Beat Rate Research Articles

Hyperglycaemia-induced impairment of the autorhythmicity and gap junction activity of mouse embryonic stem cell-derived cardiomyocyte-like cells.

Diabetes mellitus with hyperglycaemia is a major risk factor for malignant cardiac dysrhythmias. However, the underlying mechanisms remain unclear, especially during the embryonic developmental phase of the heart. This study investigated the effect of hyperglycaemia on the pulsatile activity of stem cell-derived cardiomyocytes. Mouse embryonic stem cells (mESCs) were differentiated into cardiac-like cells through embryoid body (EB) formation, in either baseline glucose or high glucose conditions. Action potentials (APs) were recorded using a voltage-sensitive fluorescent dye and gap junction activity was evaluated using scrape-loading lucifer yellow dye transfer assay. Molecular components were detected using immunocytochemistry and immunoblot analyses. High glucose decreased the spontaneous beating rate of EBs and shortened the duration of onset of quinidine-induced asystole. Furthermore, it altered AP amplitude, but not AP duration, and had no impact on neither the expression of the hyperpolarisation-activated cyclic nucleotide-gated isoform 4 (HCN4) channel nor on the EB beating rate response to ivabradine nor isoprenaline. High glucose also decreased both the intercellular spread of lucifer yellow within an EB and the expression of the cardiac gap junction protein connexin43 as well as upregulated the expression of transforming growth factor beta1 (TGF-β1) and phosphorylated Smad3. High glucose suppressed the autorhythmicity and gap junction conduction of mESC-derived cardiomyocytes, via mechanisms probably involving TGF-β1/Smad3 signalling. The results allude to glucotoxicity related proarrhythmic effects, with potential clinical implications in foetal diabetic cardiac disease.

Potential for cardiac toxicity with methylimidazolium ionic liquids

Methylimidazolium ionic liquids (MILs) are solvent chemicals used in industry. Recent work suggests that MILs are beginning to contaminate the environment and lead to exposure in the general population. In this study, the potential for MILs to cause cardiac toxicity has been examined. The effects of 5 chloride MIL salts possessing increasing alkyl chain lengths (2 C, EMI; 4 C, BMI; 6 C; HMI, 8 C, M8OI; 10 C, DMI) on rat neonatal cardiomyocyte beat rate, beat amplitude and cell survival were initially examined. Increasing alkyl chain length resulted in increasing adverse effects, with effects seen at 10−5 M at all endpoints with M8OI and DMI, the lowest concentration tested. A limited sub-acute toxicity study in rats identified potential cardiotoxic effects with longer chain MILs (HMI, M8OI and DMI) based on clinical chemistry. A 5 month oral/drinking water study with these MILs confirmed cardiotoxicity based on histopathology and clinical chemistry endpoints. Since previous studies in mice did not identify the heart as a target organ, the likely cause of the species difference was investigated. qRT-PCR and Western blotting identified a marked higher expression of p-glycoprotein-3 (also known as ABCB4 or MDR2) and the breast cancer related protein transporter BCRP (also known as ABCG2) in mouse, compared to rat heart. Addition of the BCRP inhibitor Ko143 – but not the p-glycoproteins inhibitor cyclosporin A - increased mouse cardiomyocyte HL-1 cell sensitivity to longer chain MILs to a limited extent. MILs therefore have a potential for cardiotoxicity in rats. Mice may be less sensitive to cardiotoxicity from MILs due in part, to increased excretion via higher levels of cardiac BCRP expression and/or function. MILs alone, therefore may represent a hazard in man in the future, particularly if use levels increase. The impact that MILs exposure has on sensitivity to cardiotoxic drugs, heart disease and other chronic diseases is unknown.

Action potential variability in human pluripotent stem cell-derived cardiomyocytes obtained from healthy donors.

Human pluripotent stem cells (PSC) have been used for disease modelling, after differentiation into the desired cell type. Electrophysiologic properties of cardiomyocytes derived from pluripotent stem cells are extensively used to model cardiac arrhythmias, in cardiomyopathies and channelopathies. This requires strict control of the multiple variables that can influence the electrical properties of these cells. In this article, we report the action potential variability of 780 cardiomyocytes derived from pluripotent stem cells obtained from six healthy donors. We analyze the overall distribution of action potential (AP) data, the distribution of action potential data per cell line, per differentiation protocol and batch. This analysis indicates that even using the same cell line and differentiation protocol, the differentiation batch still affects the results. This variability has important implications in modeling arrhythmias and imputing pathogenicity to variants encountered in patients with arrhythmic diseases. We conclude that even when using isogenic cell lines to ascertain pathogenicity to variants associated to arrythmias one should use cardiomyocytes derived from pluripotent stem cells using the same differentiation protocol and batch and pace the cells or use only cells that have very similar spontaneous beat rates. Otherwise, one may find phenotypic variability that is not attributable to pathogenic variants.

