Prediction of arrhythmia susceptibility through mathematical modeling and machine learning

At present, the QT interval on the electrocardiographic (ECG) waveform is the most common metric for assessing an individual’s susceptibility to ventricular arrhythmias, with a long QT, or, at the cellular level, a long action potential duration (APD) considered high risk. However, the limitations of this simple approach have long been recognized. Here, we sought to improve prediction of arrhythmia susceptibility by combining mechanistic mathematical modeling with machine learning (ML). Simulations with a model of the ventricular myocyte were performed to develop a large heterogenous population of cardiomyocytes (n = 10,586), and we tested each variant’s ability to withstand three arrhythmogenic triggers: 1) block of the rapid delayed rectifier potassium current (IKr Block), 2) augmentation of the L-type calcium current (ICaL Increase), and 3) injection of inward current (Current Injection). Eight ML algorithms were trained to predict, based on simulated AP features in preperturbed cells, whether each cell would develop arrhythmic dynamics in response to each trigger. We found that APD can accurately predict how cells respond to the simple Current Injection trigger but cannot effectively predict the response to IKr Block or ICaL Increase. ML predictive performance could be improved by incorporating additional AP features and simulations of additional experimental protocols. Importantly, we discovered that the most relevant features and experimental protocols were trigger specific, which shed light on the mechanisms that promoted arrhythmia formation in response to the triggers. Overall, our quantitative approach provides a means to understand and predict differences between individuals in arrhythmia susceptibility.

High-Throughput Drug Screening System Based on Human Induced Pluripotent Stem Cell-Derived Atrial Myocytes ∼ A Novel Platform to Detect Cardiac Toxicity for Atrial Arrhythmias

Evaluation of proarrhythmic properties is critical for drug discovery. In particular, QT prolongation in electrocardiograms has been utilized as a surrogate marker in many evaluation systems to assess the risk of torsade de pointes and lethal ventricular arrhythmia. Recently, new evaluation systems based on human iPS cell-derived cardiomyocytes have been established. On the other hand, in clinical situations, it has been reported that the incidence of atrial arrhythmias such as atrial fibrillation has been increasing every year, with the prediction of a persistent increase in the near future. As to the increased incidence of atrial arrhythmias, in addition to the increased population of geriatric patients, a wide variety of drug treatments may be related, as an experimental method to detect drug-induced atrial arrhythmia has not been established so far. In the present study, we characterized the atrial-like cardiomyocytes derived from human induced pluripotent stem cells and examined their potential for the evaluation of drug-induced atrial arrhythmia. Atrial-like cardiomyocytes were induced by adding retinoic acid (RA) during the process of myocardial differentiation, and their characteristics were compared to those of RA-free cardiomyocytes. Using gene expression and membrane potential analysis, it was confirmed that the cells with or without RA treatment have atrial or ventricular like cardiomyocytes, respectively. Using the ultra-rapid activating delayed rectifier potassium current (IKur) channel inhibitor, which is specific to atrial cardiomyocytes, Pulse width duration (PWD) 30cF prolongation was confirmed only in atrial-like cardiomyocytes. In addition, ventricular like cardiomyocytes exhibited an early after depolarization by treatment with rapidly activating delayed rectifier potassium current (IKr) channel inhibitor, which induces ventricular arrhythmia in clinical situations. Here, we have established a high-throughput drug evaluation system using human iPS cell-derived atrial-like cardiomyocytes. Based on the obtained data, the system might be a valuable platform to detect potential risks for drug-induced atrial arrhythmias.

KEY FACTORS TO THE RATE DEPENDENCE OF EARLY AFTERDEPOLARIZATIONS: A SIMULATION STUDY

Early afterdepolarizations (EADs) in cardiac myocytes have been reported to be associated with a series of cardiac arrhythmias. The generation of EADs has a high correlation with the excitation period, leading to repolarization dispersion over the cardiac tissue. However, the mechanism of EAD rate dependence has not been thoroughly revealed. In this study, the simulation approach was used to investigate the mechanism underlying EAD rate dependence. The results indicated that the gating variable of the delayed rectifier potassium current ([Formula: see text]-gate) and the intracellular sodium ion concentration ([Na[Formula: see text]][Formula: see text] were key factors contributing to EAD rate dependence. Also, different mathematical models showed different types of EAD rate dependence, which needs to be considered in the future simulation research related to EADs.

