Action Potential Shape Research Articles

Steady-state and transient effects of SK channel block and adrenergic stimulation to counteract acetylcholine-induced arrhythmogenic effects in the human atria: A computational study

Hyperactivity of the parasympathetic nervous system has been linked to the development of paroxysmal atrial fibrillation (AF). The parasympathetic neurotransmitter acetylcholine (ACh) causes a reduction in action potential (AP) duration (APD) and an increase in resting membrane potential (RMP), both of which contribute to enhance the risk for reentry. Research suggests that small-conductance calcium activated potassium (SK) channels may be an effective target for treating AF. Therapies targeting the autonomic nervous system, either alone or in combination with other drugs, have been explored and have been shown to decrease the incidence of atrial arrhythmias. This study uses computational modeling and simulation to examine the impact of SK channel block (SKb) and β-adrenergic stimulation through Isoproterenol (Iso) on countering the negative effects of cholinergic activity in human atrial cell and 2D tissue models. The steady-state effects of Iso and/or SKb on AP shape, APD at 90% repolarization (APD90) and RMP were evaluated. The ability to terminate stable rotational activity in cholinergically-stimulated 2D tissue models of AF was also investigated. A range of SKb and Iso application kinetics, which reflect varying drug binding rates, were taken into consideration. The results showed that SKb alone prolonged APD90 and was able to stop sustained rotors in the presence of ACh concentrations up to 0.01 μM. Iso terminated rotors under all tested ACh concentrations, but resulted in highly-variable steady-state outcomes depending on baseline AP morphology. Importantly, the combination of SKb and Iso resulted in greater APD90 prolongation and showed promising anti-arrhythmic potential by stopping stable rotors and preventing re-inducibility.

Lobule-Related Action Potential Shape- and History-Dependent Current Integration in Purkinje Cells of Adult and Developing Mice.

Purkinje cells (PCs) are the principal cells of the cerebellar cortex and form a central element in the modular organization of the cerebellum. Differentiation of PCs based on gene expression profiles revealed two subpopulations with distinct connectivity, action potential firing and learning-induced activity changes. However, which basal cell physiological features underlie the differences between these subpopulations and to what extent they integrate input differentially remains largely unclear. Here, we investigate the cellular electrophysiological properties of PC subpopulation in adult and juvenile mice. We found that multiple fundamental cell physiological properties, including membrane resistance and various aspects of the action potential shape, differ between PCs from anterior and nodular lobules. Moreover, the two PC subpopulations also differed in the integration of negative and positive current steps as well as in size of the hyperpolarization-activated current. A comparative analysis in juvenile mice confirmed that most of these lobule-specific differences are already present at pre-weaning ages. Finally, we found that current integration in PCs is input history-dependent for both positive and negative currents, but this is not a distinctive feature between anterior and nodular PCs. Our results support the concept of a fundamental differentiation of PCs subpopulations in terms of cell physiological properties and current integration, yet reveals that history-dependent input processing is consistent across PC subtypes.

Action potential metrics and automated data analysis pipeline for cardiotoxicity testing using optically mapped hiPSC-derived 3D cardiac microtissues.

