Electrical Burst Research Articles

SAT061 Deficiency Of ENaC Excites Pancreatic β Cells To Promote Insulin Secretion

Abstract Disclosure: Y. Que: None. X. Ma: None. Y. Wu: None. J. Chen: None. J. Guo: None. Y. Ruan: None. In response to glucose elevation or other physiological stimuli, pancreatic islet β cells are excited to mobilize intracellular Ca2+ leading to insulin secretion, which is a complex cellular event with underlying mechanisms not fully elucidated. The present study explored possible involvement of the epithelial Na+ channel (ENaC) in β cell excitability and insulin secretion. Analyzing human databases, primary rat/mouse pancreatic tissues as well as RINm5F, a rat β-cell line, we confirmed the expression of Scnn1a, Scnn1b and Scnn1g genes (encoding ENaC subunits, α, β and γ, respectively) in human and rodent β cells. To our surprise, inhibiting this Na+ channel by selective blockers, amiloride (1-10 µM) or benzamil (1 µM), did not retard insulin secretion, but instead triggered a slow membrane depolarization with electrical bursts (41.5 ± 5.9 mV, measured by patch-clamp, n = 5-6), elicited substantial Ca2+ oscillations (135.2 ± 2.5% of baseline, by Fura-2 imaging, n = 256-380) and promoted insulin secretion (158.2 ± 25.6% of control, by ELISA, t-test, p < 0.05, n = 6) in RINm5F or isolated mouse β cells. siRNA-based knockdown of ENaCα, the rate-limiting subunit of ENaC, in RINm5F cells confirmed that deficiency of ENaC induced a significant increase in insulin secretion (230.2 ± 20.2% of control, t-test, p < 0.001, n = 6). Proteomic analysis of RINm5F cells (n = 5) through mass spectrometry showed that signaling pathways key to glucose metabolism and insulin secretion were significantly activated in RINm5F cells with ENaC knockdown in comparison with control cells, consistently suggesting a role of ENaC deficiency in exciting β cells to release insulin. We next built a mouse model with β cell-specific knockout of ENaCα (Scnn1afl/fl, Ins1-Cre+), which exhibited disturbed responses in glucose tolerance test in comparison with the loxp-negative Cre control mice (Scnn1awt/wt, Ins1-Cre+). Taken together, these results have suggested an important role of ENaC in regulating the excitability of β cells and insulin secretion, which may contribute to the understanding of Na+ environment in relation to insulin homeostasis. This work was supported in part by National Natural Science Foundation of China (82071599), Areas of Excellence Scheme of Hong Kong (AoE/M-402/20) and Bai Cheng Bai Yuan Start-up Fund (I2022A008). Presentation: Saturday, June 17, 2023

Recovery kinetics of short-term depression of GABAergic and glutamatergic synapses at layer 2/3 pyramidal cells in the mouse barrel cortex.

Short-term synaptic plasticity (STP) is a widespread mechanism underlying activity-dependent modifications of cortical networks. To investigate how STP influences excitatory and inhibitory synapses in layer 2/3 of mouse barrel cortex, we combined whole-cell patch-clamp recordings from visually identified pyramidal neurons (PyrN) and parvalbumin-positive interneurons (PV-IN) of cortical layer 2/3 in acute slices with electrical stimulation of afferent fibers in layer 4 and optogenetic activation of PV-IN. These experiments revealed that electrical burst stimulation (10 pulses at 10 Hz) of layer 4 afferents to layer 2/3 neurons induced comparable short-term depression (STD) of glutamatergic postsynaptic currents (PSCs) in PyrN and in PV-IN, while disynaptic GABAergic PSCs in PyrN showed a stronger depression. Burst-induced depression of glutamatergic PSCs decayed within <4 s, while the decay of GABAergic PSCs required >11 s. Optogenetically-induced GABAergic PSCs in PyrN also demonstrated STD after burst stimulation, with a decay of >11 s. Excitatory postsynaptic potentials (EPSPs) in PyrN were unaffected after electrical burst stimulation, while a selective optogenetic STD of GABAergic synapses caused a transient increase of electrically evoked EPSPs in PyrN. In summary, these results demonstrate substantial short-term plasticity at all synapses investigated and suggest that the prominent STD observed in GABAergic synapses can moderate the functional efficacy of glutamatergic STD after repetitive synaptic stimulations. This mechanism may contribute to a reliable information flow toward the integrative layer 2/3 for complex time-varying sensory stimuli.

