Action Potential Recordings Research Articles

Train-of-four ratio, counts and post tetanic counts with the Tetragraph electromyograph in comparison to mechanomyography.

Automated EMG devices to detect compound muscle action potentials from the adductor pollicis muscle in response to ulnar nerve stimulation, regardless of hand and thumb position, may serve as a better reference ("gold standard") for clinical assessment of neuromuscular function than traditional mechanomyography (MMG) systems that need custom design and validation in lab settings. This evaluation compared the TetraGraph EMG system against a validated MMG device to investigate the accuracy and repeatability of this quantitative EMG monitor for detecting onset, offset and deep neuromuscular block. Simultaneous muscle action potential recordings from the EMG neuromuscular monitor and muscle contractions from an in-house developed MMG monitor in response to ulnar nerve stimulation were obtained from patients having elective surgery requiring neuromuscular block. Train-of-four (TOF) ratios, TOF counts, and post-tetanic counts (PTCs) were recorded simultaneously from the same hand muscle and compared. In total, 685 pairs of simultaneous TOF ratios were evaluated. The mean difference (bias) of TOF ratios between devices was small (-2.1%). TOF counts from 285 data pairs were within a count of 2 or less 96% of the time. During deep block, PTC comparisons from 215 data pairs were within a count of 2 or less 95% of the time. These findings, along with prior EMG device evaluations, indicate that real-time EMG neuromuscular monitoring technology to detect muscle action potentials from the adductor pollicis in the clinical setting is closely aligned with the force of thumb contraction determined from MMG. The accuracy of quantitative EMG technology of the TetraGraph EMG system lends strong support for this monitor, along with other similarly validated EMG monitors, to become a clinical standard for all phases (onset, depth and reversal) of neuromuscular block in clinical practice.

Calmodulin kinase II inhibition suppresses atrioventricular conduction by regulating intracellular Ca2+ homeostasis

BackgroundCa2+/calmodulin-dependent protein kinase II (CaMKII) inhibition decelerates atrioventricular node (AVN) conduction, providing a potential treatment of tachycardia. However, the effectiveness of CaMKII inhibition on tachycardia and its underlying mechanism remains unclear. ObjectiveWe aimed to assess the effectiveness of CaMKII inhibition in reducing ventricular rates during atrial fibrillation and to elucidate the underlying mechanism in affecting AVN electrophysiology. MethodsCardiac CaMKII inhibition (AC3-I) mice were used. Transesophageal atrial pacing was performed to evaluate AVN conduction function and to induce atrial fibrillation. Patch-clamp techniques were employed to record action potentials and ionic currents in AVN cells. Intracellular Ca2+ transients and sarcomere length measurements were obtained with the IonOptix system. Masson trichrome stain was used to evaluate fibrosis in the AVN region. Western blotting and immunofluorescence techniques were employed to detect connexin expression and localization. ResultsCaMKII inhibition decreased the ventricular rate during atrial fibrillation and isoproterenol-induced tachycardia. Esophageal electrocardiogram results from AC3-I mice showed longer AVN conduction than in wild-type mice. AN- and N-type AVN cells from AC3-I mice exhibited slower action potential frequencies and diastolic depolarization rates than those of wild-type mice. The study revealed that CaMKII inhibition reduced AVN cell sarcoplasmic reticulum (SR) Ca2+ content, Ca2+ release rate from the SR during diastole, Ca2+ transient amplitude, and SR Ca2+ uptake rate. In addition, CaMKII inhibition prolonged the sarcomere diastole duration and enhanced the sensitivity of sarcomeres to Ca2+. ConclusionCaMKII inhibition effectively decreases the ventricular rate during atrial fibrillation and tachycardia by slowing down AVN conduction through suppressing Ca2+ overload in AVN cells.

Magnetic Detection of Neural Activity by Nanocoil Transducers.

