Action Potential Duration Research Articles

Exosomal mir-126-3p derived from endothelial cells induces ion channel dysfunction by targeting RGS3 signaling in cardiomyocytes: a novel mechanism in Takotsubo cardiomyopathy

BackgroundTakotsubo cardiomyopathy (TTC) is marked by an acute, transient, and reversible left ventricular systolic dysfunction triggered by stress, with endothelial dysfunction being one of its pathophysiological mechanisms. However, the precise molecular mechanism underlying the interaction between endothelial cells and cardiomyocytes during TTC remains unclear. This study reveals that exosomal miRNAs derived from endothelial cells exposed to catecholamine contribute to ion channel dysfunction in the setting of TTC.MethodsHuman-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) were treated with epinephrine (Epi) or exosomes (Exo) from Epi-treated human cardiac microvascular endothelial cells (HCMECs) or Exo derived from HCMECs transfected with miR-126-3p. The immunofluorescence staining, flow cytometry, qPCR, single-cell contraction, intracellular calcium transients, patch-clamp, dual luciferase reporter assay and western blot were performed for the study.ResultsModeling TTC with high doses of epinephrine (Epi) treatment in hiPSC-CMs shows suppression of depolarization velocity (Vmax), prolongation of action potential duration (APD), and induction of arrhythmic events. Exo derived from HCMECs treated with Epi (Epi-exo) mimicked or enhanced the effects of Epi. Epi exposure led to elevated levels of miR-126-3p in both HCMECs and their exosomes. Exo enriched with miR-126-3p demonstrated similar effects as Epi-exo, establishing the crucial role of miR-126-3p in the mechanism of Epi-exo. Dual luciferase reporter assay coupled with gene mutation techniques identified that miR-126-3p was found to target the regulator of G-protein signaling 3 (RGS3) gene. Western blot and qPCR analyses confirmed that miR-126-3p-mimic reduced RGS3 expression in both HCMECs and hiPSC-CMs, indicating miR-126-3p inhibits RGS3 signaling. Additionally, miR-126-3p levels were significantly higher in the serum of TTC patients compared to healthy controls and patients who had recovered from TTC.ConclusionsOur study is the first to reveal that exosomal miR-126-3p, originating from endothelial cells, contributes to ion channel dysfunction by regulating RGS3 signaling in cardiomyocytes. These findings provide new perspectives on the pathogenesis of TTC and suggest potential therapeutic targets for treatment.Graphical

Downmodulation of potassium conductances induces epileptic seizures in cortical network models via multiple synergistic factors.

Voltage-gated potassium conductances [Formula: see text] play a critical role not only in normal neural function, but also in many neurological disorders and related therapeutic interventions. In particular, in an important animal model of epileptic seizures, 4-aminopyridine (4-AP) administration is thought to induce seizures by reducing [Formula: see text] in cortex and other brain areas. Interestingly, 4-AP has also been useful in the treatment of neurological disorders such as multiple sclerosis (MS) and spinal cord injury, where it is thought to improve action potential propagation in axonal fibers. Here, we examined [Formula: see text] downmodulation in bio-physical models of cortical networks that included different neuron types organized in layers, potassium diffusion in interstitial and larger extracellular spaces, and glial buffering. Our findings are fourfold. First, [Formula: see text] downmodulation in pyramidal and fast-spiking inhibitory interneurons led to differential effects, making the latter much more likely to enter depolarization block. Second, both neuron types showed an increase in the duration and amplitude of action potentials, with more pronounced effects in pyramidal neurons. Third, a sufficiently strong [Formula: see text] reduction dramatically increased network synchrony, resulting in seizure like dynamics. Fourth, we hypothesized that broader action potentials were likely to not only improve their propagation, as in 4-AP therapeutic uses, but also to increase synaptic coupling. Notably, graded synapses incorporating this effect further amplified network synchronization and seizure-like dynamics. Overall, our findings elucidate different effects that [Formula: see text] downmodulation may have in cortical networks, explaining its potential role in both pathological neural dynamics and therapeutic applications.Significance Statement The modulation of voltage-gated potassium-conductances [Formula: see text] is thought to play an important role in epileptic seizures and therapeutic interventions in epilepsy, multiple sclerosis and spinal-cord injury. We show that [Formula: see text] downmodulation can lead to a cascade of effects including changes in basal excitability, broadening of action potentials resulting in enhanced robustness to synaptic noise perturbations and strengthening of synaptic coupling; and differential effects in excitatory and fast-spiking inhibitory interneurons, promoting depolarization block in the latter under high downmodulation. All these effects synergistically contribute to the emergence of seizure-like dynamics in the form of almost-periodic synchronized neuronal-population spiking in cortical networks. Under appropriate levels, [Formula: see text] downmodula tion can also have therapeutic effects by improving neuronal communication via the broadening of action potentials.