Recent advances in human induced pluripotent stem cell (hiPSC)-derived cardiac microtissues provide a unique opportunity for cardiotoxic assessment of pharmaceutical and environmental compounds. Here, we developed a series of automated data processing algorithms to assess changes in action potential (AP) properties for cardiotoxicity testing in 3D engineered cardiac microtissues generated from hiPSC-derived cardiomyocytes (hiPSC-CMs). Purified hiPSC-CMs were mixed with 5-25% human cardiac fibroblasts (hCFs) under scaffold-free conditions and allowed to self-assemble into 3D spherical microtissues in 35-microwell agarose gels. Optical mapping was performed to quantify electrophysiological changes. To increase throughput, AP traces from 4x4 cardiac microtissues were simultaneously acquired with a voltage sensitive dye and a CMOS camera. Individual microtissues showing APs were identified using automated thresholding after Fourier transforming traces. An asymmetric least squares method was used to correct non-uniform background and baseline drift, and the fluorescence was normalized (ΔF/F0). Bilateral filtering was applied to preserve the sharpness of the AP upstroke. AP shape changes under selective ion channel block were characterized using AP metrics including stimulation delay, rise time of AP upstroke, APD30, APD50, APD80, APDmxr (maximum rate change of repolarization), and AP triangulation (APDtri = APDmxr-APD50). We also characterized changes in AP metrics under various ion channel block conditions with multi-class logistic regression and feature extraction using principal component analysis of human AP computer simulations. Simulation results were validated experimentally with selective pharmacological ion channel blockers. In conclusion, this simple and robust automated data analysis pipeline for evaluating key AP metrics provides an excellent in vitro cardiotoxicity testing platform for a wide range of environmental and pharmaceutical compounds.

Re-assessing the role of the helix bundle crossing in NaK.

Potassium channels play the main role in shaping action potentials across all living organisms. This function is owed to their ability to open and close in response to stimuli, and thus conduct ions in a selective and regulated fashion. Small prokaryotic potassium channels, such as KcsA, have become model systems for investigations of structure, dynamics and function. These channels are amenable to studies by various biophysical techniques and have been used to obtain insight into the molecular mechanisms of ion channel gating and selectivity. In the case of KcsA, the inner gate is allosterically coupled with the selectivity filter, and several ion conduction mechanisms across the selectivity filter have been proposed. To understand how general these gating and selectivity mechanisms are, we have used solution NMR spectroscopy and solid-supported membrane electrophysiology (SSME) to study another small prokaryotic channel NaK. While NaK maintains a very high degree of structural conservation to potassium channel pores, the conformation of the NaK selectivity filter is different. We show that while NaK still displays allosteric coupling between the selectivity filter and the helix bundle crossing, akin to what was observed for KcsA, the helix bundle crossing remains fully open in both conductive and non-conductive channels. In addition, using SSME we find that ion conduction by NaK shows K+ selectivity in the absence of an electric gradient. These observations contrast with the most recent models proposed by NMR and MD simulations on KcsA and other channels.

Biophysical properties of Nav1.5 channel in ventricular-like and atrial-like chamber cardiomyocytes derived from human induced pluripotent stem cells.

Cardiomyocytes derived from human induced pluripotent stem cells (hiPSC-CMs) is a promising way modeling cardiac diseases and investigating their associated mechanisms. Nevertheless, it is still challenging to differentiate hiPSC-CMs into specific cardiac types to investigate differences in ion channel properties such as for the NaV1.5 channel. In this context, we investigated biophysical properties of NaV1.5 in ventricular-like and atrial-like hiPSC-CMs. Atrial-like cardiomyocytes were obtained from the differentiation of hiPSCs treated with retinoic acid (RA). The quality of atrial specification was assessed by immunocytofluorescence, qPCR and western blot. Calcium and sodium currents were recorded using the Patch-clamp technique and their electrophysiological properties were investigated using optical mapping technique. hiPSC-CMs differentiation with RA induced a higher expression of mlc2a protein, a marker of atrial cardiomyocytes, and a lower expression of mlc2v and Cx43 proteins, markers of ventricular cardiomyocytes. The amplitude, the duration, and the steady-state phase of action potentials (APs) were decreased in RA-differentiated cardiomyocytes, corresponding to the atrial AP shape. Interestingly, no difference in SCN5A gene expression was detected whereas changes in NaV1.5 currents were observed. Indeed, lower current densities and a shift of the activation and inactivation to more depolarized membrane potential were measured in RA-differentiated cardiomyocytes. Furthermore, the kinetics of the fast inactivation were slower in atrial-like cardiomyocytes. These results showed that RA treatment allowed to obtain atrial-like cardiomyocytes and permitted to investigate differences in NaV1.5 channel properties between ventricular and atrial compartments. This approach with RA could represent an interesting way to investigate specific-type channelopathies.