Electrophysiological recordings from brain cells are performed routinely using implanted electrodes, but they traditionally require a wired connection to the outside of the brain. A completely passive, wireless device that does not require on-board power for active transmission but that still facilitates remote detection could open the door for mass-scale direct recording of action potentials and transform the way we acquire brain signals. We present a nanofabricated coil that forms a neuroelectromagnetic junction, yielding a highly enhanced magnetic field transduction of electrophysiology. We show that this micrometer-scale device enables remote magnetic detection of neuronal fields from the center of the coil using room temperature superconducting quantum interference device (SQUID) microscopy. Further, time-locked stimulation in conjunction with magnetometry demonstrates thresholding behavior that affirms the viability of the technology for detection with no requirement for wires or on-board power. This strategy may permit unprecedented detection of electrophysiology using magnetoencephalography and magnetic resonance imaging.

Identification and characterisation of functional Kir6.1-containing ATP-sensitive potassium channels in the cardiac ventricular sarcolemmal membrane.

The canonical Kir6.2/SUR2A ventricular KATP channel is highly ATP-sensitive and remains closed under normal physiological conditions. These channels activate only when prolonged metabolic compromise causes significant ATP depletion and then shortens the action potential to reduce contractile activity. Pharmacological activation of KATP channels is cardioprotective, but physiologically, it is difficult to understand how these channels protect the heart if they only open under extreme metabolic stress. The presence of a second KATP channel population could help explain this. Here, we characterise the biophysical and pharmacological behaviours of a constitutively active Kir6.1-containing KATP channel in ventricular cardiomyocytes. Patch-clamp recordings from rat ventricular myocytes in combination with well-defined pharmacological modulators was used to characterise these newly identified K+ channels. Action potential recording, calcium (Fluo-4) fluorescence measurements and video edge detection of contractile function were used to assess functional consequences of channel modulation. Our data show a ventricular K+ conductance whose biophysical characteristics and response to pharmacological modulation were consistent with Kir6.1-containing channels. These Kir6.1-containing channels lack the ATP-sensitivity of the canonical channels and are constitutively active. We conclude there are two functionally distinct populations of ventricular KATP channels: constitutively active Kir6.1-containing channels that play an important role in fine-tuning the action potential and Kir6.2/SUR2A channels that activate with prolonged ischaemia to impart late-stage protection against catastrophic ATP depletion. Further research is required to determine whether Kir6.1 is an overlooked target in Comprehensive in vitro Proarrhythmia Assay (CiPA) cardiac safety screens.

Sapphire-Based Optrode for Low Noise Neural Recording and Optogenetic Manipulation.

Electrophysiological recordings of neurons in deep brain regions using optogenetic stimulation are essential to understanding and regulating the role of complex neural activity in biological behavior and cognitive function. Optogenetic techniques have significantly advanced neuroscience research by enabling the optical manipulation of neural activities. Because of the significance of the technique, constant advancements in implantable optrodes that integrate optical stimulation with low-noise, large-scale electrophysiological recording are in demand to improve the spatiotemporal resolution for various experimental designs and future clinical applications. However, robust and easy-to-use neural optrodes that integrate neural recording arrays with high-intensity light emitting diodes (LEDs) are still lacking. Here, we propose a neural optrode based on Gallium Nitride (GaN) on sapphire technology, which integrates a high-intensity blue LED with a 5x2 recording array monolithically for simultaneous neural recording and optogenetic manipulation. To reduce the noise interference between the recording electrodes and the LED, which is in close physical proximity, three metal grounding interlayers were incorporated within the optrode, and their ability to reduce LED-induced artifacts during neural recording was confirmed through both electromagnetic simulations and experimental demonstrations. The capability of the sapphire optrode to record action potentials has been demonstrated by recording the firing of mitral/tuft cells in the olfactory bulbs of mice in vivo. Additionally, the elevation of action potential firing due to optogenetic stimulation observed using the sapphire probe in medial superior olive (MSO) neurons of the gerbil auditory brainstem confirms the capability of this sapphire optrode to precisely access neural activities in deep brain regions under complex experimental designs.