A new diagnostic definition for patients with lower urinary tract symptoms evaluated by corpus cavernosum electromyography: 'Pelvic autonomic dysfunction'.

This study investigates the use of corpus cavernosum electromyography (CC-EMG) recording as a diagnostic tool for identifying the presence of pelvic autonomic dysfunction (PAD) in patients with LUTS and concurrent ED. Fifty patients were included in the study, and the electromyographic and urodynamic findings were compared. The 50% relaxation degree was accepted as a threshold value for PAD. According to relaxation degree in CC-EMG recordings, patients were grouped as; Group 1 (with PAD)=23 patients with a relaxation degree of less than 50%. Group 2 (without PAD)=27 patients with a relaxation degree greater than 50%. The mean age of the patients was 58 ± 3.2 (50-71) years. The comparison of the urodynamic parameters and transrectal ultrasound revealed that the postvoiding residual urine volume (PVR) (p=0.0007), P det Qmax (p=0.0005), and P det compliance (0.003) values were statistically significantly lower in Group 2. The mean IIEF-5 and IIEF-15 scores of the patients in Group 1 were 2.5 ± 0.7 and 8.1 ± 1.1 respectively. In Group 2, mean IIEF-5 and IIEF-15 scores were 7.2 ± 2.9, and 17.2 ± 3.5 (p: 0.00023 and p: 0.0009). The mean I-PSS score was 22.8 ± 3 in Group 1 and 16 ± 2.3 in Group 2 (p=0.001). The mean relaxation degree (RD) in Group I was 30.73 ± 6.8%, and in Group 2 was 66.3 ± 5.1%. The mean amplitude values of the patients in Group I were 261.41 ± 112.97 before papaverine injection, and in Group 2 were 246 ± 101.28 respectively. The inter-group difference was statistically significant (p < 0.05). The mean amplitude value of the patients in Group I after papaverine injection was 182.73 ± 60.71, and in Group 2, that value was 83.2 ± 29.19. The inter-group difference was also significant (p < 0.05). Increased electrical bursts and decreased relaxation responses on CC-EMG indirectly indicated an increase in the contractility of the bladder sphincter. Therefore, we concluded that CC-EMG could be a diagnostic tool for PAD and cavernous autonomic dysfunction.

Stereospecific antiseizure activity in mouse and rat epilepsy models by a pyridinium inhibitor of TNFα/NFκB signaling

Epilepsy affects over 50 million people worldwide and increases the risk of death. An intrinsic state of central inflammation, mainly driven by TNFα/NFκB signaling, may contribute to the refractory nature of some epilepsies. We have therefore hypothesized that inhibitors of this signaling pathway might be therapeutic. To test this hypothesis, we have measured the antiseizure properties of the enantiomeric compounds MRS-2481 and MRS-2485 in rodent seizure model systems. In the 6 ​Hz (44 ​mA) induced seizure test in mice, the (S) species, MRS-2485, was found to have higher protective potency and lower toxicity than the (R) species MRS-2481. However, neither of these enantiomers were protective in the MES-induced seizure test. MRS-2485 was also found to be protective in the corneal kindled mouse test. Finally, MRS-2485 reduced the post-kainate rat hippocampal slice electrical burst rate and duration. We conclude that MRS-2485, the (S)-enantiomer, is a potent inhibitor of seizure activity in mouse and rat models of epilepsy.

Oscillations in K(ATP) conductance drive slow calcium oscillations in pancreatic β-cells

ATP-sensitive K+ (K(ATP)) channels were first reported in the β-cells of pancreatic islets in 1984, and it was soon established that they are the primary means by which the blood glucose level is transduced to cellular electrical activity and consequently insulin secretion. However, the role that the K(ATP) channels play in driving the bursting electrical activity of islet β-cells, which drives pulsatile insulin secretion, remains unclear. One difficulty is that bursting is abolished when several different ion channel types are blocked pharmacologically or genetically, making it challenging to distinguish causation from correlation. Here, we demonstrate a means for determining whether activity-dependent oscillations in K(ATP) conductance play the primary role in driving electrical bursting in β-cells. We use mathematical models to predict that if K(ATP) is the driver, then contrary to intuition, the mean, peak, and nadir levels of ATP/ADP should be invariant to changes in glucose within the concentration range that supports bursting. We test this in islets using Perceval-HR to image oscillations in ATP/ADP. We find that mean, peak, and nadir levels are indeed approximately invariant, supporting the hypothesis that oscillations in K(ATP) conductance are the main drivers of the slow bursting oscillations typically seen at stimulatory glucose levels in mouse islets. In conclusion, we provide, for the first time to our knowledge, causal evidence for the role of K(ATP) channels not only as the primary target for glucose regulation but also for their role in driving bursting electrical activity and pulsatile insulin secretion.