Application of Convolutional Neural Networks Using Action Potential Shape for In-Silico Proarrhythmic Risk Assessment.

This study proposes a convolutional neural network (CNN) model using action potential (AP) shapes as input for proarrhythmic risk assessment, considering the hypothesis that machine-learning features automatically extracted from AP shapes contain more meaningful information than do manually extracted indicators. We used 28 drugs listed in the comprehensive in vitro proarrhythmia assay (CiPA), consisting of eight high-risk, eleven intermediate-risk, and nine low-risk torsadogenic drugs. We performed drug simulations to generate AP shapes using experimental drug data, obtaining 2000 AP shapes per drug. The proposed CNN model was trained to classify the TdP risk into three levels, high-, intermediate-, and low-risk, based on in silico AP shapes generated using 12 drugs. We then evaluated the performance of the proposed model for 16 drugs. The classification accuracy of the proposed CNN model was excellent for high- and low-risk drugs, with AUCs of 0.914 and 0.951, respectively. The model performance for intermediate-risk drugs was good, at 0.814. Our proposed model can accurately assess the TdP risks of drugs from in silico AP shapes, reflecting the pharmacokinetics of ionic currents. We need to secure more drugs for future studies to improve the TdP-risk-assessment robustness.

Resting membrane potential is less negative in trabeculae from right atrial appendages of women, but action potential duration does not shorten with age

AimsThe incidence of atrial fibrillation (AF) increases with age. Women have a lower risk. Little is known on the impact of age, sex and clinical variables on action potentials (AP) recorded in right atrial tissue obtained during open heart surgery from patients in sinus rhythm (SR) and in longstanding AF. We here investigated whether age or sex have an impact on the shape of AP recorded in vitro from right atrial tissue. MethodsWe performed multivariable analysis of individual AP data from trabeculae obtained during heart surgery of patients in SR (n = 320) or in longstanding AF (n = 201). AP were recorded by sharp microelectrodes at 37 °C at 1 Hz. Impact of clinical variables were modeled using a multivariable mixed model regression. ResultsIn SR, AP duration at 90% repolarization (APD90) increased with age. Lower ejection fraction and higher body mass index were associated with smaller action potential amplitude (APA) and maximum upstroke velocity (Vmax). The use of beta-blockers was associated with larger APD90. In tissues from women, resting membrane potential was less negative and APA as well as Vmax were smaller. Besides shorter APD20 in elderly patients, effects of age and sex on atrial AP were lost in AF. ConclusionThe higher probability to develop AF at advanced age cannot be explained by a shortening in APD90. Less negative RMP and lower upstroke velocity might contribute to lower incidence of AF in women, which may be of clinical relevance.

Coupling the Cardiac Voltage-Gated Sodium Channel to Channelrhodopsin-2 Generates Novel Optical Switches for Action Potential Studies.

Voltage-gated sodium (Na+) channels respond to short membrane depolarization with conformational changes leading to pore opening, Na+ influx, and action potential (AP) upstroke. In the present study, we coupled channelrhodopsin-2 (ChR2), the key ion channel in optogenetics, directly to the cardiac voltage-gated Na+ channel (Nav1.5). Fusion constructs were expressed in Xenopus laevis oocytes, and electrophysiological recordings were performed by the two-microelectrode technique. Heteromeric channels retained both typical Nav1.5 kinetics and light-sensitive ChR2 properties. Switching to the current-clamp mode and applying short blue-light pulses resulted either in subthreshold depolarization or in a rapid change of membrane polarity typically seen in APs of excitable cells. To study the effect of individual K+ channels on the AP shape, we co-expressed either Kv1.2 or hERG with one of the Nav1.5-ChR2 fusions. As expected, both delayed rectifier K+ channels shortened AP duration significantly. Kv1.2 currents remarkably accelerated initial repolarization, whereas hERG channel activity efficiently restored the resting membrane potential. Finally, we investigated the effect of the LQT3 deletion mutant ΔKPQ on the AP shape and noticed an extremely prolonged AP duration that was directly correlated to the size of the non-inactivating Na+ current fraction. In conclusion, coupling of ChR2 to a voltage-gated Na+ channel generates optical switches that are useful for studying the effect of individual ion channels on the AP shape. Moreover, our novel optogenetic approach provides the potential for an application in pharmacology and optogenetic tissue-engineering.