Hepatocyte Growth Factor Delivery to Injured Cervical Spinal Cord Using an Engineered Biomaterial Protects Respiratory Neural Circuitry and Preserves Functional Diaphragm Innervation.

A major portion of spinal cord injury (SCI) cases occur in the cervical region, where essential components of the respiratory neural circuitry are located. Phrenic motor neurons (PhMNs) housed at cervical spinal cord level C3-C5 directly innervate the diaphragm, and SCI-induced damage to these cells severely impairs respiratory function. In this study, we tested a biomaterial-based approach aimed at preserving this critical phrenic motor circuitry after cervical SCI by locally delivering hepatocyte growth factor (HGF). HGF is a potent mitogen that promotes survival, proliferation, migration, repair, and regeneration of a number of different cell and tissue types in response to injury. We developed a hydrogel-based HGF delivery system that can be injected into the intrathecal space for local delivery of high levels of HGF without damaging the spinal cord. Implantation of HGF hydrogel after unilateral C5 contusion-type SCI in rats preserved diaphragm function, as assessed by invivo recordings of both compound muscle action potentials and inspiratory electromyography amplitudes. HGF hydrogel also preserved PhMN innervation of the diaphragm, as assessed by both retrograde PhMN tracing and detailed neuromuscular junction morphological analysis. Furthermore, HGF hydrogel significantly decreased lesion size and degeneration of cervical motor neuron cell bodies, as well as reduced levels surrounding the injury site of scar-associated chondroitin sulfate proteoglycan molecules that limit axon growth capacity. Our findings demonstrate that local biomaterial-based delivery of HGF hydrogel to injured cervical spinal cord is an effective strategy for preserving respiratory circuitry and diaphragm function.

Recordings of Superior Laryngeal Nerve Sensory Nerve Action Potentials in a Rat Model.

Superior laryngeal nerve (SLN) function is critical to laryngeal sensation. Sensory dysfunction in the larynx, mediated through the internal branch of the superior laryngeal nerve (iSLN), is thought to occur with aging and neurodegenerative disease. However, objective analysis of iSLN neurophysiology is difficult due to its anatomic location and small diameter. This study measures sensory nerve action potentials (SNAP) from the iSLN in a rat model. SNAP data were obtained from two adult rat strains (Sprague-Dawley, SD and Fischer 344 × Brown Norway F1 Hybrid rats, FBN). Evoked responses were obtained by stimulating the main trunk of the SLN and recording the response using a 160-μm cuff electrode placed around the iSLN. SNAP were averaged from 10 stimulations. Laryngeal adductor reflex (LAR) threshold measurements were obtained with stimulation of the iSLN and direct laryngoscopy. The sections of the iSLN were obtained for histologic analysis. SLN-evoked responses were successfully obtained in 18 hemi-laryngeal preparations (SD n = 13 and FBN n = 5) with corresponding LAR threshold measurements. Mean(±SD) SNAP latency, total duration, amplitude, negative durations, and intensity were 2.28 ms (±0.56), 2.13 ms (±0.70), 879 μV (±535), and 0.69 mA (±0.25), respectively. SLN stimulation threshold to elicit an LAR was of 0.84 mA (±0.31). It is feasible to record evoked SLN responses in two adult rat strains. This work may lead to a tractable animal model for objective measurements of SLN neurophysiology with various disease states. N/A Laryngoscope, 2024.