Neurohormonal Regulation of I Ks in Heart Failure: Implications for Ventricular Arrhythmogenesis and Sudden Cardiac Death

Heart failure (HF) results in sustained alterations in neurohormonal signaling, including enhanced signaling through the sympathetic nervous system and renin‐angiotensin‐aldosterone system pathways. While enhanced sympathetic nervous system and renin‐angiotensin‐aldosterone system activity initially help compensate for the failing myocardium, sustained signaling through these pathways ultimately contributes to HF pathophysiology. HF remains a leading cause of mortality, with arrhythmogenic sudden cardiac death comprising a common mechanism of HF‐related death. The propensity for arrhythmia development in HF occurs secondary to cardiac electrical remodeling that involves pathological regulation of ventricular ion channels, including the slow component of the delayed rectifier potassium current, that contribute to action potential duration prolongation. To elucidate a mechanistic explanation for how HF‐mediated electrical remodeling predisposes to arrhythmia development, a multitude of investigations have investigated the specific regulatory effects of HF‐associated stimuli, including enhanced sympathetic nervous system and renin‐angiotensin‐aldosterone system signaling, on the slow component of the delayed rectifier potassium current. The objective of this review is to summarize the current knowledge related to the regulation of the slow component of the delayed rectifier potassium current in response to HF‐associated stimuli, including the intracellular pathways involved and the specific regulatory mechanisms.

P938Regulation of the delayed rectifier potassium current during the adrenergic adaptation of cardiomyocytes

Funding Acknowledgements Supported by the ÚNKP-19-3 New National Excellence program of the Ministry for Innovation and Technology Introduction and aims Adaptation of the human heart to physical activity is a complex mechanism that includes the change of heart rate, morphology of the action potential (AP) among others. Stimulation of β-adrenergic receptors (β-AR) causes the shortening of the AP duration of ventricular cardiomyocytes. This is caused by the regulation of the potassium currents by the β-adrenergic signaling pathway. Our aim was to investigate the role of protein kinase A (PKA) and calcium/calmodulin-dependent protein kinase II (CaMKII) in the regulation of the slow component (IKs) of the delayed rectifier potassium current under β-AR activation. Methods Our experiments were performed on isolated canine cardiomyocytes from the left ventricle. The IKs current profile was determined under a ventricular AP. We used "AP voltage clamp" conditions in six experimental groups: Control (CTRL), β-AR stimulation with isoproterenol (ISO), CaMKII inhibition with KN-93 (KN-93), PKA inhibition with H-89 (H-89) β-AR stimulation with inhibited CaMKII (KN-93 + ISO), β-AR stimulation with inhibited PKA (H-89 + ISO). β-AR stimulation with inhibited CaMKII and PKA (KN-93 + H-89 + ISO) Results The highest current density of IKs was approximately 6 times higher and the charge delivered by IKs was about 8 times larger in the ISO group than in CTRL or KN-93 conditions. In the KN-93 + ISO group, IKs amplitude was about 60% smaller and delivered about half the total charge compared to the ISO group. In the H‑89 + ISO group, IKs was about 30% smaller and delivered 40% less total charge than in the ISO group. In the KN-93 + H-89 + ISO group the IKs did not changed sicnificantly. Conclusion Based on our results, CaMKII plays an important role in regulating IKs by β-AR stimulation.

Tyrosine-protein kinase Src regulates Kv1.5 channel activity and membrane expression through interaction with the N-terminus of the channel

Kv1.5 is a voltage-gated potassium channel that generates the ultra-rapid delayed rectifier potassium current (IKur) important in the repolarization of the atrial action potential. Malfunction of the Kv1.5 channel often results in atrial fibrillation (AFib). A reduction in Kv1.5 current (IKv1.5) occurs upon activation of the endogenous tyrosine-protein kinase Src. The Src SH3 domain binds to proline-rich motifs located within the N-terminus of Kv1.5. Disruption of these binding motifs has been involved in the development of familial AFib. The mechanism underlying the reduction of IKv1.5 upon Src activation has not yet been established and the relationship between Kv1.5 and Src is poorly understood. Therefore, the present study aims to further elucidate the mechanism behind IKv1.5 reduction. The hypothesis that Src regulates Kv1.5 activity by altering the density of mature membrane-localized channels was tested using whole-cell voltage clamp and Western blot analysis. We demonstrate that Src tonically inhibits Kv1.5 activity and decreases the density of mature membrane-localized channels. Kv1.5 channels possessing mutations within the Src binding motifs were also investigated and it was determined that each binding motif contributes to the Kv1.5-Src relationship, however, the binding of Src to an individual motif is sufficiently effective. Our findings indicate that Src regulates Kv1.5 through an interaction with the N-terminal binding motifs and suggests that the inhibition of forward trafficking may be involved in the underlying mechanism. (Supported by the Heart and Stroke foundation of Canada and The Canadian Institutes of Health Research).