Chronic stress facilitates bursting electrical activity in pituitary corticotrophs.

Coordination of an appropriate stress response is dependent upon anterior pituitary corticotroph excitability in response to hypothalamic secretagogues and glucocorticoid negative feedback. A key determinant of corticotroph excitability is large conductance calcium- and voltage-activated (BK) potassium channels that are critical for promoting corticotrophin-releasing hormone (CRH)-induced bursting that enhances adrenocorticotrophic hormone secretion. Previous studies revealed hypothalamic-pituitary-adrenal axis hyperexcitability following chronic stress (CS) is partly a function of increased corticotroph output. Thus, we hypothesise that chronic stress promotes corticotroph excitability through a BK-dependent mechanism. Corticotrophs from CS mice displayed significant increase in spontaneous bursting, which was suppressed by the BK blocker paxilline. Mathematical modelling reveals that the time constant of BK channel activation, plus properties and proportion of BK channels functionally coupled to L-type Ca2+ channels determines bursting activity. Surprisingly, CS corticotrophs (but not unstressed) display CRH-induced bursting even when the majority of BK channels are inhibited by paxilline, which modelling suggests is a consequence of the stochastic behaviour of a small number of BK channels coupled to L-type Ca2+ channels. Our data reveal that changes in the stochastic behaviour of a small number of BK channels can finely tune corticotroph excitability through stress-induced changes in BK channel properties. Importantly, regulation of BK channel function is highly context dependent allowing dynamic control of corticotroph excitability over a large range of time domains and physiological challenges in health and disease. This is likely to occur in other BK-expressing endocrine cells, with important implications for the physiological processes they regulate and the potential for therapy. KEY POINTS: Chronic stress (CS) is predicted to modify the electrical excitability of anterior pituitary corticotrophs. Electrophysiological recordings from isolated corticotrophs from CS male mice display spontaneous electrical bursting behaviour compared to the tonic spiking behaviour of unstressed corticotrophs. The increased spontaneous bursting from CS corticotrophs is BK-dependent and mathematical modelling reveals that the time constant of activation, properties and proportion of BK channels functionally coupled to L-type calcium channels determines the promotion of bursting activity. CS (but not unstressed) corticotrophs display corticotrophin-releasing hormone-induced bursting even when the majority of BK channels are pharmacologically inhibited, which can be explained by the stochastic behaviour of a small number of BK channels with distinct properties. Corticotroph excitability can be finely tuned by the stochastic behaviour of a small number of BK channels dependent on their properties and functional co-localisation with L-type calcium channels to control corticotroph excitability over diverse time domains and physiological challenges.

Fast-slow analysis demonstrates mechanism for stochastic endocrine cell electrical bursts

Findings in a mathematical model for cells in the pituitary gland highlight the role of certain potassium channels in electrical activity and hormone release.

Revisiting brain stimulation in Parkinson's disease.

Electrical burst stimulation rescues movement effectively in parkinsonian mice.

Fast-slow analysis of a stochastic mechanism for electrical bursting.

Electrical bursting oscillations in neurons and endocrine cells are activity patterns that facilitate the secretion of neurotransmitters and hormones and have been the focus of study for several decades. Mathematical modeling has been an extremely useful tool in this effort, and the use of fast-slow analysis has made it possible to understand bursting from a dynamic perspective and to make testable predictions about changes in system parameters or the cellular environment. It is typically the case that the electrical impulses that occur during the active phase of a burst are due to stable limit cycles in the fast subsystem of equations or, in the case of so-called "pseudo-plateau bursting," canards that are induced by a folded node singularity. In this article, we show an entirely different mechanism for bursting that relies on stochastic opening and closing of a key ion channel. We demonstrate, using fast-slow analysis, how the short-lived stochastic channel openings can yield a much longer response in which single action potentials are converted into bursts of action potentials. Without this stochastic element, the system is incapable of bursting. This mechanism can describe stochastic bursting in pituitary corticotrophs, which are small cells that exhibit a great deal of noise as well as other pituitary cells, such as lactotrophs and somatotrophs that exhibit noisy bursts of electrical activity.