Abstract Mo053: Acacetin's Antiarrhythmic Potential in Human-Induced Pluripotent Stem Cell-Derived Cardiomyocytes Harboring Hypertrophic and Dilated Cardiomyopathy-Related Mutations

Background: Hypertrophic cardiomyopathy (HCM) is often caused by mutations such as p.R943X in the Myosin Binding Protein C3 (MYBPC3), leading to early and delayed afterdepolarizations, myofibrillar disarray, and calcium dysfunction, which may induce lethal arrhythmias. Dilated cardiomyopathy (DCM) can also result from mutations, such as the p.R14del mutation in the phospholamban (PLN) protein, which is crucial for calcium signaling regulation, leading to arrhythmogenesis when provoked. This study explores the arrhythmogenic potential in human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) carrying the HCM MYBPC3 p.R943X mutation and the DCM PLN p.R14del mutation. Methods: We utilized a 384-well optical physiological system to record action potentials (APs) in hiPSC-CMs carrying the MYBPC3 p.R943X mutation and the DCM PLN p.R14del mutation, including their isogenic controls. Action potential metrics were measured under spontaneous and electrically paced conditions. The effects of Acacetin, NS-5806, nifedipine, and GS-967 on the recorded AP types were analyzed to understand the underlying channelopathies and arrhythmogenic potential. Special attention was given to the variability between the HCM MYBPC3 p.R943X and DCM PLN p.R14del cell lines' susceptibility to arrhythmogenesis and their isogenic controls. Results: The HCM MYBPC3 p.R943X hiPSC-CMs exhibited a significant increase in early afterdepolarizations, non-sustained and sustained ventricular tachycardia, and AP notches compared to their isogenic control. AP notches and arrhythmogenic events were notably provoked by NS-5806 in both the HCM MYBPC3 p.R943X and DCM PLN p.R14del hiPSC-CMs. However, these were significantly ameliorated (p<0.05) by Acacetin administration (5 µM), highlighting its potential as an effective antiarrhythmic in managing both HCM- and DCM-induced arrhythmias. GS-967 also showed efficacy in reducing early afterdepolarizations (EADs) in the HCM model. Conclusions: This study provides novel insights into the arrhythmogenic potential of hiPSC-CMs carrying MYBPC3 p.R943X and PLN p.R14del mutations, underscoring the significant role of Acacetin in attenuating these effects. It paves the way for developing targeted therapies that address the complex molecular mechanisms underlying arrhythmogenesis in cardiomyopathies. The findings suggest that Acacetin could be a promising candidate for the pharmacologic treatment of cardiomyopathies characterized by arrhythmogenic risks.

NeuroRoots, a bio-inspired, seamless brain machine interface for long-term recording in delicate brain regions.

Scalable electronic brain implants with long-term stability and low biological perturbation are crucial technologies for high-quality brain-machine interfaces that can seamlessly access delicate and hard-to-reach regions of the brain. Here, we created "NeuroRoots," a biomimetic multi-channel implant with similar dimensions (7 μm wide and 1.5 μm thick), mechanical compliance, and spatial distribution as axons in the brain. Unlike planar shank implants, these devices consist of a number of individual electrode "roots," each tendril independent from the other. A simple microscale delivery approach based on commercially available apparatus minimally perturbs existing neural architectures during surgery. NeuroRoots enables high density single unit recording from the cerebellum in vitro and in vivo. NeuroRoots also reliably recorded action potentials in various brain regions for at least 7 weeks during behavioral experiments in freely-moving rats, without adjustment of electrode position. This minimally invasive axon-like implant design is an important step toward improving the integration and stability of brain-machine interfacing.

Multifunctional Nanomesh Enables Cellular-Resolution, Elastic Neuroelectronics.

Silicone-based devices have the potential to achieve an ideal interface with nervous tissue but suffer from scalability, primarily due to the mechanical mismatch between established electronic materials and soft elastomer substrates. This study presents a novel approach using conventional electrode materials through multifunctional nanomesh to achieve reliable elastic microelectrodes directly on polydimethylsiloxane (PDMS) silicone with an unprecedented cellular resolution. This engineered nanomesh features an in-plane nanoscale mesh pattern, physically embodied by a stack of three thin-film materials by design, namely Parylene-C for mechanical buffering, gold (Au) for electrical conduction, and Poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate) (PEDOT:PSS) for improved electrochemical interfacing. Nanomesh elastic neuroelectronics are validated using single-unit recording from the small and curvilinear epidural surface of mouse dorsal root ganglia (DRG) with device self-conformed and superior recording quality compared to plastic control devices requiring manual pressing is demonstrated. Electrode scaling studies from in vivo epidural recording further revealed the need for cellular resolution for high-fidelity recording of single-unit activities and compound action potentials. In addition to creating a minimally invasive device to effectively interface with DRG sensory afferents at a single-cell resolution, this study establishes nanomeshing as a practical pathway to leverage traditional electrode materials for a new class of elastic neuroelectronics.