Levosimendan increases the phosphorylation state of phospholamban in the isolated human atrium

Levosimendan (up to 10 µM) given alone failed to increase force of contraction in isolated electrically stimulated (1 Hz) left atrial (LA) preparations from wild-type mice. Only in the additional presence of 0.1 µM rolipram, an inhibitor of the activity of phosphodiesterase IV, levosimendan increased force of contraction in LA and increased the phosphorylation state of phospholamban at amino acid serine 16. Levosimendan alone increased the beating rate in isolated spontaneously beating right atrial preparations from mice and this effect was potentiated by rolipram. The positive inotropic and the positive chronotropic effects of levosimendan in mouse atrial preparations were attenuated by 10 µM propranolol. Finally, we studied the contractile effects of levosimendan in isolated electrically stimulated (1 Hz) right atrial preparations from the human atrium (HAP), obtained during cardiac surgery. We detected concentration-dependent positive inotropic effects of levosimendan alone that reached plateau at 1 µM levosimendan in HAP (n = 11). Levosimendan shortened time of tension relaxation in HAP. Cilostamide (1 µM), an inhibitor of phosphodiesterase III, or propranolol (10 µM) blocked the positive inotropic effect of levosimendan in HAP. Levosimendan (1 µM) alone increased in HAP the phosphorylation state of phospholamban. In conclusion, we present evidence that levosimendan acts via phosphodiesterase III inhibition in the human atrium leading to phospholamban phosphorylation and thus explaining the positive inotropic effects of levosimendan in HAP.

Cellular Biomechanic Impairment in Cardiomyocytes Carrying the Progeria Mutation: An Atomic Force Microscopy Investigation.

Given the clinical effect of progeria syndrome, understanding the cell mechanical behavior of this pathology could benefit the patient's treatment. Progeria patients show a point mutation in the lamin A/C gene (LMNA), which could change the cell's biomechanical properties. This paper reports a mechano-dynamic analysis of a progeria mutation (c.1824 C > T, p.Gly608Gly) in neonatal rat ventricular myocytes (NRVMs) using cell indentation by atomic force microscopy to measure alterations in beating force, frequency, and contractile amplitude of selected cells within cell clusters. Furthermore, we examined the beating rate variability using a time-domain method that produces a Poincaré plot because beat-to-beat changes can shed light on the causes of arrhythmias. Our data have been further related to our cell phenotype findings, using immunofluorescence and calcium transient analysis, showing that mutant NRVMs display changes in both beating force and frequency. These changes were associated with a decreased gap junction localization (Connexin 43) in the mutant NRVMs even in the presence of a stable cytoskeletal structure (microtubules and actin filaments) when compared with controls (wild type and non-treated cells). These data emphasize the kindred between nucleoskeleton (LMNA), cytoskeleton, and the sarcolemmal structures in NRVM with the progeria Gly608Gly mutation, prompting future mechanistic and therapeutic investigations.

Effects of Adrenomedullin on Atrial Electrophysiology and Pulmonary Vein Arrhythmogenesis.

Adrenomedullin, a peptide with vasodilatory, natriuretic, and diuretic effects, may be a novel agent for treating heart failure. Heart failure is associated with an increased risk of atrial fibrillation (AF), but the effects of adrenomedullin on atrial arrhythmogenesis remain unclear. This study investigated whether adrenomedullin modulates the electrophysiology of the atria (AF substrate) or pulmonary vein (PV; AF trigger) arrhythmogenesis. Conventional microelectrode or whole-cell patch clamps were used to study the effects of adrenomedullin (10, 30, and 100 pg/mL) on the electrical activity, mechanical response, and ionic currents of isolated rabbit PV and sinoatrial node tissue preparations and single PV cardiomyocytes. At 30 and 100 pg/mL, adrenomedullin significantly reduced the spontaneous beating rate of the PVs from 2.0 ± 0.4 to 1.3 ± 0.5 and 1.1 ± 0.5 Hz (reductions of 32.9% ± 7.1% and 44.9 ± 8.4%), respectively, and reduced PV diastolic tension by 12.8% ± 4.1% and 14.5% ± 4.1%, respectively. By contrast, adrenomedullin did not affect sinoatrial node beating. In the presence of L-NAME (a nitric oxide synthesis inhibitor, 100 μM), adrenomedullin (30 pg/mL) did not affect the spontaneous beating rate or diastolic tension of the PVs. In the single-cell experiments, adrenomedullin (30 pg/mL) significantly reduced the L-type calcium current (ICa-L) and reverse-mode current of the sodium-calcium exchanger (NCX). Adrenomedullin reduces spontaneous PV activity and PV diastolic tension by reducing ICa-L and NCX current and thus may be useful for treating atrial tachyarrhythmia.