From decimeter-scale elevated ionic conductivity regions in the cloud to lightning initiation

In this work, we represent the lightning initiation scenario as a sequence of two transitions of discharge activity to progressively larger spatial scales: the first one is from small-scale avalanches to intermediate-scale streamers; and the second one is from streamers to the lightning seed. We postulate the existence of ion production centers in the cloud, whose occurrence is caused by electric field bursts accompanying hydrometeor collisions (or near collisions) in the turbulent thundercloud environment. When a new ion production center is created inside (fully or partially) the residual ion spot left behind by a previously established center, there is a cumulative effect in the increasing of ion concentration. As a result, the essentially non-conducting thundercloud becomes seeded by elevated ion-conductivity regions (EICRs) with spatial extent of 0.1–1 m and a lifetime of 1–10 s. The electric field on the surface of an EICR (due to its conductivity being at least 4 orders of magnitude higher than ambient) is a factor of 3 or more higher than ambient. For a maximum ambient electric field of 100 kV/m typically measured in thunderclouds, such field enhancement is sufficient for initiation of positive streamers and their propagation over distances of the order of decimeters, and this will be happening naturally, without any external agents (e.g., superenergetic cosmic ray particles) or extraordinary in-cloud conditions, such as very high potential differences or very large hydrometeors. Provided that each EICR generates at least one streamer during its lifetime, the streamers will form a 3D network, some parts of which will contain hot channel segments created via the cumulative heating and/or thermal-ionizational instability. These hot channel segments will polarize, interact with each other, and cluster, forming longer conducting structures in the cloud. When the ambient potential difference bridged by such a conducting structure exceeds 3 MV, we assume that the lightning seed, capable of self-sustained bidirectional extension, is formed.

Comparative Analysis of Cortical Auditory Evoked Potential in Cochlear Implant Users.

The primary goal of the study was to investigate electrical cortical auditory evoked potentials (eCAEPs) at maximum comfortable level (MCL) and 50% MCL on three cochlear implant (CI) electrodes and compare them with the acoustic CAEP (aCAEPs), in terms of the amplitude and latency of the P1-N1-P2 complex. This was achieved by comparing the eCAEP obtained with the method described and stimulating single electrodes, via the fitting software spanning the cochlear array and the aCAEP obtained using the HEARLab system at four speech tokens. Twenty MED-EL (MED-EL Medical Electronics, Innsbruck, Austria) CI adult users were tested. CAEP recording with HEARLab System was performed with speech tokens /m/, /g/, /t/, and /s/ in free field, presented at 55 dB SPL. eCAEPs were recorded with an Evoked Potential device triggered from the MAX Programming Interface (MED-EL Medical Devices) with 70 msec electrical burst at 0.9 Hz at the apical (1), middle (6), and basal (10 or 11) CI electrode at their MCL and 50% MCL. CAEP responses were recorded in 100% of the test subjects for the speech token /t/, 95% for the speech tokens /g/ and /s/, and 90% for the speech token /m/. For eCAEP recordings, in all subjects, it was possible to identify N1 and P2 peaks when stimulating the apical and middle electrodes. This incidence of detection decreased to an 85% chance of stimulation at 50% MCL on the same electrodes. A P1 peak was less evident for all electrodes. There was an overall increase in latency for stimulation at 50% MCL compared with MCL. There was a significant difference in the amplitude of adjacent peaks (P1-N1 and N1-P2) for 50% MCL compared with MCL. The mean of the maximum cross-correlation values were in the range of 0.63 to 0.68 for the four speech tokens. The distribution of the calculated time shift, where the maximum of the cross-correlation was found, was distributed between the speech tokens. The speech token /g/ had the highest number of valid cross-correlations, while the speech token /s/ had the lowest number. This study successfully compared aCAEP and eCAEP in CI users. Both acoustic and electrical P1-N1-P2 recordings obtained were clear and reliable, with good correlation. Latency increased with decreasing stimulation level, while amplitude decreased. eCAEP is potentially a better option to verify speech detection at the cortical level because it (1) uses direct stimulation and therefore creates less interference and delay of the sound processor and (2) creates more flexibility with the recording setup and stimulation setting. As such, eCAEP is an alternative method for CI optimization.