Influence of Apamin on the Extracellularly Recorded Action Potentials Profiles of Subepicardial Cardiomyocytes of the Rat Heart in Myocardial Infarction

Influence of Apamin on the Extracellularly Recorded Action Potentials Profiles of Subepicardial Cardiomyocytes of the Rat Heart in Myocardial Infarction

Inhibitory modulation of action potentials in crayfish motor axons by fluoxetine.

The goal of this report is to explore how K2P channels modulate axonal excitability by using the crayfish ventral superficial flexor preparation. This preparation allows for simultaneous recording of motor nerve extracellular action potentials (eAP) and intracellular excitatory junctional potential (EJP) from a muscle fiber. Previous pharmacological studies have demonstrated the presence of K2P-like channels in crayfish. Fluoxetine (50µM) was used to block K2P channels in this study. The blocker caused a gradual decline, and eventually complete block, of motor axon action potentials. At an intermediate stage of the block, when the peak-to-peak amplitude of eAP decreased to ∼60%-80% of the control value, the amplitude of the initial positive component of eAP declined at a faster rate than that of the negative peak representing sodium influx. Furthermore, the second positive peak following this sodium influx, which corresponds to the after-hyperpolarizing phase of intracellularly recorded action potentials (iAP), became larger during the intermediate stage of eAP block. Finally, EJP recorded simultaneously with eAP showed no change in amplitude, but did show a significant increase in synaptic delay. These changes in eAP shape and EJP delay are interpreted as the consequence of depolarized resting membrane potential after K2P channel block. In addition to providing insights to possible functions of K2P channels in unmyelinated axons, results presented here also serve as an example of how changes in eAP shape contain information that can be used to infer alterations in intracellular events. This type of eAP-iAP cross-inference is valuable for gaining mechanistic insights here and may also be applicable to other model systems.

An Experimental Study of Incremental Buckling-Resistant Inchworm-Type Insertion of Microwire Neural Electrodes

Abstract Chronically implanting microelectrodes for high-resolution action potential recording is critical for understanding the brain. The smallest and most flexible electrodes, most suitable for chronic recordings, are also the most difficult to insert due to buckling against the thin but hard-to-penetrate brain meninges. To address such implantation challenges without introducing further damage to the brain, this paper presents our design and prototype of an inchworm-type insertion device that conducts a grip-feed-release incremental motion for planar microelectrode insertion. To optimize the operating parameters of the developed inchworm insertion device, experimental studies were conducted on PVC-based brain-mimicking phantom to investigate the effects of (1) incremental insertion depth, (2) inserter drive shaft rotary speed, and (3) the resulting inchworm insertion speed, on the phantom (1) penetration rupture force and (2) dimpling depth at rupture. Analysis showed that all three factors had a statistically significant impact on the rupture force and dimpling depth. A moderate level of the resulting insertion speed yielded the lowest rupture force and dimpling depth at rupture. Low insertion speed levels were associated with higher rupture force while high insertion speeds led to a large variance in dimpling depth and potential insertion failure. To achieve such a moderate insertion speed, it would be preferred for both the incremental insertion depth and the drive shaft rotary speed to be at a moderate level. Such findings lay the foundation for enabling previously impossible buckling-free insertion of miniaturized flexible planar microelectrodes deep into the brain.