Laser-Induced Action Potential-Like Measurements of Cardiomyocytes on Microelectrode Arrays for Increased Predictivity of Safety Pharmacology.

Life-threatening drug-induced cardiac arrhythmia is often preceded by prolonged cardiac action potentials (AP), commonly accompanied by small proarrhythmic membrane potential fluctuations. The shape and time course of the repolarizing fraction of the AP can be pivotal for the presence or absence of arrhythmia. Microelectrode arrays (MEA) allow easy access to cardiotoxic compound effects via extracellular field potentials (FP). Although a powerful and well-established tool in research and cardiac safety pharmacology, the FP waveform does not allow to infer the original AP shape due to the extracellular recording principle and the resulting intrinsic alternating current (AC) filtering. A novel device, described here, can repetitively open the membrane of cardiomyocytes cultivated on top of the MEA electrodes at multiple cultivation time points, using a highly focused nanosecond laser beam. The laser poration results in transforming the electrophysiological signal from FP to intracellular-like APs (laser-induced AP, liAP) and enables the recording of transcellular voltage deflections. This intracellular access allows a better description of the AP shape and a better and more sensitive classification of proarrhythmic potentials than regular MEA recordings. This system is a revolutionary extension to the existing electrophysiological methods, permitting accurate evaluation of cardiotoxic effect with all advantages of MEA-based recordings (easy, acute, and chronic experiments, signal propagation analysis, etc.).

Morphological Determinants of Cell-to-Cell Variations in Action Potential Dynamics in Substantia Nigra Dopaminergic Neurons.

Action potential (AP) shape is a critical electrophysiological parameter, in particular because it strongly modulates neurotransmitter release. As it greatly varies between neuronal types, AP shape is also used to distinguish neuronal populations. For instance, AP duration ranges from hundreds of microseconds in cerebellar granule cells to 2-3 ms in SNc dopaminergic (DA) neurons. While most of this variation across cell types seems to arise from differences in the voltage- and calcium-gated ion channels expressed, a few studies suggested that dendritic morphology also affects AP shape. AP duration also displays significant variability in a same neuronal type, although the determinants of these variations are poorly known. Using electrophysiological recordings, morphological reconstructions, and realistic Hodgkin-Huxley modeling, we investigated the relationships between dendritic morphology and AP shape in rat SNc DA neurons from both sexes. In this neuronal type where the axon arises from an axon-bearing dendrite (ABD), the duration of the somatic AP could be predicted from a linear combination of the ABD and non-ABDs' complexities. Dendrotomy experiments and simulation showed that these correlations arise from the causal influence of dendritic topology on AP duration, due in particular to a high density of sodium channels in the somatodendritic compartment. Surprisingly, computational modeling suggested that this effect arises from the influence of sodium currents on the decaying phase of the AP. Consistent with previous findings, these results demonstrate that dendritic morphology plays a major role in defining the electrophysiological properties of SNc DA neurons and their cell-to-cell variations.SIGNIFICANCE STATEMENT Action potential (AP) shape is a critical electrophysiological parameter, in particular because it strongly modulates neurotransmitter release. AP shape (e.g., duration) greatly varies between neuronal types but also within a same neuronal type. While differences in ion channel expression seem to explain most of AP shape variation across cell types, the determinants of cell-to-cell variations in a same neuronal type are mostly unknown. We used electrophysiological recordings, neuronal reconstruction, and modeling to show that, because of the presence of sodium channels in the somatodendritic compartment, a large part of cell-to-cell variations in somatic AP duration in substantia nigra pars compacta dopaminergic neurons is explained by variations in dendritic topology.