Bursting oscillations in a piecewise system with time delay under periodic excitation: Theoretical and experimental observation of real electrical bursting signals using microcontroller

Bursting oscillations in a piecewise system with time delay under periodic excitation: Theoretical and experimental observation of real electrical bursting signals using microcontroller

Pulsed radiofrequency for chronic pain: In vitro evidence of an electroporation mediated calcium uptake

Pulsed radiofrequency (PRF) treatments for chronic pain consist in the delivery of a train of sinusoidal electric bursts to the targeted nerve. Despite numerous clinical evidence of its efficiency, the mechanism of action of PRF remains unclear. Since most of the reported biological effects of PRF can be initiated by a calcium influx into the neurons, we hypothesized that PRF may induce a mild electroporation effect causing a calcium uptake. To test this hypothesis, HEK-293 cells were exposed to PRF bursts and cytosolic calcium and Yo-Pro-1 uptake were monitored. After a single burst, calcium peaks were observed for electric fields above 480 V/cm while the uptake of Yo-pro-1 was insignificant. After a train of 120 bursts, the electric fields required to induce a calcium and Yo-pro-1 uptake decreased to 330 V/cm and 880 V/cm respectively. Calcium peaks were not detected when cells were treated in calcium free media. The temperature increase during the treatments was lower than 5 °C in all cases. Finally, the cell response for different burst frequencies and extracellular media conductivities correlated with the induced transmembrane voltage calculated with a numerical model. Our results support the hypothesis of an electroporation mediated calcium influx.

Prenatal exposure to nicotine disrupts synaptic network formation by inhibiting spontaneous correlated wave activity

Correlated spontaneous activity propagating over a wide region of the central nervous system is expressed during a specific period of embryonic development. We previously demonstrated using an optical imaging technique with a voltage-sensitive dye that this wave-like activity, which we referred to as the depolarization wave, is fundamentally involved in the early process of synaptic network formation. We found that the in ovo application of bicuculline/strychnine or d-tubocurarine, which blocked the neurotransmitters mediating the wave, significantly reduced functional synaptic expression in the brainstem sensory nucleus. This result, particularly for d-tubocurarine, an antagonist of nicotinic acetylcholine receptors, suggested that prenatal nicotine exposure associated with maternal smoking affects the development of neural circuit formation by interfering with the correlated wave. In the present study, we tested this hypothesis by examining the effects of nicotine on the correlated activity and assessing the chronic action of nicotine in ovo on functional synaptic expression along the vagal sensory pathway. In ovo observations of chick embryo behavior and electrical recording using in vitro preparations showed that the application of nicotine transiently increased embryonic movements and electrical bursts associated with the wave, but subsequently inhibited these activities, suggesting that the dominant action of the drug was to inhibit the wave. Optical imaging with the voltage-sensitive dye showed that the chronic exposure to nicotine in ovo markedly reduced functional synaptic expression in the higher-order sensory nucleus of the vagus nerve, the parabrachial nucleus. The results suggest that prenatal nicotine exposure disrupts the initial formation of the neural circuitry by inhibiting correlated spontaneous wave activity.

Challenging the challenge: A randomized controlled trial evaluating the inflammatory response and pain perception of healthy volunteers after single-dose LPS administration, as a potential model for inflammatory pain in early-phase drug development