Mitragynine, an euphoric compound inhibits hERG1a/1b channel current and upregulates the complexation of hERG1a-Hsp90 in HEK293-hERG1a/1b cells

Mitragyna speciosa Korth (M. speciosa) has been widely used as a recreational product, however, there are growing concerns on the abuse potentials and toxicity of the plant. Several poisoning and fatal cases involving kratom and mitragynine have been reported but the underlying causes remain unclear. The human ether-a-go-go-related gene 1 (hERG1) encodes the pore-forming subunit underlying cardiac rapidly delayed rectifier potassium current (IKr). Pharmacological blockade of the IKr can cause acquired long QT syndrome, leading to lethal cardiac arrhythmias. This study aims to elucidate the mechanisms of mitragynine-induced inhibition on hERG1a/1b current. Electrophysiology experiments were carried out using Port-a-Patch system. Quantitative RT-PCR, Western blot analysis, immunofluorescence and co-immunoprecipitation methods were used to determine the effects of mitragynine on hERG1a/1b expression and hERG1-cytosolic chaperones interaction. Mitragynine was found to inhibit the IKr current with an IC50 value of 332.70 nM. It causes a significant reduction of the fully-glycosylated (fg) hERG1a protein expression but upregulates both core-glycosylated (cg) expression and hERG1a-Hsp90 complexes, suggesting possible impaired hERG1a trafficking. In conclusion, mitragynine inhibits hERG1a/1b current through direct channel blockade at lower concentration, but at higher concentration, it upregulates the complexation of hERG1a-Hsp90 which may be inhibitory towards channel trafficking.

The molecular determinants of R-roscovitine block of hERG channels

Human ether-à-go-go-related gene (Kv11.1, or hERG) is a potassium channel that conducts the delayed rectifier potassium current (IKr) during the repolarization phase of cardiac action potentials. hERG channels have a larger pore than other K+channels and can trap many unintended drugs, often resulting in acquired LQTS (aLQTS). R-roscovitine, a cyclin-dependent kinase (CDK) inhibitor that also inhibits L-type calcium channels, inhibits open hERG channels but does not become trapped in the pore. Two-electrode voltage clamp recordings from Xenopus oocytes expressing wild-type (WT) or mutant (T623A, S624A, Y652A, F656A) hERG channels demonstrated that, compared to WT hERG, T623A, Y652A, and F656A inhibition by 200 μM R-roscovitine was ~ 48 %, 29 %, and 73 % weaker, respectively. In contrast, S624A hERG was inhibited more potently than WT hERG, with an ~ 34 % stronger inhibition. These findings were further supported by the IC50 values, which were increased for T623A, Y652A and F656A (by ~5.5, 2.75, and 42 fold respectively) and reduced 1.3 fold for the S624A mutant. Our data suggest that while T623, Y652, and F656 are critical for R-roscovitine-mediated inhibition, S624 may not be. This relatively unique feature, coupled with R-roscovitine’s tolerance in clinical trials, could guide future drug screens. We discuss our findings and how they lend support for the recent Comprehensive In Vitro Proarrhythmia Assay (CiPA) guidelines on the re-evaluation of potentially useful drugs that had failed testing due to unintended interactions with hERG.

Excavatolide-B Enhances Contextual Memory Retrieval via Repressing the Delayed Rectifier Potassium Current in the Hippocampus

Memory retrieval dysfunction is a symptom of schizophrenia, autism spectrum disorder (ASD), and absence epilepsy (AE), as well as an early sign of Alzheimer’s disease. To date, few drugs have been reported to enhance memory retrieval. Here, we found that a coral-derived natural product, excavatolide-B (Exc-B), enhances contextual memory retrieval in both wild-type and Cav3.2−/− mice via repressing the delayed rectifier potassium current, thus lowering the threshold for action potential initiation and enhancing induction of long-term potentiation (LTP). The human CACNA1H gene encodes a T-type calcium channel (Cav3.2), and its mutation is associated with schizophrenia, ASD, and AE, which are all characterized by abnormal memory function. Our previous publication demonstrated that Cav3.2−/− mice exhibit impaired contextual-associated memory retrieval, whilst their retrieval of spatial memory and auditory cued memory remain intact. The effect of Exc-B on enhancing the retrieval of context-associated memory provides a hope for novel drug development.