Spatial uniformity of action potentials indicates base-to-apex depolarization and repolarization of rainbow trout (Oncorhynchus mykiss) ventricle.

The spatial pattern of electrical activation is crucial for a full understanding of fish heart function. However, it remains unclear whether there is regional variation in action potential (AP) morphologies and underlying ion currents. Because the direction of depolarization and spatial differences in the durations of ventricular APs set limits to potential patterns of ventricular repolarization, we determined AP morphologies, underlying ion currents and ion channel expression in four different ventricular regions (spongy myocardium; and apex, base and middle of the compact myocardium), and correlated them with in vivo electrocardiograms (ECGs) in rainbow trout (Oncorhynchus mykiss). ECGs recorded from three leads indicated that the depolarization and repolarization of APs propagate from base to apex, and the main depolarization axis of the ventricle is between +90 and +120deg. AP shape was uniform across the whole ventricle, and little regional differences were found in the density of repolarizing K+ currents or depolarizing Ca2+ and Na+ currents and the underlying transcripts of ion channels, providing compelling evidence for the suggested excitation pattern. The spatial uniformity of AP durations and base-to-apex propagation of activation with a relatively slow velocity of propagation indicates no special ventricular conduction pathway in the trout ventricle such as the His-Purkinje system of mammalian hearts. The sequence of repolarization is solely determined by activation time without being affected by regional differences in AP duration.

Abstract P1090: Polycystin-1 Regulates Calcium Handling And Contractility In Human IPSC-derived Ventricular Cardiomyocytes

Mutations in polycystin-1 (PC1) cause 85% of autosomal dominant polycystic kidney disease cases. This disease is characterized by progressive renal disease and multiple cardiovascular alterations. Evidence suggests that cardiovascular disease precedes the onset of renal failure. We recently demonstrated that PC1 ablation shortens cardiomyocyte action potential, decreases Ca 2+ transient amplitude, and reduces contractility in adult cardiomyocytes isolated from cardiomyocyte-specific PC1 knock-out mice compared to control mice. Alterations in Ca 2+ cycling in PC1-deficient cardiomyocytes were demonstrated to be due to action potential duration shortening and impaired sarcoendoplasmic reticulum ATPase activity. However, mouse and human cardiomyocytes have considerable differences in their action potential shapes and the ion channels involved. It remains unknown whether PC1 regulates these events in human cardiomyocytes. Here, we sought to investigate whether PC1 regulates excitation-contraction coupling in human ventricular cardiomyocytes. We differentiated commercially available human-induced pluripotent stem cell lines (hiPSC) into ventricular cardiomyocytes (hiPSC-CM) using published protocols (measurements were performed 60-90 days after differentiation). We developed a protocol to simultaneously measure action potentials (FluoVolt), intracellular Ca 2+ (Calbryte-590) and contractility using the IonOptix algorithm to detect changes in contrast as contractility measurements. PC1 ablation using siRNA decreased action potential duration and impaired Ca 2+ transients and contractility in hiPSC-CMs. Because hiPSC-CMs have several limitations due to their immature state, we employed a well-validated method to improve their maturation that utilizes a Matrigel mattress and hormone supplementation (dexamethasone; Dexa / thyroid hormone; T3; PMID: 28974554; developed by Dr. Bjorn Knollmann’s lab). Using the Matrigel mattress/Dexa/T3 technique, we increased hiPSC-CM maturation measured as follows: cardiomyocytes were oriented and contracted in the longitudinal axis, ryanodine receptor type 2 and alpha-actinin localized with a striated pattern, exhibited robust inward K + rectifier currents (I K1 ) and fired action potentials that resemble human adult cardiomyocytes. In this model, PC1 knockdown also decreased action potential duration and impaired Ca 2+ transients and decreased contractility measured as fractional shortening. Altogether, this evidence shows for the first time that PC1 regulates human cardiomyocyte physiology and suggests that cardiac alterations in patients with autosomal dominant polycystic kidney disease may emerge as a result of impaired cardiomyocyte contractile function.