Downregulation of HDAC2 attenuates ventricular arrhythmias in a mouse model of cardiac hypertrophy through upregulation of KCHIP2 expression.

Decrease in repolarizing K+ currents, particularly the fast component of transient outward K+ current (Ito,f), prolongs action potential duration (APD) and predisposes the heart to ventricular arrhythmia during cardiac hypertrophy. Histone deacetylases (HDACs) have been suggested to participate in the development of cardiac hypertrophy, and class I HDAC inhibition has been found to attenuate pathological remodeling. This study investigated the potential therapeutic effects of HDAC2 on ventricular arrhythmia in pressure overload-induced cardiac hypertrophy. An in vivo cardiac hypertrophic model was produced by performing transverse aortic constriction (TAC) surgery, and an in vitro cardiomyocyte hypertrophy model by stimulating neonatal rat ventricular myocytes (NRVMs) with phenylephrine (PE). HDAC2 expression was upregulated in TAC mouse hearts and in PE-stimulated cardiomyocytes. Susceptibility to ventricular arrhythmia was increased in TAC mice, while Ito,f was decreased and APD was prolonged in TAC cardiomyocytes. Heart-specific knockdown of HDAC2 (HKD) by RNA interference increased Ito,f, shortened APD and decreased susceptibility to ventricular arrhythmia. Concomitantly, HKD increased the expression of the obligatory β subunit of Ito,f, KChIP2, which is downregulated in hypertrophic hearts. The effects of HKD on KChIP2 expression, Ito,f and APD were also observed in PE-stimulated cardiomyocytes. Mechanistically, HKD increased H3K4me3 abundance and H3K4me3 enrichment at the Kcnip2 promoter in cardiomyocytes. HKD also decreased the expression of KDM5, the H3K4me3 demethylase, which resulted in H3K4me3 upregulation. While investigating the regulatory mechanisms underlying the effect of HDAC2 on KDM5 stability, we identified CNOT4 as the active KDM5 ubiquitinase in cardiomyocytes. HKD increased CNOT4 expression and CNOT4-KDM5 interactions, and thus enhanced the polyubiquitinated degradation of KDM5. HDAC2 inhibition serves as a novel therapeutic strategy for preventing cardiac hypertrophy-associated electrophysiological remodeling. Furthermore, we identified a novel signaling pathway of CNOT4-mediated KDM5 degradation contributing to the upregulation of H3K4me3-mediated KChIP2 expression in response to HDAC2 inhibition.

Low-dose quinine targets KCNH6 to potentiate glucose-induced insulin secretion.

Insulin secretion is mainly regulated by two electrophysiological events, depolarization initiated by the closure of ATP-sensitive K+ (KATP) channels and repolarization mediated by K+ efflux. Quinine, a natural component commonly used for the treatment of malaria, has been reported to directly stimulate insulin release and lead to hypoglycemia in patients during treatment through inhibiting KATP channels. In this study, we verified the insulinotropic effect of quinine on the isolated mouse pancreatic islets. We also revealed that low-dose quinine (<20 µM) did not directly provoke Ca2+ spikes or insulin secretion under low-glucose conditions but potentiated Ca2+ influx and insulin secretion induced by high glucose, which cannot be explained by KATP inhibition. KCNH6 (hERG2) is a voltage-dependent K+ (Kv) channel that plays a critical role in the repolarization of pancreatic β cells. Patch clamp experiments showed that quinine inhibited hERG channels at low micromolar concentrations. However, whether quinine can target KCNH6 to potentiate glucose-induced insulin secretion remains unclear. Here, we showed that in vivo administration of low-dose quinine (25mg/kg) improved glucose tolerance and increased glucose-induced insulin release in wild-type control mice but not in Kcnh6-β-cell-specific knockout (βKO) mice. Consistently, in vitro treatment of primary islet β cells with low-dose quinine (10 µM) prolonged action potential duration and augmented glucose-induced Ca2+ influx in the wild-type control group but not in the Kcnh6-βKO group. Our results demonstrate that KCNH6 plays an important role in low-dose quinine-potentiated insulin secretion and provide new insights into KCNH6-targeted drug development.

Melatonin lessens the susceptibility to atrial fibrillation in sleep deprivation by ameliorating Ca2+ mishandling in response to mitochondrial oxidative stress.