Polystyrene microplastics weaken the predator-induced defenses of Daphnia magna: Evidences from the changes in morphology and behavior

Polystyrene microplastics are ubiquitous in freshwater ecosystems and have significant impacts on freshwater organisms. Stable interspecific relationships, including the predation and defense relationships between predator and prey, play an extremely important role in maintaining the health of aquatic ecosystems. However, it still remains unknown whether polystyrene microplastics can interfere with predator-induced defenses of prey, especially in behavior change. Therefore, we studied the effects of different particle sizes and concentrations of polystyrene microplastics on the induced defenses related to morphology, reproduction, and behavior of Daphnia magna exposed to the predation risks from a species of zooplanktivorous fish Rhodeus ocellatus. Results showed that polystyrene microplastics weakened the predator-induced defenses of D. magna, including morphology, reproduction, and behavior. Polystyrene microplastics did not affect the beat rate of the second antennae (swimming activity) and thoracic appendage (filter-feeding activity) of D. magna, but R. ocellatus kairomone reduced Daphnia swimming activity. Moreover, in the absence of R. ocellatus predation risks, polystyrene microplastics did not alter the vertical distribution of D. magna in the water column, whereas D. magna exposed to R. ocellatus kairomone stayed deeper in the water; however, when both polystyrene microplastics and fish kairomone were present, D. magna inhabited in shallower water. Furthermore, small-sized microplastics interfered with the inducible behavioral defense of D. magna more strongly than large-sized polystyrene microplastics. Such findings suggested that polystyrene microplastics can weaken the predator-induced defenses of Daphnia, which may increase their risk of predation and alter the population dynamics of zooplankton.

Cardiovascular concentration-effect relationships of amodiaquine and its metabolite desethylamodiaquine: Clinical and preclinical studies.

Amodiaquine is a 4-aminoquinoline used extensively for the treatment and prevention of malaria. Orally administered amodiaquine is largely converted to the active metabolite desethylamodiaquine. Amodiaquine can cause bradycardia, hypotension, and electrocardiograph QT interval prolongation, but the relationship of these changes to drug concentrations is not well characterized. We conducted a secondary analysis of a pharmacokinetic study of the cardiac safety of amodiaquine (10mg base/kg/day over 3days) in 54 Kenyan adults (≥18 years) with uncomplicated malaria. Nonlinear mixed effects modelling was used to assess amodiaquine and desethylamodiaquine concentration-effect relationships for vital sign (pulse rate, blood pressure) and electrocardiograph interval (QT, QRS, PR) outcomes. We also measured the spontaneous beating heart rate after cumulative dosing of amodiaquine and desethylamodiaquine in isolated mouse atrial preparations. Amodiaquine and desethylamodiaquine caused concentration-dependent mean decreases in pulse rate (1.9 beats/min per 100 nmol/L; 95% confidence interval: 1.5-2.4), supine systolic blood pressure (1.7mmHg per 100 nmol/L; 1.2-2.1), erect systolic blood pressure (1.5mmHg per 100 nmol/L; 1.0-2.0) and erect diastolic blood pressure (1.4mmHg per 100 nmol/L; 1.0-1.7). The mean QT interval prolongation was 1.4 ms per 100 nmol/L irrespective of correction factor after adjustment for residual heart rate dependency. There was no significant effect of drug concentration on postural change in blood pressure or PR and QRS intervals. In mouse atria, the spontaneous beating rate was significantly reduced by amodiaquine (n= 6) and desethylamodiaquine (n= 8) at 3μmol/L (amodiaquine: 10±2%; desethylamodiaquine: 12 ±3%) and 10μmol/L (amodiaquine: 50 ±7%; desethylamodiaquine: 46 ±6%) concentrations with no significant difference in potency between the 2 compounds. Amodiaquine and desethylamodiaquine have concentration-dependent effects on heart rate, blood pressure, and ventricular repolarization.