Sensitivity to extracellular potassium underlies type-intrinsic differences in retinal ganglion cell excitability.

Neuronal type-specific physiologic heterogeneity can be driven by both extrinsic and intrinsic mechanisms. In retinal ganglion cells (RGCs), which carry visual information from the retina to central targets, evidence suggests intrinsic properties shaping action potential (AP) generation significantly impact the responses of RGCs to visual stimuli. Here, we explored how differences in intrinsic excitability further distinguish two RCG types with distinct presynaptic circuits, alpha ON-sustained (αON-S) cells and alpha OFF-sustained (αOFF-S) cells. We found that αOFF-S RGCs are more excitable to modest depolarizing currents than αON-S RGCs but excitability plateaued earlier as depolarization increased (i.e., depolarization block). In addition to differences in depolarization block sensitivity, the two cell types also produced distinct AP shapes with increasing stimulation. αOFF-S AP width and variability increased with depolarization magnitude, which correlated with the onset of depolarization block, while αON-S AP width and variability remained stable. We then tested if differences in depolarization block observed in αON-S and αOFF-S RGCs were due to sensitivity to extracellular potassium. We found αOFF-S RGCs more sensitive to increased extracellular potassium concentration, which shifted αON-S RGC excitability to that of αOFF-S cells under baseline potassium conditions. Finally, we investigated the influence of the axon initial segment (AIS) dimensions on RGC spiking. We found that the relationship between AIS length and evoked spike rate varied not only by cell type, but also by the strength of stimulation, suggesting AIS structure alone cannot fully explain the observed differences RGC excitability. Thus, sensitivity to extracellular potassium contributes to differences in intrinsic excitability, a key factor that shapes how RGCs encode visual information.

Microphysiological Modeling of the Structure and Function of Neuromuscular Transmitter Release Sites.

The general mechanism of calcium-triggered chemical transmitter release from neuronal synapses has been intensely studied, is well-known, and highly conserved between species and synapses across the nervous system. However, the structural and functional details within each transmitter release site (or active zone) are difficult to study in living tissue using current experimental approaches owing to the small spatial compartment within the synapse where exocytosis occurs with a very rapid time course. Therefore, computer simulations offer the opportunity to explore these microphysiological environments of the synapse at nanometer spatial scales and on a sub-microsecond timescale. Because biological reactions and physiological processes at synapses occur under conditions where stochastic behavior is dominant, simulation approaches must be driven by such stochastic processes. MCell provides a powerful simulation approach that employs particle-based stochastic simulation tools to study presynaptic processes in realistic and complex (3D) geometries using optimized Monte Carlo algorithms to track finite numbers of molecules as they diffuse and interact in a complex cellular space with other molecules in solution and on surfaces (representing membranes, channels and binding sites). In this review we discuss MCell-based spatially realistic models of the mammalian and frog neuromuscular active zones that were developed to study presynaptic mechanisms that control transmitter release. In particular, these models focus on the role of presynaptic voltage-gated calcium channels, calcium sensors that control the probability of synaptic vesicle fusion, and the effects of action potential waveform shape on presynaptic calcium entry. With the development of these models, they can now be used in the future to predict disease-induced changes to the active zone, and the effects of candidate therapeutic approaches.