The antiarrhythmic effect of melatonin(MLT) has been demonstrated in several studies; however, this hypothesis has recently been contested. Our research seeks to determine if exogenous MLT supplementation can reduce atrial fibrillation (AF) susceptibility due to sleep deprivation (SD) by addressing Ca2+ mishandling and atrial mitochondrial oxidative stress. Adult rats received daily MLT or vehicle injections and were exposed to a modified water tank. We evaluated MLT's impact on AF susceptibility by analyzing atrial electrical and structural changes, calcium handling, and oxidative stress markers. Techniques used included electrophysiological recording, echocardiography, optical mapping, histopathology, and molecular assays to understand MLT's protective effects against sleep deprivation-induced AF. Our findings indicate that MLT treatment effectively mitigates SD-induced AF, safeguards against atrial structural alterations, diminishes mitochondrial oxidative stress and normalizes calcium dynamics. Notably, MLT corrected calcium transient duration (CaD), action potential duration (APD), and conduction heterogeneity, shortened calcium transient refractoriness, and improved arrhythmogenic atrial alternans and spatially discordant alternans, thereby lowering the arrhythmogenic potential of the atria during sleep deprivation. In terms of mechanisms, MLT prevents SD-induced activation of ROS/CaMKII in atrial cardiomyocytes, reversing calcium transient refractoriness and inhibiting arrhythmogenic alternans. MLT significantly decreases the susceptibility to SD-induced AF by ameliorating mitochondrial oxidative stress and Ca2+ mishandling. These findings suggest a potential therapeutic application of MLT as an antiarrhythmic intervention for SD-related AF and underscore the need for further investigation, including clinical studies, to validate these mechanisms.

Indoxyl Sulfate Induces Ventricular Arrhythmias Attenuated by Secretoneurin in Right Ventricular Outflow Tract Cardiomyocytes.

Ventricular arrhythmias (VAs) are major causes of sudden cardiac death in chronic kidney disease (CKD) patients. Indoxyl sulfate (IS) is one common uremic toxin found in CKD patients. This study investigated whether IS could induce VAs via increasing right ventricular outflow tract (RVOT) arrhythmogenesis. Using conventional microelectrodes and whole-cell patch clamps, we studied the action potentials (APs) and ionic currents of isolated rabbit RVOT tissue preparations and single cardiomyocytes before and after IS (0.1 and 1.0μM). Calcium fluorescence imaging was performed in RVOT cardiomyocytes treated with and without IS (1.0μM) to evaluate the calcium transient and the calcium leak. In rabbit RVOT tissues, IS (0.1 and 1.0μM) attenuated the contractility and shortened the AP durations in a dose-dependent manner. In addition, IS (0.1 and 1.0μM) enhanced the pro-arrhythmia effects of isoproterenol (ISO, 1.0μM) and rapid ventricular pacing in RVOT (before versus after ISO, 25% versus 83%, N = 12). In RVOT cardiomyocytes, IS (1.0μM) significantly decreased the L-type calcium currents but increased the sodium-calcium exchanger and sodium window currents. Cardiomyocytes treated with IS (1.0μM) had lower calcium transients but higher diastolic calcium and calcium leak than those without IS treatment. Pretreatment with secretoneurin (SN, 30nM, a potent neuropeptide, suppressing CaMKII) or KN-93 (0.1μM, a CaMKII inhibitor) prevented IS-induced ionic current changes and arrhythmogenesis. In conclusion, IS modulates RVOT electrophysiology and arrhythmogenesis via enhanced CaMKII activity, which is attenuated by SN, leading to a novel therapeutic target for CKD arrhythmias.

Mechanistic Insights into Melatonin's Antiarrhythmic Effects in Acute Ischemia-Reperfusion-Injured Rabbit Hearts Undergoing Therapeutic Hypothermia.

The electrophysiological mechanisms underlying melatonin's actions and the electrophysiological consequences of superimposed therapeutic hypothermia (TH) in preventing cardiac ischemia-reperfusion (IR) injury-induced arrhythmias remain largely unknown. This study aimed to unveil these issues using acute IR-injured hearts. Rabbits were divided into heart failure (HF), HF+melatonin, control, and control+melatonin groups. HF was induced by rapid right ventricular pacing. Melatonin was administered orally (10 mg/kg/day) for four weeks, and IR was created by 60-min coronary artery ligation and 30-min reperfusion. The hearts were then excised and Langendorff-perfused for optical mapping studies at normothermia, followed by TH. Melatonin significantly reduced ventricular fibrillation (VF) maintenance. In failing hearts, melatonin reduced the spatially discordant alternans (SDA) inducibility mainly by modulating intracellular Ca2+ dynamics via upregulation of sarcoplasmic reticulum Ca2+-ATPase (SERCA2a) and calsequestrin 2 and attenuating the downregulation of phosphorylated phospholamban protein expression. In control hearts, melatonin improved conduction slowing and reduced dispersion of action potential duration (APDdispersion) by upregulating phosphorylated connexin 43, attenuating the downregulation of SERCA2a and phosphorylated phospholamban and attenuating the upregulation of phosphorylated Ca2+/calmodulin-dependent protein kinase II. TH significantly retarded intracellular Ca2+ decay slowed conduction, and increased APDdispersion, thereby facilitating SDA induction, which counteracted the beneficial effects of melatonin in reducing VF maintenance.

Dynamical effects of mechano-chemo-transduction on cardiac alternans.

