterised marketed drugs, belonging to different arrhythmogenic risk classes.

Isoform changes of action potential regulators in the ventricles of arrhythmogenic phospholamban-R14del humanized mouse hearts

Arrhythmogenic cardiomyopathy (ACM) is characterized by life-threatening ventricular arrhythmias and sudden cardiac death and affects hundreds of thousands of patients worldwide. The deletion of Arginine 14 (p.R14del) in the phospholamban (PLN) gene has been implicated in the pathogenesis of ACM. PLN is a key regulator of sarcoplasmic reticulum (SR) Ca2+ cycling and cardiac contractility. Despite global gene and protein expression studies, the molecular mechanisms of PLN-R14del ACM pathogenesis remain unclear. Using a humanized PLN-R14del mouse model and human induced pluripotent stem cell derived cardiomyocytes (iPSC-CMs), we investigated the transcriptome-wide mRNA splicing changes associated with the R14del mutation. We identified >200 significant alternative splicing (AS) events and distinct AS profiles were observed in the right (RV) and left (LV) ventricles in PLN-R14del compared to WT mouse hearts. Enrichment analysis of the AS events showed that the most affected biological process was associated with “cardiac cell action potential”, specifically in the RV. We found that splicing of 2 key genes, Trpm4 and Camk2d, which encode proteins regulating calcium homeostasis in the heart, were altered in PLN-R14del mouse hearts and human iPSC-CMs. Bioinformatical analysis pointed to the tissue-specific splicing factors Srrm4 and Nova1 as likely upstream regulators of the observed splicing changes in the PLN-R14del cardiomyocytes. Our findings suggest that aberrant splicing may affect Ca2+-homeostasis in the heart, contributing to the increased risk of arrythmogenesis in PLN-R14del ACM.

Hydrocotyle bonariensis Comm ex Lamm (Araliaceae) leaves extract inhibits IKs not IKr potassium currents: Potential implications for anti-arrhythmic therapy

Background and aimHydrocotyle bonariensis Comm ex Lamm (Araliaceae) is one of these plants sufficiently exploited in traditional African medicine for its hypotensive effect. However, the pharmacological effects of those plants on cardiac functions are not well known. The potassium currents IKs and IKr, responsible for the repolarization of cardiac cell action potential, strongly influence the human cardiac rhythm. Therefore, modulators of these currents have a beneficial or undesirable medical importance in relation to cardiac arrhythmias. In order to optimize the therapeutic use of this medicinal plant, we studied the effects of hydro-ethanolic leaf extract of Hydrocotyle bonariensis on both potassium currents. Experimental procedureThe patch clamp experiments for IK currents recording were performed on the HEK 293 (Human Embryonic Kidney 293) cell line, stably transfected with either KCNQ1 and KCNE1 genes encoding the channel responsible for the "IKs" current (HEK293 IKs), or with hERG (human ether-a-go-go related gene) gene encoding "IKr" current (HEK293 IKr). Results and conclusionThis study revealed that the hydro-ethanolic leaf extract of H. bonariensis significantly inhibits the slow potassium component (IKs) without altering the fast potassium component (IKr). The extract at 0.5 mg/ml decreases IKs conductance by 24 ± 4.1% (n = 6) without modifying its activation threshold suggesting a direct blockade of the slow potassium channel. This selective action of the extract on the IKs current reflects a class III anti-arrhythmic effect.

A Graphical-Based Mathematical Model for Simulating Excitability in a Single Cardiac Cell

Excitability is a phenomenon seen in different kinds of systems, e.g., biological systems. Cardiac cells and neurons are well-known examples of excitable biological systems. Excitability as a crucial property should be involved in mathematical models of cardiac cells, along with the other biological properties. Excitability of mathematical cardiac-cell models is usually investigated in the phase plane (or the phase space) which is not applicable with simple mathematical analysis. Besides, the possible roles of each model parameter in the excitability property cannot be investigated explicitly and independently using phase plane analysis. In this paper, we present a new graphical-based method for designing excitability of a single cardiac cell. Each parameter in the presented approach not only has electrophysiological interpretation but also its role in regulating excitability is evident and can be analysed explicitly. Our approach is simpler and more tractable by mathematical analysis than the phase plane method. Another advantage of our approach is that the other important feature of the cardiac cell action potential, i.e., plateau morphology, can be designed and regulated separately from the excitability property. To evaluate our presented approach, we applied it for simulating excitability in well-known complex electrophysiological models of ventricular and atrial cells. Results show that our model can simulate excitability and time evolution of the plateau phase simultaneously.