Human in vitro neurocardiac coculture (ivNCC) assay development for evaluating cardiac contractility modulation.

Two of the most prominent organ systems, the nervous and the cardiovascular systems, are intricately connected to maintain homeostasis in mammals. Recent years have shown tremendous efforts toward therapeutic modulation of cardiac contractility and electrophysiology by electrical stimulation. Neuronal innervation and cardiac ganglia regulation are often overlooked when developing in vitro models for cardiac devices, but it is likely that peripheral nervous system plays a role in the clinical effects. We developed an in vitro neurocardiac coculture (ivNCC) model system to study cardiac and neuronal interplay using human induced pluripotent stem cell (hiPSC) technology. We demonstrated significant expression and colocalization of cardiac markers including troponin, α-actinin, and neuronal marker peripherin in neurocardiac coculture. To assess functional coupling between the cardiomyocytes and neurons, we evaluated nicotine-induced β-adrenergic norepinephrine effect and found beat rate was significantly increased in ivNCC as compared to monoculture alone. The developed platform was used as a nonclinical model for the assessment of cardiac medical devices that deliver nonexcitatory electrical pulses to the heart during the absolute refractory period of the cardiac cycle, that is, cardiac contractility modulation (CCM) therapy. Robust coculture response was observed at 14 V/cm (5V, 64 mA), monophasic, 2ms pulse duration for pacing and 20 V/cm (7V, 90 mA) phase amplitude, biphasic, 5.14 ms pulse duration for CCM. We observed that the CCM effect and kinetics were more pronounced in coculture as compared to cardiac monoculture, supporting a hypothesis that some part of CCM mechanism of action can be attributed to peripheral nervous system stimulation. This study provides novel characterization of CCM effects on hiPSC-derived neurocardiac cocultures. This innervated human heart model can be further extended to investigate arrhythmic mechanisms, neurocardiac safety, and toxicity post-chronic exposure to materials, drugs, and medical devices. We present data on acute CCM electrical stimulation effects on a functional and optimized coculture using commercially available hiPSC-derived cardiomyocytes and neurons. Moreover, this study provides an in vitro human heart model to evaluate neuronal innervation and cardiac ganglia regulation of contractility by applying CCM pulse parameters that closely resemble clinical setting. This ivNCC platform provides a potential tool for investigating aspects of cardiac and neurological device safety and performance.

Inhibitory Effect of a Late Sodium Current Blocker, NCC-3902, on the Automaticity of the Guinea Pig Pulmonary Vein Myocardium.

The effect of blocking the persistent component of the sodium channel current (late INa) on the automatic activity of the isolated guinea pig pulmonary vein myocardium was examined. NCC-3902 blocked late INa, but did not affect other major ion channel currents stably expressed in cell lines. In isolated pulmonary vein cardiomyocytes, NCC-3902 blocked the late INa induced by a ramp depolarizing voltage clamp pulse similar to that of the pacemaker depolarization observed in the pulmonary vein myocardium. In isolated pulmonary vein tissue, NCC-3902 decreased the frequency of automatic firing of the myocardium through a reduction of the pacemaker depolarization slope. In isolated pulmonary vein cardiomyocytes, NCC-3902 significantly reduced the firing frequency of Ca2+ transients, but had no effect on Ca2+ sparks. NCC-3902 affected neither the spontaneous beating rate of the right atrium nor the contractile force of the ventricular myocardium. Selective blockers of late INa like NCC-3902, which inhibit the automatic activity of the pulmonary vein myocardium, appear to be promising as drugs for the pharmacological treatment of atrial fibrillation.

Importance of beating rate control for the analysis of drug effects on contractility in human induced pluripotent stem cell-derived cardiomyocytes