In every heartbeat, cardiac muscle cells perform excitation-Ca2+ signaling-contraction (EC) coupling to pump blood against the vascular resistance. Cardiomyocytes can sense the mechanical load and activate mechano-chemo-transduction (MCT) mechanism, which provides feedback regulation of EC coupling. MCT feedback is important for the heart to upregulate contraction in response to increased load to maintain cardiac output. MCT feedback enhances the L-type Ca2+ current, sensitizes ryanodine receptors (RyRs), and augments SERCA pump activity, thereby maintaining contraction amplitude despite increased load. However, under certain conditions, MCT feedback can also promote cardiac alternans, seen as beat-to-beat variations in action potential duration, Ca2+ transients, and contraction strength, which is a precursor to arrhythmias. While alternans can arise from instabilities in either membrane voltage or intracellular Ca2+ cycling, underlying mechanisms of MCT-induced alternans, particularly electromechanically discordant alternans where stronger beats are paradoxically associated with shorter action potentials, remain unclear. In this study, we used a mathematical model of the ventricular myocyte to investigate the effects of MCT feedback on the dynamical system that generates alternans. We systematically analyzed how MCT feedback, acting through L-type Ca2+ channels (LTCCs), RyRs, or SERCA, affects the stability of membrane voltage and Ca2+ cycling, as well as the coupling between them. Our results show that MCT feedback can generally promote both concordant and discordant alternans in action potential and Ca2+ transients, depending on the underlying instability mechanism. We found that MCT feedback through RyRs predominantly increases Ca2+ instability, while LTCC and SERCA feedback have complex effects due to the interplay between stability and coupling alterations. We also showed how to determine underlying mechanisms from experimental and clinical observations. Our modeling studies provide new insights into the complex dynamics underlying cardiac alternans and highlight the importance of MCT feedback in the development of life-threatening arrhythmias in the heart under mechanical load.

Evaluation of electrophysiological changes before and after pramipexole treatment in patients diagnosed with idiopathic restless legs syndrome

Background and purpose – Restless legs syndrome (RLS) is a neurological disorder characterized by an irresistible urge to move the legs, frequently accompanied by unpleasant sensations. In this study, we aimed to determine the changes in electrophysiological tests including Fwave responses and H-reflex parameters in patients with idiopathic RLS after pramipexole medication. Methods –This prospective study was conducted with twenty newly diagnosed, idiopathic RLS patients. The electrophysiological parameters, specifically F-wave duration (FWD), the duration of related compound muscle action potential (CMAPD), and FWD/ CMAPD ratio, the ratio of maximum H-reflex amplitude(H) to maximum M-response amplitude (M)(H/M) were recorded in patients with idiopathic RLS before and after pramipexole medication. Results – The results revealed significant improvements in specific electrophysiolo gical parameters after pramipexole use. We observed a significant decrease in FWD, CMAPD, and FWD/CMAPD values after pramipexole treatment suggesting a reduction in motor neuron excitability. We also obser ved a significant decrease in H/M ratio after pramipexole medication indicating reduced spinal motor neuron excitability. Conclusion – This study demonstrated significant improvement in specific electrophysiological parameters following pramipexole treatment in patients with idiopathic RLS. The findings provide better understanding of the effects of pramipexole on nerve function and may contribute to the development of a monitoring tool and lead to more targeted treatments and interventions for RLS, improving the overall management and care for patients’ outcomes and quality of life with this debilitating condition.

Shank3 Overexpression Leads to Cardiac Dysfunction in Mice by Disrupting Calcium Homeostasis in Cardiomyocytes.

SH3 and multiple ankyrin repeat domains 3 (Shank3) proteins play crucial roles as neuronal postsynaptic scaffolds. Alongside neuropsychiatric symptoms, individuals with SHANK3 mutations often exhibit symptoms related to dysfunctions in other organs, including the heart. However, detailed insights into the cardiac functions of Shank3 remain limited. This study aimed to characterize the cardiac phenotypes of Shank3-overexpressing transgenic mice and explore the underlying mechanisms. Cardiac histological analysis, electrocardiogram and echocardiogram recordings were conducted on Shank3-overexpressing transgenic mice. Electrophysiological properties, including action potentials and L-type Ca²⁺ channel (LTCC) currents, were measured in isolated cardiomyocytes. Ca²⁺ homeostasis was assessed by analyzing cytosolic Ca²⁺ transients and sarcoplasmic reticulum Ca²⁺ contents. Depolarization-induced cell shortening was examined in cardiomyocytes. Immunoprecipitation followed by mass spectrometry-based identification was employed to identify proteins in the cardiac Shank3 interactome. Western blot and immunocytochemical analyses were conducted to identify changes in protein expression in Shank3-overexpressing transgenic cardiomyocytes. The hearts of Shank3-overexpressing transgenic mice displayed reduced weight and increased fibrosis. In vivo, sudden cardiac death, arrhythmia, and contractility impairments were identified. Shank3-overexpressing transgenic cardiomyocytes showed prolonged action potential duration and increased LTCC current density. Cytosolic Ca²⁺ transients were increased with prolonged decay time, while sarcoplasmic reticulum Ca²⁺ contents remained normal. Cell shortening was augmented in Shank3-overexpressing transgenic cardiomyocytes. The cardiac Shank3 interactome comprised 78 proteins with various functions. Troponin I levels were down-regulated in Shank3-overexpressing transgenic cardiomyocytes. This study revealed cardiac dysfunction in Shank3-overexpressing transgenic mice, potentially attributed to changes in Ca²⁺ homeostasis and contraction, with a notable reduction in troponin I.