When the clock strikes: Modeling the relation between circadian rhythms and cardiac arrhythmias

It has recently been observed that the occurrence of sudden cardiac death has a close statistical relationship with the time of day, viz., ventricular fibrillation is most likely to occur between 12am-6am, with 6pm-12am being the next most likely period. Consequently there has been significant interest in understanding how cardiac activity is influenced by the circadian clock, i.e., temporal oscillations in physiological activity with a period close to 24 hours and synchronized with the day-night cycle. Although studies have identified the genetic basis of circadian rhythm at the intracellular level, the mechanisms by which they influence cardiac pathologies are not yet fully understood. Evidence has suggested that diurnal variations in the conductance properties of ion channel proteins that govern the excitation dynamics of cardiac cells may provide the crucial link. In this paper, we investigate the relationship between the circadian rhythm as manifested in modulations of ion channel properties and the susceptibility to cardiac arrhythmias by using a mathematical model that describes the electrical activity in ventricular tissue. We show that changes in the channel conductance that lead to extreme values for the duration of action potentials in cardiac cells can result either in abnormally high-frequency reentrant activity or spontaneous conduction block of excitation waves. Both phenomena increase the likelihood of wavebreaks that are known to initiate potentially life- threatening arrhythmias. Thus, disruptive cardiac excitation dynamics are most likely to occur in time-intervals of the day-night cycle during which the channel properties are closest to these extreme values, providing an intriguing relation between circadian rhythms and cardiac pathologies.

Implications of Kir/Nav1.5 Reciprocity for Propagating Action Potentials in Cardiac Cells

The main ion channels involved in the propagation of action potentials in cardiac cells are the voltage-gated sodium channels, the inward rectifier potassium channels, and the inter-cellular gap junction channels. Sodium channels have been shown to be localized to the intercalated disks along with Cx43 channels. Recent evidence of reciprocity in the co-localized expression of Kir2.1 and Nav1.5 channels in cardiac myocytes suggest that ionic currents due to these two channels are in some way calibrated to each other. In isolated cells, the fast sodium current is much larger than IK1 resulting in a large depolarization reserve. Using simulations of chains of cardiac cells, we show that the depolarization reserve for conducting action potentials is significantly smaller than in isolated cells. We also studied the changes in the depolarization reserve with variations in gap junction channel density and explored the conditions under which propagation slowed or failed. These insights will allow a better understanding of the effects of Na channel blockers as well as regional differences in action potential conduction in the heart.

Map-based model of the cardiac action potential

A simple computationally efficient model which is capable of replicating the basic features of cardiac cell action potential is proposed. The model is a four-dimensional map and demonstrates good correspondence with real cardiac cells. Various regimes of cardiac activity, which can be reproduced by the proposed model, are shown. Bifurcation mechanisms of these regimes transitions are explained using phase space analysis. The dynamics of 1D and 2D lattices of coupled maps which model the behavior of electrically connected cells is discussed in the context of synchronization theory.

Osmotic modulation of slowly activating IKs in guinea-pig ventricular myocytes

The aims of the study were to determine the effects of anisosmotic bathing solution on selected properties of I(Ks), the slowly activating delayed-rectifier K(+) current important for repolarization of the action potential in cardiac cells. Guinea-pig ventricular myocytes were voltage-clamped using either the ruptured-patch or perforated-patch technique, and the amplitude, time course, and voltage dependence of I(Ks) were determined before [isosmotic (1T)] and during superfusion of hyposmotic (<1T) or hyperosmotic (>1T) bathing solution. Hyposmotic solution increased the amplitude of I(Ks), and hyperosmotic solution decreased it. Anisosmotic-induced changes in I(Ks) amplitude were complete in 2-5 min, well-maintained, reversible, and not accompanied by significant changes in I(Ks) time course and voltage dependence. There was little difference in the results obtained with the ruptured-patch technique and those obtained with the perforated-patch technique. The amplitude of I(Ks) was sensitive to small (±10%) changes in osmolarity, maximally increased by hyposmotic solution with T < 0.7, and strongly decreased by hyperosmotic solution with T > 1.5. Experimental data on a plot of relative (1T = 1.0) I(Ks) amplitude vs. the reciprocal of relative osmolarity are well-described by a Hill equation that has a lower asymptote of 0.0, an upper asymptote of 2.0, and a slope factor of 1.87 ± 0.07. Modulation of I(Ks) amplitude by anisosmotic solution is independent of patch configuration, unaccompanied by changes in current gating, and well-described by a Hill dose-response relation that predicts relatively strong responses of I(Ks) to small perturbations in external osmolarity.