Background and aimsFollowing an infection, cytokines not only regulate the acute immune response, but also contribute to symptoms such as inflammatory hyperalgesia. We aimed to characterize the acute inflammatory response induced by a human endotoxemia model, and its effect on pain perception using evoked pain tests in two different dose levels. We also attempted to determine whether combining a human endotoxemia challenge with measurement of pain thresholds in healthy subjects could serve as a model to study drug effects on inflammatory pain. Methods and resultsThis was a placebo-controlled, randomized, cross-over study in 24 healthy males. Twelve subjects were administered a bolus of 1 ng/kg LPS intravenously, and twelve 2 ng/kg LPS. Before days of placebo/LPS administration, subjects completed a full study day without study drug administration, but with identical pain threshold testing. Blood sampling and evoked pain tests (electrical burst and -stair, heat, pressure, and cold pressor test) were performed pre-dose and at frequent intervals up to 10hr post-dose. Data were analysed with a repeated-measures ANCOVA. For both dose levels, LPS induced an evident acute inflammatory response, but did not significantly affect any of the pain modalities. In a post-hoc analysis, lowering of pain thresholds was observed in the first 3 h after dosing, corresponding with the peak of the acute inflammatory response around 1–3 h post-dose. ConclusionMild acute systemic inflammation, as induced by 1 ng/kg and 2 ng/kg LPS intravenous administration, did not significantly change pain thresholds in this study. The endotoxemia model in combination with evoked pain tests is not suitable to study acute inflammatory hyperalgesia in healthy males.

Cardiac overexpression of perilipin 2 induces atrial steatosis, connexin 43 remodeling, and atrial fibrillation in aged mice

Atrial fibrillation (AF) is prevalent in patients with obesity and diabetes, and such patients often exhibit cardiac steatosis. Since the role of cardiac steatosis per se in the induction of AF has not been elucidated, the present study was designed to explore the relation between cardiac steatosis and AF. Transgenic (Tg) mice with cardiac-specific overexpression of perilipin 2 (PLIN2) were housed in the laboratory for more than 12 mo before the study. Electron microscopy of the atria of PLIN2-Tg mice showed accumulation of small lipid droplets around mitochondrial chains, and five- to ninefold greater atrial triacylglycerol (TAG) content compared with wild-type (WT) mice. Electrocardiography showed significantly longer RR intervals in PLIN2-Tg mice than in WT mice. Transesophageal electrical burst pacing resulted in significantly higher prevalence of sustained (>5 min) AF (69%) in PLIN2-Tg mice than in WT mice (24%), although it was comparable in younger (4-mo-old) mice. Connexin 43 (Cx43), a gap junction protein, was localized at the intercalated disks in WT atria but was heterogeneously distributed on the lateral side of cardiomyocytes in PLIN2-Tg atria. Langendorff-perfused hearts using the optical mapping technique showed slower and heterogeneous impulse propagation in PLIN2-Tg atria compared with WT atria. Cardiac overexpression of hormone-sensitive lipase in PLIN2-Tg mice resulted in atrial TAG depletion and amelioration of AF susceptibility. The results suggest that PLIN2-induced steatosis is associated with Cx43 remodeling, impaired conduction propagation, and higher incidence of AF in aged mice. Therapies targeting cardiac steatosis could be potentially beneficial against AF in patients with obesity or diabetes.

Chronic intermittent tachypacing by an optogenetic approach induces arrhythmia vulnerability in human engineered heart tissue.

AimsChronic tachypacing is commonly used in animals to induce cardiac dysfunction and to study mechanisms of heart failure and arrhythmogenesis. Human induced pluripotent stem cells (hiPSC) may replace animal models to overcome species differences and ethical problems. Here, 3D engineered heart tissue (EHT) was used to investigate the effect of chronic tachypacing on hiPSC-cardiomyocytes (hiPSC-CMs).Methods and resultsTo avoid cell toxicity by electrical pacing, we developed an optogenetic approach. EHTs were transduced with lentivirus expressing channelrhodopsin-2 (H134R) and stimulated by 15 s bursts of blue light pulses (0.3 mW/mm2, 30 ms, 3 Hz) separated by 15 s without pacing for 3 weeks. Chronic optical tachypacing did not affect contractile peak force, but induced faster contraction kinetics, shorter action potentials, and shorter effective refractory periods. This electrical remodelling increased vulnerability to tachycardia episodes upon electrical burst pacing. Lower calsequestrin 2 protein levels, faster diastolic depolarization (DD) and efficacy of JTV-519 (46% at 1 µmol/L) to terminate tachycardia indicate alterations of Ca2+ handling being part of the underlying mechanism. However, other antiarrhythmic compounds like flecainide (69% at 1 µmol/L) and E-4031 (100% at 1 µmol/L) were also effective, but not ivabradine (1 µmol/L) or SEA0400 (10 µmol/L).ConclusionWe demonstrated a high vulnerability to tachycardia of optically tachypaced hiPSC-CMs in EHT and the effective termination by ryanodine receptor stabilization, sodium or hERG potassium channel inhibition. This new model might serve as a preclinical tool to test antiarrhythmic drugs increasing the insight in treating ventricular tachycardia.