Artemisinin-Quinidine Combination for Suppressing Ventricular Tachyarrhythmia in an Ex Vivo Model of Brugada Syndrome.

The ionic mechanism underlying Brugada syndrome (BrS) arises from an imbalance in transient outward current flow between the epicardium and endocardium. Previous studies report that artemisinin, originally derived from a Chinese herb for antimalarial use, inhibits the Ito current in canines. In a prior study, we showed the antiarrhythmic effects of artemisinin in BrS wedge preparation models. However, quinidine remains a well-established antiarrhythmic agent for treating BrS. Therefore, this study aims to investigate the efficacy of combining artemisinin with low-dose quinidine in suppressing ventricular tachyarrhythmia (VTA) in experimental canine BrS models. Transmural pseudo-electrocardiogram and epicardial/endocardial action potential (AP) were recorded from coronary-perfused canine right ventricular wedge preparation. To mimic the BrS model, acetylcholine (3 μM), calcium channel blocker verapamil (1 μM), and Ito agonist NS5806 (6-10 μM) were administered until VTA was induced. Subsequently, low-dose quinidine (1-2 μM) combined with artemisinin (100 μM) was perfused to mitigate VTA. Key parameters, including AP duration, J wave area, notch index, and T wave dispersion, were measured. After administering the provocation agents, all sample models exhibited prominent J waves and VTA. Artemisinin alone (100-150 μM) suppressed VTA and restored the AP dome in all three preparations. Its infusion resulted in reductions in the J wave area and epicardial notch index. Consequently, low-dose quinidine (1-2 μM) did not fully restore the AP dome in all six sample models. However, when combined with additional artemisinin (100 μM), low-dose quinidine effectively suppressed VTA in all six models and restored the AP dome while also decreasing the J wave area and epicardial notch index. Low-dose quinidine alone fails to fully alleviate VTA in the BrS wedge model. However, its combination with artemisinin effectively suppresses VTA. Artemisinin may reduce the therapeutic dose of quinidine, potentially minimizing its associated adverse effects.

K+ currents in ventricular cardiomyocytes of p.N98S-calmodulin mutant mice.

Missense mutations in calmodulin (CaM)-encoding genes are associated with life-threatening ventricular arrhythmia syndromes. Here, we investigated a role of cardiac K+ channel dysregulation in arrhythmogenic long QT syndrome (LQTS) using a knock-in mouse model heterozygous for a recurrent mutation (p.N98S) in the Calm1 gene (Calm1N98S/+). Single-cell patch-clamp technique and whole-heart optical voltage mapping were used to assess action potentials and whole-cell currents. Ventricular action potential duration (APD) at baseline was similar between genotypes. The β-adrenergic agonist isoproterenol prolonged APD in myocytes and isolated perfused hearts from Calm1N98S/+, but not wild-type (Calm1+/+), mice. Current density-voltage relationships for the small-conductance calcium-activated K+ (SK) current and the inward rectifier K+ current did not significantly differ between Calm1+/+ and Calm1N98S/+ ventricular cardiomyocytes ± isoproterenol. Peak densities of other voltage-gated K+ currents were significantly larger in Calm1N98S/+ versus Calm1+/+ cells at voltages ≥40 mV both without and with isoproterenol. Isoproterenol reduced outward KATP currents more in Calm1N98S/+ versus Calm1+/+ myocytes. Dialysis of Calm1+/+ cardiomyocytes with exogenous wild-type or N98S-CaM protein (5 μmol/l) via the pipette respectively increased and eliminated SK currents. The specific SK channel inhibitor apamin did not significantly alter APD of Calm1+/+ or Calm1N98S/+ hearts ± isoproterenol. Thus, dysregulation of SK or voltage-gated K+ channels does not contribute to the β-adrenergic-induced LQTS of Calm1N98S/+ mice, possibly because cardiomyocyte content of endogenous N98S-CaM and/or its affinity for CaM binding domains may be too low to modulate channel properties. The larger KATP current inhibition by isoproterenol may delay Calm1N98S/+ myocyte repolarization at low intracellular [ATP].