Cardiac contractility evaluation using human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) has recently attracted much attention as a clinical cardiotoxicity predictive model. Most studies on this were conducted under spontaneous beating conditions and involved video-based analyses. Cardiac contractility is known to be influenced by beating rates; accordingly, beating rate control is recommended to accurately analyze the effects of drugs on cardiac contractility. Therefore, we investigated the relationship between contraction parameters and beating rates of cardiac cell sheet tissues by directly measuring the contraction force and compared the effects of ion channel drugs (mexiletine, ranolazine, and dofetilide) on contraction parameters under spontaneous beating conditions with those under pacing (1 Hz) conditions. To characterize the contraction/relaxation kinetics, we introduced a novel analysis tool, called a “C-V loop,” a plot of contraction force versus force-changing rate (“velocity”). When we increased the beating rate, the contraction force, force-changing rate, and relaxation time markedly decreased. The occurrence frequencies of beating arrest and irregular beats at high concentration ranges of mexiletine and ranolazine were more suppressed in paced samples than in spontaneously beating ones. We also found that relaxation time increased by treatment with dofetilide and contraction amplitude decreased in a concentration-dependent manner by mexiletine treatment only in the samples under pacing. These drug responses were consistent with the previous reports using human samples. These results indicated that beating rate control is necessary to stably evaluate the effects of drugs on contractility and that tests under 1-Hz pacing are more relevant to clinical settings.

Hybrid Cell Analysis System to Assess Structural and Contractile Changes of Human iPSC-Derived Cardiomyocytes for Preclinical Cardiac Risk Evaluation.

Cardiac contractility assessment is of immense importance for the development of new therapeutics and their safe transition into clinical stages. While human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) hold promise to serve as a human-relevant model in preclinical phases of drug discovery and safety pharmacology, their maturity is still controversial in the scientific community and under constant development. We present a hybrid contractility and impedance/extracellular field potential (EFP) technology, adding significant pro-maturation features to an industry-standard 96-well platform. The impedance/EFP system monitors cellular functionality in real-time. Besides the beat rate of contractile cells, the electrical impedance spectroscopy readouts detect compound-induced morphological changes like cell density and integrity of the cellular monolayer. In the other component of the hybrid cell analysis system, the cells are cultured on bio-compliant membranes that mimic the mechanical environment of real heart tissue. This physiological environment supports the maturation of hiPSC-CMs in vitro, leading to more adult-like contractile responses including positive inotropic effects after treatment with isoproterenol, S-Bay K8644, or omecamtiv mecarbil. Parameters such as the amplitude of contraction force (mN/mm2) and beat duration also reveal downstream effects of compounds with influence on electrophysiological properties and calcium handling. The hybrid system provides the ideal tool for holistic cell analysis, allowing preclinical cardiac risk assessment beyond the current perspectives of human-relevant cell-based assays.

Radiation-induced electrophysiological remodelling in neonatal rat ventricular cardiomyocytes

Abstract Background Ventricular tachycardia (VT) carries a relatively high risk of recurrence and significantly impacts patients' quality of life, despite state of the art treatment including antiarrhythmic drug therapy and catheter ablation. Cardiac stereotactic body radiotherapy (SBRT) has emerged as a promising non-invasive treatment option for refractory VT. A marked reduction in VT episodes is observed within days of treating high-risk patients with a single fraction SBRT. The cellular mechanisms underlying the clinically demonstrated antiarrhythmic effect are still poorly understood. Radiation-induced fibrosis is observed several months after irradiation, suggesting other mechanisms that explain the immediate benefit of radiotherapy on the VT substrate. Purpose We hypothesized that early antiarrhythmic effects of single-fraction radiation are produced by direct molecular and functional electrophysiological changes, independent of fibrosis. Our aim was to investigate the effects of 20 Gy ionizing photon radiation on the electrophysiology of ventricular neonatal rat cardiomyocytes at time periods from 24 hours to 96 hours after irradiation. Methods Ventricular neonatal rat cardiomyocytes were isolated, cultivated for 24 to 96 hours and then exposed to photon beam with a single dose of 20 Gy. Expression changes of ion channels, calcium handling proteins and structural proteins affecting cellular electrophysiology were assessed via RT-qPCR and Western blotting. Beat frequency was evaluated by video analysis and action potentials (APD) were recorded using the whole-cell voltage clamp technique in current-clamp mode. Intracellular calcium transients in Fura-2 AM loaded cardiomyocytes were recorded with an IonOptix recording system. Results Irradiation lead to significant changes in the mRNA and protein level of ion channels, with most distinct changes being observed 96h after radiation. Increased mRNA and protein expression of (Cav1.2) and of the potassium channels Kcnd3 (Kv4.3), Kcnh2 (Kv11.1), Kcnq1 (Kv7.1), Kcnk2 (K2P2.1) and Kcnj2 (Kir2.1) was observed. Additionally, Connexin 43 (Cx43) was upregulated. Irradiated cardiomyocytes showed a significant increase in beat rate over time. APD was significantly shortened, with a significantly increased upstroke velocity. Regarding calcium handling, significant upregulations of Ryr2 (RYR2) and Slc8a1 (NCX) transcript and protein level was detected. Additionally, calcium handling properties were altered. Conclusion Our results suggest that acute irradiation effects are associated with electrophysiological remodelling in cardiomyocytes. Changes of cellular ion channel expression and calcium homeostasis lead to normalization and shortening of heart-failure associated APD prolongation. These results reveal new insights into acute antiarrhythmic effects on cardiomyocytes after SBRT-therapy. Funding Acknowledgement Type of funding sources: None.