Sensitive Intracellular Recording of Action Potentials by Supramolecular Self‐Assembled Electrodes

AbstractDeveloping accurate electrophysiological recording platforms is crucial for investigating cellular electrical activities, understanding the mechanisms underlying cardiovascular disease, and advancing therapeutic strategies. While the successful recording of intracellular electrophysiological status has been achieved by 3D micro/nano devices, their preparation through demanding micro/nanofabrication and poorly controllable solvothermal synthesis is an impediment to widespread utilization. Easier‐to‐access recording electrodes are needed. To address this challenge, a simple and efficient supramolecular self‐assembly strategy that allows access to novel 3D electrodes is introduced. It relies on small organic molecules that can be self‐assembled efficiently into various 3D nanostructures, including lamellated nanosheets, thin nanobelts, and short rod‐like structures. Electrodes prepared through the present self‐assembly process are considerably simpler and more cost‐effective than conventional 3D micro/nanoelectrodes prepared through fabrication or solvothermal synthesis. It is demonstrated that the present electrodes allow for higher quality and prolonged intracellular recordings of action potentials as compared to similarly sized conventional planar electrodes. The noted improvements are attributed to the fact that 3D geometrical electrodes enhance the cell‐electrode interface, ensuring tight coupling and effective electroporation. This work showcases how supramolecular self‐assembly might emerge as a promising and powerful electrode preparation strategy that may facilitate low‐cost electrophysiological studies.

In vivo magnetic recording of single-neuron action potentials Abbreviated title: In vivo magnetic single-neuron action potentials.

Measuring fast neuronal signals is the domain of electrophysiology and magnetophysiology. While electrophysiology is easier to perform, magnetophysiology avoids tissue-based distortions and measures a signal with directional information. At the macroscale, magnetoencephalography (MEG) is established, and at the mesoscale, visually evoked magnetic fields have been reported. At the microscale however, while benefits of recording magnetic counterparts of electric spikes would be numerous, they are also highly challenging in vivo. Here, we combine magnetic and electric recordings of neuronal action potentials in anesthetized rats using miniaturized giant magneto-resistance (GMR) sensors. We reveal the magnetic signature of action potentials of well-isolated single units. The recorded magnetic signals showed a distinct waveform and considerable signal strength. This demonstration of in vivo magnetic action potentials opens a wide field of possibilities to profit from the combined power of magnetic and electric recordings and thus to significantly advance the understanding of neuronal circuits.

HiPSC-derived cardiomyocyte cultivation and measurement of electrophysiological properties on gelatine/glucose/polypyrrole scaffold

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – EU funding. Main funding source(s): EU Horizon 2020 Framework Programme (H2020) under Grant agreements No. 953138 (EMAPS-Cardio) Purpose Cardiotoxicity is one of the main side effects limiting development of new medicines and resulting in withdrawal of drugs from the market. There are different mechanisms of drug-induced cardiotoxicity, including arrhythmia, disruption of mitochondria function, apoptosis, altered growth factor signalling [1]. Heart on a chip model for drug testing seems attractive approach to evaluate the possible cardiotoxic effects on cell biological, but mostly electrophysiological properties. Our aim was to create a 3D system with a biocompatible scaffold, coated with a conductive layer, suitable for culturing and maturation of human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CMs) and compatible with electrophysiology parameter measurement techniques. Methods Elastic scaffold was fabricated by electrospinning of gelatin-glucose microfibers followed by the vapor-phase and electrochemical deposition of conductive polymer polypyrrole (PPy) to create electroconductive layer. hiPSC-CMs at different maturation levels (at 6 vs 14 weeks) were used to test biocompatibility of the scaffold. To increase cell attachment, scaffold was treated with 0.1 M HCl and coated with Geltrex. Cells were stained with fluorescent dyes (Calcein AM, DAPI). Electrophysiological properties were analysed using intracellular calcium imaging (Cal520) and action potential recordings with sharp electrodes. HiPSC-CMs on coverslips were used as a 2D control. Results Calcein AM staining showed that high number of hiPSC-CMs attached to gelatine/glucose/PPy scaffold proving that the scaffold is biocompatible and cells remain viable even after long term incubation (> 2 weeks). HiPSC-CMs at lower maturity (6 weeks) had better attachment properties to it, indicating that more mature cardiomyocytes (14 weeks) gradually lose their capacity to attach to PPy surface. Intracellular calcium imaging showed that both types of cells generated synchronous spontaneous calcium oscillations on gelatine/glucose/PPy scaffolds. Action potential recordings in 3D were similar to those in 2D, and confirmed differences in hiPSC-CMs maturation levels – at lower maturation level hiPSC-CMs had lower upstroke velocity as compared to more matured hiPSC-CMs. Conclusions Gelatine/glucose/PPy scaffold is biocompatible for hiPSC-CMs cultivation, and suitable for evaluation of cardiomyocyte electrophysiological properties. Such electromechanoactive scaffold can be further employed for development of heart-on-a-chip model.