Sequence-Dependent Repolarization Is Modulated by Endogenous Action Potential Duration Gradients Rather Than Electrical Coupling in Ventricular Myocardium.

Previous studies suggest the relationship between activation time (AT) and action potential duration (APD) in the heart is dependent on electrotonic coupling, but this has not been directly tested. This study assessed whether acute changes in electrical coupling, or other determinants of conduction or repolarization, modulate APD heterogeneity. Langendorff-perfused guinea pig hearts were epicardially paced and optically mapped after treatment with the gap junction uncoupler carbenoxolone, ephaptic uncoupler mannitol, ephaptic enhancer dextran 2MDa, sodium channel inhibitor flecainide, or rapid component of the delayed rectifier potassium channel inhibitor E4031. SDof APD and the AT-APD slope and coefficient of determination were quantified as metrics of APD heterogeneity. SD of APD increased with carbenoxolone, mannitol, and altered activation sequence. The AT-APD slope was insensitive to carbenoxolone, mannitol, dextran, flecainide, or E4031 but changed in response to activation sequence. The coefficient of determination did not change with carbenoxolone; decreased with mannitol, E4031, and activation sequence; but increased with dextran and flecainide. APD heterogeneity changes were dependent on whether the estimation used SD of APD or the AT-APD relationship. The pacing stimulus increased APD at the site of stimulation, revealing a confounding stimulus effect on APD within the measurement area. Simulations predict that the stimulus artifact and endogenous APD gradients are stronger determinants of APD heterogeneity than AT. APD dependence on conduction is relatively small. Furthermore, APD heterogeneity within a mapping field of view is dependent on endogenous gradients, the stimulus artifact, and the experimental approach, rather than electrical coupling.

Non-invasive Assessment of HydroQUInidine EffecT in Brugada Syndrome (QUIET BrS).

Hydroquinidine reduces arrhythmic events in patients with Brugada syndrome (BrS). The mechanism by which it exerts antiarrhythmic benefit and its electrophysiological effects on BrS substrate remain incompletely understood. This study aimed to determine the effect of Hydroquinidine on ventricular depolarisation and repolarisation in patients with BrS in vivo. Twelve patients with BrS underwent electrocardiogram (standard, high-lead and signal averaged) and electrocardiographic imaging (ECGi) at baseline and "on-treatment" with Hydroquinidine 300mg twice daily. ST-segment elevation, activation time (AT), repolarisation time (RT) and activation-recovery intervals (ARI) were computed for the ventricles and the right ventricular outflow tract (RVOT). Serum Hydroquinidine levels were determined, and adverse drug events captured through medication survey. Hydroquinidine increased RT (301.1 ± 24.1ms vs 348.8 ± 28.3ms, p <0.001), repolarisation gradients (1.1 ± 0.4 ms/mm vs 1.6 ± 0.4 ms/mm, p<0.001) and ARI (241.3 ± 18.1 ms vs 284.8 ± 21.5 ms, p<0.001) in the RVOT, with a greater change in the RVOT compared to the rest of the ventricles. In contrast, activation parameters did not change significantly on-treatment with Hydroquinidine, although there was a subtle increase in ST-segment elevation over the RVOT (1.5 ± 0.7mV vs 1.8 ± 0.8mV; P<0.001). Hydroquinidine levels did not correlate with electrophysiological changes or occurrence of adverse drug reactions. One patient developed frequent non-sustained ventricular tachycardia on-treatment with Hydroquinidine. Hydroquinidine primarily effects ventricular repolarisation and action potential duration (indicated by ARI) in patients with BrS and demonstrates regional variation with more significant changes in the RVOT.

Modeling Fibroblast-Cardiomyocyte Interactions: Unveiling the Role of Ion Currents in Action Potential Modulation.

Fibrotic cardiomyopathy represents a significant pathological condition characterized by the interaction between cardiomyocytes and fibroblasts in the heart, and it currently lacks an effective cure. In vitro platforms, such as engineered heart tissue (EHT) developed through the co-culturing of cardiomyocytes and fibroblasts, are under investigation to elucidate and manipulate these cellular interactions. We present the first integration of mathematical electrophysiological models that encapsulate fibroblast-cardiomyocyte interactions with experimental EHT studies to identify and modulate the ion channels governing these dynamics. Our findings resolve a long-standing debate regarding the effect of fibroblast coupling on cardiomyocyte action potential duration (APD). We demonstrate that these seemingly contradictory outcomes are contingent upon the specific properties of the cardiomyocyte to which the fibroblast is coupled, particularly the relative magnitudes of the fast Na+ and transient outward K+ currents within the cardiomyocyte. Our results emphasize the critical importance of detailed ionic current representation in cardiomyocytes for accurately predicting the interactions between cardiomyocytes and fibroblasts in EHT. Surprisingly, complex ion channel-based models of fibroblast electrophysiology did not outperform simplified resistance-capacitance models in this analysis. Collectively, our findings highlight the promising potential of synergizing in vitro and in silico approaches to identify therapeutic targets for cardiomyopathies.