Elevated thyroid-stimulating hormone (TSH) is associated with atrial fibrillation and directly modulates electrophysiological properties of atrial cardiomyocytes

Abstract Background Thyroid dysfunction has a strong association with cardiac arrhythmias. However, subclinical hypothyroidism (SH) is often neglected in studies. SH is characterized by increased values of TSH and normal values of T3/T4. Recent studies indicate that high TSH levels are related to an increased incidence of cardiac electrical abnormalities, especially atrial fibrillation (AF). Purpose The present work aims to investigate direct effects of TSH on ion channel remodelling in cardiomyocytes and a possible causative role in the increased occurrence of arrhythmia in SH. Methods A retrospective monocentric study analysed medical records, ECGs and echocardiographies of 3134 patients between 2007 to 2020. Inclusion criteria comprised patient age ≥18 years, normal fT4, TSH values 4–75 mU/l and available ECGs. The patient cohort was divided into two groups based on current treatment recommendations for SH (TSH level 4–9.9 mU/l; 10–75 mU/l). Direct effects on ion channel remodelling were analysed in HL-1 and neonatal cardiomyocytes with confirmed expression of TSH receptors. TSH concentrations of 15, 30, and 60 mU/l simulated different degrees of hypothyroidism. RT-qPCR analyses, western blots and immunofluorescence stainings were performed after incubation with respective TSH concentrations for 24h. In vitro video analyses of automatism in NRCM before and after TSH incubation were performed. Statistical comparisons were calculated using student's t-tests and the chi-square test. In addition, a multivariate analysis of factors affecting the occurrence of AF in patients with SH was carried out. Results Elevated TSH levels were associated with higher prevalence of AF. In the group of patients with a TSH value of 10–75 mU/l, AF was observed in 42.8% of cases compared to 31.7% in patients with lower TSH levels (p<0.0001). In vitro, TSH application to cardiomyocytes led to significant changes in mRNA and protein expression of key ion channels and regulatory proteins. Incubation of HL-1 cells with 15 mU/l TSH showed a decrease in Kcne1 protein, while 60 mU/l TSH caused an upregulation of K2P3.1 (TASK-1), Kv7.1, and KCa2.1 expression. To further elucidate the TSH signaling cascade in cardiomyocytes, we investigated TSH effects on PKAα, CaMKIIδ and CaMKIIγ. PKAα was upregulated after 24h incubation with 60 mU/l TSH and exhibited an increase of the phosphorylated state. Finally, neonatal cardiomyocytes showed an increased beating rate after incubation with TSH. Conclusion The prevalence of atrial fibrillation is significantly increased in patients with SH. Our findings further reveal a direct effect of TSH on ion channel expression in cardiomyocytes as potential proarrhythmic mechanism. TSH induced changes in the expression of ion channels that determine action potential duration. Subclinical hypothyroidism contributes to the development of atrial fibrillation. Underlying mechanisms and therapeutic implications require further research. Funding Acknowledgement Type of funding sources: Public Institution(s). Main funding source(s): Heidelberg University Medical School - Olympia Morata

Role of Endothelin-1 in Right Atrial Arrhythmogenesis in Rabbits with Monocrotaline-Induced Pulmonary Arterial Hypertension.