Defining candidate drug characteristics for Long-QT (LQT3) syndrome

Mutations of the SCN5A gene can significantly alter the function of cardiac myocyte sodium channels leading to increased risk of ventricular arrhythmia. Over the past decade, detailed Markov models of the action potential of cardiac cells have been developed. In such models, the effects of a drug can be treated as alterations in on- and off rates between open and inactivated states on one hand, and blocked states on the other hand. Our aim is to compute the rates specifying a drug in order to: (a) restore the steady-state open probability of the mutant channel to that of normal wild type channels; and (b) minimize the difference between whole cell currents in drugged mutant and wild type cells. The difference in the electrochemical state vector of the cell can be measured in a norm taking all components and their dynamical properties into account. Measured with this norm, the difference between the state of the mutant and wild-type cell was reduced by a factor of 36 after the drug was introduced and by factors of 4 over mexitiline and 25 over lidocaine. The results suggest the potential to synthesize more effective drugs based on mechanisms of action of existing compounds.

The Role of Kinesin I and a Small Gtpase in the Forward Trafficking of Kv1.5 Channels

Kv channels play important roles in the repolarization phase of the action potential in cardiac cells. The regulation of functional Kv1.5 surface expression has been reported to be modulated by retrograde trafficking through dynein motor but little is known about regulation by forward trafficking. Here, we use electrophysiological and immunocytochemical methods to investigate the mechanisms and regulation of anterograde trafficking of newly synthesized Kv1.5 channel proteins in cultured cells and in adult cardiomyocytes. Over-expression of a kinesin I isoform (Kif5b) increased outward K+ current by two fold in cultured cells stably expressing Kv1.5. This enhancement of Kv1.5 current by Kif5b was blocked by a six hour treatment with Brefeldin A. Over-expression of Kif5b increased Kv1.5 current additively with inhibition of endocytosis by p50 over-expression and dynamin inhibitory peptide. Deletion of a specific SH3-binding domain in Kv1.5 that is essential for internalization of the channel similarly enhanced Kif5b-induced Kv1.5 current. Expression of a dominant negative Kif5b mutant prior to induction of Kv1.5 in a tetracycline-inducible system almost completely blocked Kv1.5 current. These results demonstrate that Kif5b is required for the forward trafficking of newly synthesized Kv1.5 channel to the plasma membrane. This work has been extended to adult rat cardiomyocytes transfected with Kif5b constructs and wild-type and dominant negative Rab-type small GTPases. Results indicate that newly synthesized Kv1.5 traffics via a non-conventional pathway and on to the plasmalemma in a Kif5b-dependent process.

Induction of NO synthase 2 in ventricular cardiomyocytes incubated with a conventional bicarbonate dialysis bath

In hypotension-prone patients, conventional bicarbonate dialysis (BD) causes a reduced cardiovascular tolerance to the treatment with respect to acetate-free biofiltration (AFB). One possible explanation is an overproduction of endogenous NO (nitric oxide) due to the residual quote of acetate (4 mM) in the BD dialysate formulation. NO overload might cause the impairment of cardiovascular reactivity observed during BD. In this study, a potential direct impact of the residual quote of acetate on the cardiac cells was investigated. Ventricular cardiac myocytes isolated from adult rat hearts were treated with three different dialysis baths with or without acetate: BD, AFB and AFB + 4 mM of acetate (AFB(+)). Corresponding levels of expression of the inducible NO synthase 2 (NOS2) were assessed after the treatments along with the measurement of single-cell action potential (AP). Incubation with acetate-containing dialysis solutions significantly enhanced (P < 0.05) the expression of NOS2 mRNA (BD: 1.11 +/- 0.31; AFB(+): 0.73 +/- 0.04, NOS2/GAPDH intensitometric ratio) with respect to the acetate-free bath (AFB: 0.03 +/- 0.01). Accordingly, protein translation was also enhanced (BD: 0.176 +/- 0.021; AFB(+): 0.135 +/- 0.009, NOS2/alpha-tubuline intensitometric ratio) with respect to AFB (0.002 +/- 0.001, P < 0.05). Measurement of the AP indicates that acetate-containing solutions determine a shortening of the repolarization phase as compared to treatment with AFB (BD: 95 +/- 13; AFB(+): 76 +/- 10; AFB: 162 +/- 16 ms). These findings show that the residual quote of acetate of the BD bath formulation affects the expression of NOS2 and the duration of AP in cardiac cells. This might cause the cardiac contractile impairment in unstable patients during BD treatment.