Alleviating the Effects of Short QT Syndrome Type 3 by Allele-Specific Suppression of the KCNJ2 Mutant Allele.

Short QT syndrome type 3 (SQTS3 or SQT3), which is associated with life-threatening cardiac arrhythmias, is caused by heterozygous gain-of-function mutations in the KCNJ2 gene. This gene encodes the pore-forming α-subunit of the ion channel that carries the cardiac inward rectifier potassium current (IK1). These gain-of-function mutations either increase the amplitude of IK1 or attenuate its rectification. The aim of the present in silico study is to test to which extent allele-specific suppression of the KCNJ2 mutant allele can alleviate the effects of SQT3, as recently demonstrated in in vitro studies on specific heterozygous mutations associated with long QT syndrome type 1 and 2 and short QT syndrome type 1. To this end, simulations were carried out with the two most recent comprehensive models of a single human ventricular cardiomyocyte. These simulations showed that suppression of the mutant allele can, at least partially, counteract the effects of the mutation on IK1 and restore the action potential duration for each of the four SQT3 mutations that are known by now. We conclude that allele-specific suppression of the KCNJ2 mutant allele is a promising technique in the treatment of SQT3 that should be evaluated in in vitro and in vivo studies.

The properties of laryngeal electromyography in the non-paralyzed sides of patients with idiopathic vocal cord paralysis.

To evaluate the injuries of the recurrent laryngeal nerve (RLN), superior laryngeal nerve (SLN), and their innervated laryngeal muscles on the non-paralyzed sides in patients with idiopathic vocal cord paralysis (IVCP). Eighty-four cases of patients with IVCP were evaluated using stroboscopic laryngoscopy, voice analysis, and laryngeal electromyography(LEMG). Concurrently, twenty-eight cases involving healthy volunteers without vocal cord paralysis were enrolled and examined using LEMG during the same period. Subsequently, comparisons of LEMG results were conducted between the paralyzed and non-paralyzed sides, and among the non-paralyzed sides.Furthermore, a comparison was conducted between the LEMG results of the non-paralyzed sides in patients with IVCP and the corresponding ipsilateral sides in healthy volunteers.These comparisons were stratified based on the side of paralysis and the duration of the disease course (less than 3 months versus more than 3 months) for IVCP patients. Initially, the amplitude of the RLN, the action potential durations (APDs) and amplitude of the thyroarytenoid muscle (TM), and the amplitude of the posterior cricoarytenoid muscle (PCM) on the paralyzed side were significantly lower than those on the non-paralyzed side. Conversely, the latency of the RLN on the paralyzed side was significantly longer compared to the non-paralyzed side (P < 0.05). However, there were no significant differences in the latency and amplitude of the SLN, nor in the APD and amplitude of the CM between the paralyzed and non-paralyzed sides (P > 0.05). Furthermore, no significant differences were observed in the LEMG results of the non-paralyzed sides in IVCP patients, irrespective of the paralyzed side or the duration of the disease course (P > 0.05), and no significant differences of the LEMG results were observed when comparing the non-paralyzed sides of patients with IVCP to the ipsilateral sides of healthy volunteers (P > 0.05). However, when the disease course is less than 3 months, the amplitudes of the SLN and CM in the non-paralyzed side (right side) of left-sided IVCP patients are significantly lower than those in the non-paralyzed side (left side) of right-sided IVCP patients (SLN, 6.23 ± 4.42 mv vs. 10.21 ± 7.56 mv, t=-2.296, P = 0.028; CM, 0.49 ± 0.17 mv vs. 0.60 ± 0.19 mv, t=-2.207, P = 0.032), of which the amplitude of the CM is significantly reduced compared to the right side of healthy controls (0.49 ± 0.17 mv vs. 0.61 ± 0.21 mv, t=-2.423, P = 0.019), indicating that the SLN and CM on the non-paralyzed side are vulnerable to damage. Moreover, in patients with right-sided IVCP persisting for more than 3 months, there was a significant reduction in the amplitudes of the SLN and RLN, as well as the TM and PCM on the non-paralyzed side (left side), compared to the corresponding ipsilateral sides in healthy volunteers (P < 0.05). This observation indicates that the LEMG results for the non-paralyzed side in IVCP patients are not entirely normal, particularly in cases of right-sided IVCP with a disease duration exceeding three months. In patients with IVCP, it is imperative to conduct a simultaneous analysis of LEMG results from both the paralyzed and non-paralyzed sides after determining the affected side and the progression of the paralysis. This thorough examination is crucial for accurately evaluating the condition and prognosis of individuals with IVCP.

PDE8 Inhibition and Its Impact on ICa,L in Persistent Atrial Fibrillation: Evaluation of PDE8 as a Potential Drug Target.