P1595Increased in vivo perpetuation of whole-heart ventricular arrhythmia in heterozygous Na+/Ca2+-exchanger knockout mice

Abstract Background Commonly, innovative antiarrhythmic strategies are derived from single cell studies that frequently yield promising in vitro findings. However, these results may differ on the whole-heart level, since multicellular electrophysiology is characterized by several emergent features. In previous cellular studies, we have identified the Na+/Ca2+-exchanger (NCX) as a promising target for an innovative antiarrhythmic strategy, as NCX upregulation is present in major cardiac diseases (e.g. heart failure) and promotes independently cellular early and late afterdepolarizations (EADs and DADs). Vice versa, we found that genetic and pharmacological NCX inhibition protects against EADs and DADs. To date, it is unknown, whether the concept of NCX inhibition indeed beneficially applies to the whole-heart level. Thus, we here investigate the in vivo inducibility and perpetuation of whole-heart arrhythmia using a heterozygous NCX-knockout mouse (KO) model that is protected against EADs and DADs on the cellular level. Methods/Results Programmed electrical right ventricular stimulation (PVS) and burst stimulation were performed in KO (n=22) and wild-type (n=34) mice by an octapolar mouse electrophysiological catheter introduced via the right jugular vein. Inducibility for ventricular tachycardia (VT) during PVS was similar in WT (73.5%) compared to KO (90.0%) (p=0.1707). With burst stimulation, VT inducibility was higher in KO (KO: 68.2%; WT: 32.4%; p=0.0134). During PVS, KO exhibited increased VT perpetuation as reflected in a significantly prolonged mean (in s; KO: 0.89±0.93; WT: 0.39±0.41; p=0.0097) and cumulative VT duration (in s; KO: 19.54±27.98; WT: 4.46±6.35; p=0.0019). Analysis of animals that were inducible for VT consistently yielded similar results. The ventricular refractory period (VRP) (in ms; KO: 15.1±3.5; WT: 18.7±4.1; p=0.0050) and the QTc interval were shortened in KO (in ms; KO: 46.5±5.8; WT: 53.2±5.9; p=0.0001). Conclusions As opposed to findings on the single cell level, KO mice exhibited an increased in vivo arrhythmia burden on the whole-heart level during PVS. This mainly resulted from increased perpetuation of artificially induced VTs, since the inducibility of VTs was not significantly increased in KO with PVS. As a mechanistic explanation of these surprising results, we found significantly reduced VRP and QTc durations in KO in line with the previously demonstrated action potential shortening in single KO cardiomyocytes, which promotes the perpetuation of VTs. We conclude that genetic NCX inhibition can protect from proarrhythmic cellular triggers like EADs and DADs that can initiate VT. However, VTs may perpetuate longer in KO most likely due to reduced refractory periods. This finding carries important translational limitations for the antiarrhythmic concept of NCX inhibition and demonstrates that the value of novel innovative strategies needs evaluation on both the cellular and the whole-heart level. Acknowledgement/Funding None

P6593Comparison of the termination effects between multi-segment continuous illumination and anti-tachycardia pacing on myocardial infarction ventricular arrhythmia