Atrial arrhythmias are considered prominent phenomena in pulmonary arterial hypertension (PAH) resulting from atrial electrical and structural remodeling. Endothelin (ET)-1 levels correlate with PAH severity and are associated with atrial remodeling and arrhythmia. In this study, hemodynamic measurement, western blot analysis, and histopathology were performed in the control and monocrotaline (MCT, 60 mg/kg)-induced PAH rabbits. Conventional microelectrodes were used to simultaneously record the electrical activity in the isolated sinoatrial node (SAN) and right atrium (RA) tissue preparations before and after ET-1 (10 nM) or BQ-485 (an ET-A receptor antagonist, 100 nM) perfusion. MCT-treated rabbits showed an increased relative wall thickness in the pulmonary arterioles, mean cell width, cross-sectional area of RV myocytes, and higher right ventricular systolic pressure, which were deemed to have PAH. Compared to the control, the spontaneous beating rate of SAN–RA preparations was faster in the MCT-induced PAH group, which can be slowed down by ET-1. MCT-induced PAH rabbits had a higher incidence of sinoatrial conduction blocks, and ET-1 can induce atrial premature beats or short runs of intra-atrial reentrant tachycardia. BQ 485 administration can mitigate ET-1-induced RA arrhythmogenesis in MCT-induced PAH. The RA specimens from MCT-induced PAH rabbits had a smaller connexin 43 and larger ROCK1 and phosphorylated Akt than the control, and similar PKG and Akt to the control. In conclusion, ET-1 acts as a trigger factor to interact with the arrhythmogenic substrate to initiate and maintain atrial arrhythmias in PAH. ET-1/ET-A receptor/ROCK signaling may be a target for therapeutic interventions to treat PAH-induced atrial arrhythmias.

Neural synchronization is strongest to the spectral flux of slow music and depends on familiarity and beat salience.

Neural activity in the auditory system synchronizes to sound rhythms, and brain-environment synchronization is thought to be fundamental to successful auditory perception. Sound rhythms are often operationalized in terms of the sound's amplitude envelope. We hypothesized that - especially for music - the envelope might not best capture the complex spectro-temporal fluctuations that give rise to beat perception and synchronized neural activity. This study investigated (1) neural synchronization to different musical features, (2) tempo-dependence of neural synchronization, and (3) dependence of synchronization on familiarity, enjoyment, and ease of beat perception. In this electroencephalography study, 37 human participants listened to tempo-modulated music (1-4 Hz). Independent of whether the analysis approach was based on temporal response functions (TRFs) or reliable components analysis (RCA), the spectral flux of music - as opposed to the amplitude envelope - evoked strongest neural synchronization. Moreover, music with slower beat rates, high familiarity, and easy-to-perceive beats elicited the strongest neural response. Our results demonstrate the importance of spectro-temporal fluctuations in music for driving neural synchronization, and highlight its sensitivity to musical tempo, familiarity, and beat salience.

Micropatterned Hydrogels with Highly Ordered Cellulose Nanocrystals for Visually Monitoring Cardiomyocytes.

Cardiac microphysiological systems are accurate in vitro platforms that reveal the biological mechanisms underlying cardiopathy, accelerating pharmaceutical research in this field. Current cardiac microphysiological devices and organs-on-chips consist of several layers prepared with complex, multi-step processes. Incorporating inorganic photonic crystals may cause long-term biocompatibility issues. Herein, micropatterned hydrogels with anisotropic structural colors are prepared by locking shear-oriented tunicate cellulose nanocrystals (TCNCs) in hydrogel networks through in situ polymerization, allowing the visualization and monitoring of cardiomyocytes. The anisotropic hydrogels are composed of highly ordered TCNCs with bright interference color and micro-grooved methacrylated gelatin with excellent biocompatibility. The microgroove patterns induce cardiomyocyte alignment and the autonomous beating of cardiomyocytes causes the hydrogels to deform, dynamically shifting the interference color. These micropatterned hydrogels could noninvasively monitor real-time changes of cardiomyocytes under pharmaceutical treatment and electrical stimulation through wavelength shifts in the transmittance spectra. This system provides a new way to detect the beat rate of cardiac tissue and it may contribute to high throughput develop.

Space microgravity improves proliferation of human iPSC-derived cardiomyocytes.

SummaryIn microgravity, cells undergo profound changes in their properties. However, how human cardiac progenitors respond to space microgravity is unknown. In this study, we evaluated the effect of space microgravity on differentiation of human induced pluripotent stem cell (hiPSC)-derived cardiac progenitors compared with 1G cultures on the International Space Station (ISS). Cryopreserved 3D cardiac progenitors were cultured for 3 weeks on the ISS. Compared with 1G cultures, the microgravity cultures had 3-fold larger sphere sizes, 20-fold higher counts of nuclei, and increased expression of proliferation markers. Highly enriched cardiomyocytes generated in space microgravity showed improved Ca2+ handling and increased expression of contraction-associated genes. Short-term exposure (3 days) of cardiac progenitors to space microgravity upregulated genes involved in cell proliferation, survival, cardiac differentiation, and contraction, consistent with improved microgravity cultures at the late stage. These results indicate that space microgravity increased proliferation of hiPSC-cardiomyocytes, which had appropriate structure and function.