Atrial fibrillation (AF) poses a significant therapeutic challenge with drug interventions showing only limited success. Phosphodiesterases (PDE) regulate cardiac electrical stability and may represent an interesting target. Recently, PDE8 inhibition was proposed as an antiarrhythmic intervention by increasing L-type Ca 2+ current (I Ca,L ) and action potential duration (APD). However, the effect size of PDE8 inhibition on I Ca,L and APD seems discrepant and effects on force are unknown. We investigated the impact of PDE8 inhibition on force using PF-04957325 in right atrial appendages, obtained from patients in sinus rhythm (SR) and with persistent AF (peAF) undergoing cardiac surgery. A computational model was used to predict the effects of PDE8 inhibition on APD in SR and peAF. Results showed no increase in force after exposure to increasing concentrations of the PDE8 inhibitor PF-04957325 in either SR or peAF tissues. Furthermore, PDE8 inhibition did not affect the potency or efficacy of norepinephrine-induced inotropic effects in either group. Arrhythmic events triggered by norepinephrine were observed in both SR and peAF, but their frequency remained unaffected by PF-04957325 treatment. Computational modeling predicted that the reported increase in I Ca,L induced by PDE8 inhibition would lead to substantial APD prolongation at all repolarization states, particularly in peAF. Our findings indicate that PDE8 inhibition does not significantly impact force or arrhythmogenicity in human atrial tissue.

A simulation study of the impact of drug-IKr binding mechanisms on biomarkers of proarrhythmic risk reveals a crucial role in reverse use-dependence of action potential duration and a marked influence on the vulnerable window.

In silico human models are being used more and more to predict the potential proarrhythmic risk of compounds. It has been shown that incorporation of the dynamics of drug-hERG channel interactions can have an important impact on the action potential duration (APD) at normal heart rates. Our aim is to investigate the relevance of drug dynamics on other important biomarkers of proarrhythmic risk. We use the state-of-the-art mathematical models of the cardiac electrophysiological activity to simulate TRIaD biomarkers, namely Triangulation, Reverse use-dependency, electrical Instability of the action potential and Dispersion, together with the vulnerable window to unidirectional block. They were simulated in control conditions and in the presence of an extensive set of 114 in silico IKr blockers with different kinetics and affinities to conformational states of the channel and 10 well-known real IKr blockers at the concentration leading to a 25 % prolongation of the APD. Our results show that drug binding dynamics to hERG are crucial for the reverse use-dependence of APD, the slope of the APD restitution curve as a function of the root square of the cycle length ranging from 0 to 5.6 ms/ms (2.1 ms/ms in control conditions). The vulnerable window for unidirectional block and the transmural action potential duration dispersion markedly depended on the drug binding mechanisms and kinetics, although to a lesser extent. Virtual drugs led to increments of these two biomarkers from 25 % to 200 %. On the contrary, temporal instability and, beat-to-beat instability, are less dependent on the dynamics of drug binding. The results obtained with the models of real IKr blockers are in line with those obtained with the virtual drugs. Our study highlights the importance of considering the drug binding mechanism, as well as the kinetics, to assess the effects of IKr blockers. Moreover, adoption of in silico models mimicking these characteristics would contribute to the improvement of the prediction of the proarrhythmic risk of new compounds.

Aminophylline at clinically relevant concentrations affects inward rectifier potassium current in healthy porcine and failing human cardiomyocytes in a similar manner

Aminophylline, a bronchodilator mainly used to treat severe asthma attacks, may induce arrhythmias. Unfortunately, the underlying mechanism is not well understood. We have recently described a significant, on average inhibitory effect of aminophylline on inward rectifier potassium current IK1, known to substantially contribute to arrhythmogenesis, in rat ventricular myocytes at room temperature. This study was aimed to examine whether a similar effect may be observed under clinically relevant conditions. Experiments were performed using the whole cell patch clamp technique at 37°C on enzymatically isolated healthy porcine and failing human ventricular myocytes. The effect of clinically relevant concentrations of aminophylline (10-100µM) on IK1 did not significantly differ in healthy porcine and failing human ventricular myocytes. IK1 was reversibly inhibited by ~20 and 30% in the presence of 30 and 100µM aminophylline, respectively, at -110 mV; an analogical effect was observed at -50 mV. To separate the impact of IK1 changes on AP configuration, potentially interfering ionic currents were blocked (L-type calcium and delayed rectifier potassium currents). A significant prolongation of AP duration was observed in the presence of 100µM aminophylline in porcine cardiomyocytes which well agreed with the effect of a specific IK1 inhibitor Ba2+ (10µM) and with the result of simulations using a porcine ventricular cell model. We conclude that the observed effect of aminophylline on healthy porcine and failing human IK1 might be involved in its proarrhythmic action. To fully understand the underlying mechanism, potential aminophylline impact on other ionic currents should be explored.