The interpretation of field and LEAP potentials recorded from cardiomyocyte monolayers

Abstract Funding Acknowledgements None. Multi-electrode arrays (MEA) are the method of choice for electrophysiological characterization of cardiomyocyte monolayers. The field potentials recorded using a MEA are like extracellular electrograms recorded from ventricular myocardium using conventional electrodes. Nevertheless, different criteria are often used to interpret field potential and extracellular electrogram, which hampers correct interpretation and translation to the patient. To validate criteria for extracellular electrograms analysis applicable to field potentials, we used neonatal rat cardiomyocytes to generate monolayers. We recorded field potentials using a MEA and simultaneously recorded action potentials using sharp micro-electrodes. In parallel we recreated our experimental setting in silico and performed simulations. We demonstrate that the amplitude of the local RS-complex of a field potential correlated with conduction velocity, but only in the lower ranges of conduction velocities. The time of the peak of the T-wave of the field potentials correlated well with APD90 and the time of the steepest upslope with APD50. By injecting current into the monolayer, we could record a local extracellular action potential (LEAP). The LEAP was of low amplitude and highly depolarized, and the shape differed markedly from the shape of the local action potential. Criteria for interpreting extra cellular electrograms should also be applied to field potentials. This will provide a strong basis for analysis of heterogeneity in conduction velocity and repolarization in cultured monolayers of cardiomyocytes especially at low conduction velocities. A LEAP is not a recording of the local action potential but is generated by intracellular current provided by neighboring cardiomyocytes.

Loss of protein phosphatase 2A regulatory subunit PPP2R5A is associated with increased incidence of stress-induced proarrhythmia.

Protein phosphatase 2A (PP2A) is a serine/threonine-selective holoenzyme that controls Ca2+ homeostasis and contractility of the heart via dephosphorylation of regulatory proteins. In some genetically modified mouse models with increased arrhythmogenicity, a reduced expression of the regulatory subunit B56α of PP2A was found as a concomitant effect. Whether there is a general correlation between the abundance of B56α and the promotion of cardiac arrhythmogenesis remains unclear. The aim of this study was therefore to investigate the role of PP2A-B56α in the propensity for arrhythmic activity in the heart. The experimental analysis of this question has been addressed by using a mouse model with deletion of the PP2A-B56α gene, PPP2R5A (KO), in comparison to wild-type animals (WT). Evidence for arrhythmogenicity was investigated in whole animal, isolated heart and cardiomyocytes by ECG, recording of monophasic action potential (MAP) induced by programmed electrical stimulation (PES), measurement of Ca2+ transients under increased pacing frequencies and determination of total K+ channel currents (I K). ECG measurements showed a prolongation of QT time in KO vs. WT. KO mice exhibited a higher rate of premature ventricular contractions in the ECG. MAP measurements in Langendorff-perfused KO hearts showed increased episodes of ventricular tachyarrhythmia induced by PES. However, the KO hearts showed values for MAP duration that were similar to those in WT hearts. In contrast, KO showed more myocardial cells with spontaneous arrhythmogenic Ca2+ transient events compared to WT. The whole-cell patch-clamp technique applied to ventricular cardiomyocytes revealed comparable peak potassium channel current densities between KO and WT. These findings support the assumption that a decrease or even the loss of PP2A-B56α leads to an increased propensity of triggered arrhythmias. This could be based on the increased spontaneous Ca2+ tansients observed.