Infestation of sporocysts of parasite Leucochloridium in the snails Succinea daucina and Indosuccinea semiserica

Aim: The aim of the present study was to evaluate the differential beating rate of the parasite Leucochloridium within the tentacle of the infected snail Succinea daucina and Indosuccinea semiserica under different photoperiodic conditions. Methodology: In this study, the rhythmic beating pulse of sporocyst of the parasite invading the tentacle of the snail was recorded from multiple snails, periodically in light and dark hours. The numbers of beat per minute were subjected to univariate ANOVA using day and photoperiod as explanatory variables. A two-sample t-test was performed with the mean of pulse rate in light and dark phase to estimate the significant differences between the same. Results: The observations revealed that sporocyst brood sac of I. semiserica beat mean 99.23 ± 1.91 SE per minute ( 90 – 101) in day hours and mean 76.57 ± 4.64 SE per minute (48 – 88) in the night hours (n = 7 observations). In S. daucina, the sporocyst brood sac beat 65.85 ± 2.17 SE per minute (25 – 94) during day and 30.5 ± 3.55 SE per minute (0 – 80) in night (n = 9 separate observations). The beats of the sporocyst varied significantly diurnally and with the consecutive days revealed through t-test and ANOVA. Interpretation: Apparently, the colourful sporocyst broodsac of Leucochloridium faster beating rate under light may attract definite host, the insectivorous birds which prey on the Leucochloridium sporocycst disrupting the snail tentacle. Thus, the pulsating sporocyst inside the tentacle makes the infected host snail more conspicuous for the birds, linking the snail- Leucochloridium- bird cycle. Key words: Indosuccinea semiserica, Intermediate host, Land snail, Leucochloridium sp., Succinea daucina, Trematoda

Effect of macrophage migration inhibitory factor on pulmonary vein arrhythmogenesis through late sodium current.

Macrophage migration inhibitory factor (MIF), a pleiotropic inflammatory cytokine, is highly expressed in patients with atrial fibrillation (AF). Inflammation increases the risk of AF and is primarily triggered by pulmonary vein (PV) arrhythmogenesis. This study investigated whether MIF can modulate the electrical activity of the PV and examined the underlying mechanisms of MIF. A conventional microelectrode, a whole-cell patch clamp, western blotting, and immunofluorescent confocal microscopy were used to investigate electrical activity, calcium (Ca2+) regulation, protein expression, ionic currents, and cytosolic reactive oxygen species (ROS) in rabbit PV tissue and isolated single cardiomyocytes with and without MIF incubation (100 ng/mL, treated for 6 h). The MIF (100 ng/mL)-treated PV tissue (n = 8) demonstrated a faster beating rate (1.8 ± 0.2 vs. 2.6 ± 0.1 Hz, P < 0.05), higher incidence of triggered activity (12.5 vs. 100%, P < 0.05), and premature atrial beat (0 vs. 100%, P < 0.05) than the control PV tissue (n = 8). Compared with the control PV cardiomyocytes, MIF-treated single PV cardiomyocytes had larger Ca2+ transients (0.6 ± 0.1 vs. 1.0 ± 0.1, ΔF/F0, P < 0.05), sarcoplasmic reticulum Ca2+ content (0.9 ± 0.20 vs. 1.7 ± 0.3 mM of cytosol, P < 0.05), and cytosolic ROS (146.8 ± 5.3 vs. 163.7 ± 3.8, ΔF/F0, P < 0.05). Moreover, MIF-treated PV cardiomyocytes exhibited larger late sodium currents (INa-Late), L-type Ca2+ currents, and Na+/Ca2+ exchanger currents than the control PV cardiomyocytes. KN93 [a selective calcium/calmodulin-dependent protein kinase II (CaMKII) blocker, 1 μM], ranolazine (an INa-Late inhibitor, 10 μM), and N-(mercaptopropionyl) glycine (ROS inhibitor, 10 mM) reduced the beating rates and the incidence of triggered activity and premature captures in the MIF-treated PV tissue. Macrophage migration inhibitory factor increased PV arrhythmogenesis through Na+ and Ca2+ dysregulation through the ROS activation of CaMKII signalling, which may contribute to the genesis of AF during inflammation. Anti-CaMKII treatment may reverse PV arrhythmogenesis. Our results clearly reveal a key link between MIF and AF and offer a viable therapeutic target for AF treatment.