Abstract Introduction Anti-tachycardia pacing (ATP) therapy is an important treatment for current ICD termination of ventricular arrhythmia (VA). But there was concern about excessive frequency and voltage can cause arrhythmia, pain and damage. Optogenetic allowed modulation of electrical excitation with light in myocardium over-expressing light-sensitive protein, and optical illumination developed a non-contact and pain-free approach to research and treatment of cardiac disorder. Purpose Explore the effects of optogenetic termination and electric termination of myocardial infarction VA in vivo. Methods Rats (N=10) accepted jugular vein injection with 1×1012 vector genomes of (AAV-CAG-hChR2(H134R)-mcherry) to achieve cardiac transgenic expression of light sensitive protein Channelrhodopsin-2 (ChR2). After thoracotomy, myocardial infarction was induced by ligation of the left anterior descending coronary artery, and then VA was induced by electrical burst stimulation (10 v, 50 Hz, 2 s). Before the myocardial infarction operation, every heart was illuminated by 473 nm laser or electrical stimulated on the right ventricle in a train of 30 pulses at 8 HZ to test the threshold of light intensity or electrical pacing voltage. The VA termination process during 20 s after burst stimulation was investigated in different performance: optical termination, 1 s constant illumination repeated in 4 episodes with 1 s interval (470 nm, 20 times threshold intensity); electrical termination, ATP in 8 pulses of 8hz with 2ms duration in 4 times pacing threshold; natural recovery from VA, without optical or electrical intervention. Recovery time was defined as the time from the end of the burst stimulation to the recovery of sinus rhythm, and the termination rate was the percentage of sinus rhythm recovery among 11s after the end of burst stimulation with or without any intervention. Results We analyzed the recovery time and the termination rate (N=10, n=127 episodes per mode). The sequence of recovery time was optical termination (7.328±0.3623 s) &lt; electric termination (10.31±0.4482 s) &lt; natural recovery (12.97±0.3834 s) (figure D). We confirmed that the termination rate of optical illumination (86.14±4.145%) was higher than those of ATP (63.5±6.371%) and natural recovery (47.71±5.476%) (figure E), and there was no significant difference between the last two process (one-way of ANOVA; p=0.124). Comparison of three modes Conclusions Optogenetic manipulation shortened the recovery time and increased the termination rate in myocardial infarction ventricular arrhythmia, which may provide a potential approach for translational arrhythmia therapy. Acknowledgement/Funding The national natural science foundation of China (81772044)

P6595The optogenetic defibrillation of ventricular arrhythmia in myocardial infarction rats in vivo

Abstract Introduction Optogenetics is a low-invasive, flexible and highly selective intervention that enables electrical excitation with light on myocardium overexpressing light-sensitive proteins. Optical illumination can control the simultaneous exciting of the whole myocardium under the spot, which is more conducive to recovery from electrical disturbance to sinus rhythm. Purpose We explored optogenetic defibrillation for different illumination parameters how to affect defibrillation rates and the possible mechanism of continuous illumination defibrillation. Methods Systemic delivery via right jugular vein injection of (AAV9-CAG-hChR2(H134R)-mCherry) were performed in juvenile SD rats to achieve the light sensitive protein Channelrhodopsin-2 (ChR2) transfer throughout the whole heart. We intubated and ventilated rats, opened chest and recorded the ECG. After ligation of the left anterior descending coronary artery, ventricular arrhythmia was induced by electrical burst stimulation (10v, 50Hz, 2s). Cardiac epicardium illumination with 470nm blue laser was performed to investigate the effects of optogenetic defibrillation and its underlying mechanism. Every heart accepted 30 pulses of 20ms duration on 8Hz to test the light intensity threshold for 1:1 capture. Different illumination modes of multiple light intensity (2,4,8,10,20 times threshold intensity), pulse duration (20, 50, 200, 500 and 1000ms) and illumination position (RV apex, RV, RVOT, septum, LV) were applied in each attempt for 4 repetitions with 1 s interval. Results We demonstrated that ventricular arrhythmias could be terminated by illumination of the right ventricle at 20 times threshold intensity in 1s (figure A) with the successful defibrillation rate of 95±2.673% (mean ±SEM; N=7). Herein, the successful optogenetic defibrillation rate was strongly depending on light intensity (N=5, n=50 episodes, p=0.0118) and duration of illumination (N=5, n=50 episodes, p&lt;0.0001) (figure B.C). Notably when there were higher intensity and longer pulse duration, the higher defibrillation rate appeared. There was no significant difference in the defibrillation rate among different illumination positions (N=5, n=25episodes per position, p=0.1177) (figure D). To explore the underlying mechanism of optogenetic defibrillation, we performed the same illumination mode during sinus rhythm in 2 rats (figure E. F. G). We observed that higher light intensity and longer pulse duration were more conducive to induce an episode of higher frequency focal excitement. Views of optogenetic defibrillation Conclusions We demonstrated that optogenetic defibrillation is a highly effective intervention and the possible mechanism is partly attributed to overdrive suppression. We believe that optogenetic approach is potentially to be translated into more efficient and pain-free clinical termination of ventricular arrhythmia. Acknowledgement/Funding The national natural science foundation of China (81772044)