Optogenetic manipulation of cardiac electrical dynamics using sub-threshold illumination: dissecting the role of cardiac alternans in terminating rapid rhythms

Abstract Funding Acknowledgements Type of funding sources: None. Cardiac action potential (AP) shape and propagation are regulated by several key dynamic factors such as ions channel recovery and intracellular Ca2+-cycling. Experimental methods for manipulating AP electrical dynamics commonly use ion channel inhibitors that lack spatial and temporal specificity. In this work, we propose a novel approach based on optogenetics to manipulate cardiac electrical activity employing a light-modulated depolarizing current with intensities that are too low to elicit APs (sub-threshold illumination) but are sufficient to fine-tune AP electrical dynamics. We investigated the effects of sub-threshold illumination in isolated cardiomyocytes and whole hearts by using transgenic mice constitutively expressing a light-gated ion channel (channelrhodopsins-2, ChR2). We find that ChR2-mediated depolarizing current prolongs APs and reduces conduction velocity (CV) in a space-selective and reversible manner. Sub-threshold manipulation also affects the dynamics of cardiac electrical activity, increasing the magnitude of cardiac alternans. We use an optical system that uses real-time feedback control to generate re-entrant circuits with user-defined cycle lengths to explore the role that cardiac alternans plays in spontaneous termination of ventricular tachycardias (VTs). We find that VT stability significantly decreases during sub-threshold illumination primarily due to an increase in the amplitude of electrical oscillations, which implies that cardiac alternans may be beneficial in the context of self-termination of VT.

Electrocardiographic marker of the cardiac action potential triangulation induced by antiarrhythmic drugs in perfused guinea-pig heart.

What is the central question of this study? Can the triangular appearance of ventricular action potential, indicating proarrhythmic profile of antiarrhythmic agent, be approximated by specific changes on an electrocardiogram (ECG)? What are the main finding and its importance? The triangulation of the ventricular action potential seen when antiarrhythmic drugs induce a greater lengthening of the late repolarization compared to the initial repolarization in epicardium is closely approximated by a greater prolongation of the T wave upslope relative to the interval between the J point and the start of the T wave (the JTstart interval) on the ECG. These findings may improve the power of ECG assessments in predicting the drug-induced arrhythmia resulting from slowed phase 3 repolarization. Antiarrhythmic drugs prescribed to treat atrial fibrillation can occasionally precipitate ventricular tachyarrhythmia through a prominent slowing of the phase 3 repolarization. The latter results in the triangular shape of ventricular action potential, indicating high arrhythmic risk. However, clinically, the utility of triangulation assessments for predicting arrhythmia is limited owing to the invasive nature of the ventricular action potential recordings. This study examined whether the triangulation effect can be detected indirectly from electrocardiogram (ECG) analysis. Epicardial monophasic action potentials and the ECG were simultaneously recorded in perfused guinea-pig hearts. With antiarrhythmics (dofetilide, quinidine, procainamide and flecainide), a prolongation of the initial repolarization seen in the action potential recordings was closely approximated by lengthening of the interval between the J point and the start of the T wave (the JTstart interval) on the ECG, whereas a prolongation of the late repolarization was paralleled by widening of the T wave upslope. Dofetilide, quinidine and procainamide induced a prominent slowing of the phase 3 repolarization in epicardium, leading to triangulation of the action potential. These effects were accompanied by a greater prolongation of the T wave upslope compared to the JTstart interval. Flecainide elicited a proportional prolongation of the initial and the late ventricular repolarization, and therefore failed to induce triangulation, based on analysis of both epicardial action potential and ECG profiles. Collectively, these findings suggest that the ratio between the durations of the T wave upslope and the JTstart interval may represent the ECG metric of the ventricular action potential triangulation induced by antiarrhythmic drugs.

Proarrhythmic risk assessment of drugs by dVm /dt shapes using the convolutional neural network.

Comprehensive in vitro Proarrhythmia Assay (CiPA) projects for assessing proarrhythmic drugs suggested a logistic regression model using qNet as the Torsades de Pointes (TdP) risk assessment biomarker, obtained from in silico simulation. However, using a single in silico feature, such as qNet, cannot reflect whole characteristics related to TdP in the entire action potential (AP) shape. Thus, this study proposed a deep convolutional neural network (CNN) model using differential action potential shapes to classify three proarrhythmic risk levels: high, intermediate, and low, considering both characteristics related to TdP not only in the depolarization phase but also the repolarization phase of AP shape. We performed an in silico simulation and got AP shapes with drug effects using half‐maximal inhibitory concentration and Hill coefficients of 28 drugs released by CiPA groups. Then, we trained the deep CNN model with the differential AP shapes of 12 drugs and tested it with those of 16 drugs. Our model had a better performance for classifying the proarrhythmic risk of drugs than the traditional logistic regression model using qNet. The classification accuracy was 98% for high‐risk level drugs, 94% for intermediate‐risk level drugs, and 89% for low‐risk level drugs.