#1621 Quantifying ureter smooth muscle electrophysiology from calcium transient images to understand abnormal peristaltic contraction

Abstract Background and Aims Abnormal peristaltic contraction of the ureter smooth muscle (USM) causes acute kidney stone episodes. Intracellular electrical activities like membrane depolarization and action potentials play important roles in modulating the USM contraction by releasing intracellular calcium from the sarcoplasmic reticulum. Therefore, an electrophysiological study will help to assess the USM cell's electrical activities and in diagnosing abnormal USM contraction. The objective of this study is to quantify the contribution of ionic currents in shaping experimental calcium transient profiles using in-silico electrophysiological modeling. Method The simultaneous experimental recording of action potential (AP) and intracellular calcium transient images from the mouse ureter is obtained. The single isolated USM cell model comprises several voltage-gated ion channels, such as two voltage-gated calcium (T—type, and L—type) channels, one voltage-gated fast potassium (KA) channel, one calcium-dependent large conductance potassium channel, and an HCN channel. To describe the calcium-dependent gating of Ca2+-dependent potassium channels and to update the equilibrium potential of the Ca2+ ion, the intracellular Ca2+ concentration is updated during the simulation period. Results Simulation of simultaneous recordings of AP and cytosolic calcium [Ca2+]i are done on a single isolated cell. The model shows [Ca2+]i as a function of synaptic input-induced AP to simulate extracted experimental data, where Ca2+ transient is recorded simultaneously during AP in mouse USM cells. Fig. 1 shows both experimental and simulation of AP (A) and Calcium transient (B) in the USM cell. In our model, the radius “r” and time constant τ of the shell influence the Ca2+ transient profile. In the USM cell model, the submembrane calcium transient occurs from a depth of 0.1 μm to a depth of 0.6 μm. We have investigated whether Ca2+ current via the L-type Ca2+ channel is responsible for the firing of APs with fast upstroke generation. The AP and calcium transients are demolished with the absence of the L-type Ca2+ channel. Conclusion From this study, it is found that inhibition of the L-type Ca2+ channel not only prevented AP generation, it also reduced the cytosolic Ca2+ transient. This study supports the application of L-type Ca2+ channel inhibitor as a potential drug for abnormal peristaltic contraction of the ureter smooth muscle.

Atypical Antiadrenergic Effect of Refralon as a Mechanism of High Antiarrhythmic Effectiveness.

We studied the effect of Refralon on the electrophysiological properties of the supraventricular myocardium against the background of adrenergic (epinephrine) influence in the zone of the pulmonary veins, the area where 50-90% of atrial arrhythmias is triggered. The experiments were carried out on isolated tissue preparations of Wistar rats. The multichannel microelectrode array technique was used to record action potentials simultaneously in the atrium and in the ostium and distal parts of the pulmonary veins. Epinephrine application (12-50 nM) led to depolarization of the resting potential and the conduction block in the distal part of the pulmonary veins. Refralon (30 μg/kg) restored the resting potential in the distal part of the pulmonary veins. Against the background of epinephrine, Refralon did not significantly change the duration of the action potential at 90% repolarization in comparison with control. At the same time, the comparison drug E-4031 against the background of epinephrine significantly increased the duration of action potential in the atrium and in the ostium of the pulmonary veins, and sotalol increased it only in the ostium. Neither E-4031, nor sotalol restored conduction in their distal part. Refralon has a biphasic effect under conditions of adrenergic stimulation: the fast component is responsible for stabilizing the resting potential in the pulmonary vein and reduces the dispersion of action potential duration in the atrium and pulmonary vein and is also quickly washed away, and the slow component is responsible for the increase of the action potential duration and is slowly washed away.