Optogenetic manipulation of cardiac electrical dynamics using sub-threshold illumination: dissecting the role of cardiac alternans in terminating rapid rhythms

Cardiac action potential (AP) shape and propagation are regulated by several key dynamic factors such as ion channel recovery and intracellular Ca2+ cycling. Experimental methods for manipulating AP electrical dynamics commonly use ion channel inhibitors that lack spatial and temporal specificity. In this work, we propose an approach based on optogenetics to manipulate cardiac electrical activity employing a light-modulated depolarizing current with intensities that are too low to elicit APs (sub-threshold illumination), but are sufficient to fine-tune AP electrical dynamics. We investigated the effects of sub-threshold illumination in isolated cardiomyocytes and whole hearts by using transgenic mice constitutively expressing a light-gated ion channel (channelrhodopsin-2, ChR2). We find that ChR2-mediated depolarizing current prolongs APs and reduces conduction velocity (CV) in a space-selective and reversible manner. Sub-threshold manipulation also affects the dynamics of cardiac electrical activity, increasing the magnitude of cardiac alternans. We used an optical system that uses real-time feedback control to generate re-entrant circuits with user-defined cycle lengths to explore the role of cardiac alternans in spontaneous termination of ventricular tachycardias (VTs). We demonstrate that VT stability significantly decreases during sub-threshold illumination primarily due to an increase in the amplitude of electrical oscillations, which implies that cardiac alternans may be beneficial in the context of self-termination of VT.

Electrical properties of the sensory neuron and defense reactions of mollusc Lymnaea stagnalis at conditions of prolonged hyperglycemia

Hemolymph glucose level rise (from 0.12 (0.05; 0.18) to 4.10 (3.18; 6.08) mmol/L) modify the defensive behaviour of molluscs. This results in increase of the degree of animal’s body retracted into the shell and in reinforcement of weak defense reactions in response to tentacles’ tactile stimulation. No fluctuations in the tentacles length and the duration of the latent period of protraction were found. At experimental hyperglycemia (incubation of the isolated CNS preparation in 10 mmol/L D-glucose solution for 2 h), the basic electrical characteristics of FMRFamide-containing neuron RPaD1, involved in sensory stimuli detection and heart beat regulation in Lymnaea, were changed. Membrane depolarisation, accompanied by firing rate increase were observed, while current-voltage curve characteristics, as well as membrane resistance, capacity and time constant remains unchanged in comparison with control. An increase in rising and falling phases duration, undershoot amplitude were noted, while other parameters of the RPaD1 spike remained unchanged. Based on the revealed features of the action potential shape at hyperglycemia, it is assumed that these changes can be caused by the activation of the Na+-glucose co-transporter and ATP-sensitive K+-channels of the RPaD1 membrane. The action of glucose should be considered as a metabolic signal also in relation to non-feeding neurons in the mollusc brain.

Late Sodium Current of the Heart: Where Do We Stand and Where Are We Going?

Late sodium current has long been linked to dysrhythmia and contractile malfunction in the heart. Despite the increasing body of accumulating information on the subject, our understanding of its role in normal or pathologic states is not complete. Even though the role of late sodium current in shaping action potential under physiologic circumstances is debated, it’s unquestioned role in arrhythmogenesis keeps it in the focus of research. Transgenic mouse models and isoform-specific pharmacological tools have proved useful in understanding the mechanism of late sodium current in health and disease. This review will outline the mechanism and function of cardiac late sodium current with special focus on the recent advances of the area.