Ventricular Effective Refractory Period Research Articles

Defining cardiac stability during Langendorff perfusion

The Langendorff heart preparation is a valuable ex vivo technique for studying cardiac electrophysiology. For the past century it has been used in toxicology studies, drug testing, and disease modeling and has contributed significantly to understanding cardiac physiology, development, and pathological states. This study aimed to investigate the stability and reproducibility of the modern Langendorff heart preparation under various experimental conditions and to establish cardiac metric standards with corresponding exclusion criteria. Isolated hearts were extracted from adult Sprague Dawley rats and retrograde perfused with a modified Krebs-Henseleit buffer. A cohort of isolated hearts were used to establish a control baseline of multiple physiological parameters measured over time (e.g., electrocardiogram, flow rate, electrophysiology, electrical and calcium dynamics). These control baseline values were then compared to hearts exposed to the most common media additives in Langendorff studies (e.g., blebbistatin(−/−), bovine serum albumin, and voltage and calcium indicator dye). Over a three-hour period, cardiac metrics were monitored in stable Langendorff-perfused hearts. The addition of blebbistatin, DMSO or dyes did not change EP or ECG measurements as compared to control baseline. The most significant drift in all the EP values was ventricular effective refractory period (VERP). With or without blebbistatin, VERP increased from 40 to 60ms over the course of three hours. In that time heart rate and flow also decreased slightly. Blebbistatin was associated with an increased action potential duration but did not alter calcium transient duration. It was also observed that 1% circulating bovine serum albumin quenched the rh237 voltage dye signal 80% as measured by signal to noise. Constant pressure Langendorff-perfusion with a crystalloid buffer provides a reliable and reproducible model for cardiac electrophysiology research with a few caveats. The total length of study should be carefully considered to account for adaptations in cardiac physiology during longer perfusion times. As an excitation-contraction uncoupler, blebbistatin reduces motion artifact but delays action potential duration. Although BSA is used to minimize cardiac edema, it interferes with membrane bound voltage dyes. This study sets physiological parameters as a useful guide for future cardiac studies that use the isolated heart as a research model. American Physiological Society, R01HD108839 to N.G.P. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Atorvastatin, etanercept and the nephrogenic cardiac sympathetic remodeling in chronic renal failure rats.

Chronic renal failure (CRF) patients are predisposed to arrhythmias, while the detailed mechanisms are unclear. We hypothesized the chronic inflammatory state of CRF patients may lead to cardiac sympathetic remodeling, increasing the incidence of ventricular arrhythmia (VA) and sudden cardiac death. And explored the role of atorvastatin and etanercept in this process. A total of 48 rats were randomly divided into sham operation group (Sham group), CRF group, CRF + atorvastatin group (CRF + statin group), and CRF + etanercept group (CRF + rhTNFR-Fc group). Sympathetic nerve remodeling was assessed by immunofluorescence of growth-associated protein 43 (GAP-43) and tyrosine hydroxylase positive area fraction. Electrophysiological testing was performed to assess the incidence of VA by assessing the ventricular effective refractory period and ventricular fibrillation threshold. The levels of tumor necrosis factor-alpha (TNF-α) and interleukin-1beta were determined by Western blotting and enzyme-linked immunosorbent assay. Echocardiogram showed that compared with the Sham group, left ventricular end-systolic diameter and ventricular weight/body weight ratio were significantly higher in the CRF group. Hematoxylin-eosin and Masson staining indicated that myocardial fibers were broken, disordered, and fibrotic in the CRF group. Western blotting, enzyme-linked immunosorbent assay, immunofluorescence and electrophysiological examination suggested that compared with the Sham group, GAP-43 and TNF-α proteins were significantly upregulated, GAP-43 and tyrosine hydroxylase positive nerve fiber area was increased, and ventricular fibrillation threshold was significantly decreased in the CRF group. The above effects were inhibited in the CRF + statin group and the CRF + rhTNFR-Fc group. In CRF rats, TNF-α was upregulated, cardiac sympathetic remodeling was more severe, and the nephrogenic cardiac sympathetic remodeling existed. Atorvastatin and etanercept could downregulate the expression of TNF-α or inhibit its activity, thus inhibited the above effects, and reduced the occurrence of VA and sudden cardiac death.

Vericiguat suppresses ventricular tachyarrhythmias inducibility in a rabbit myocardial infarction model.

The VICTORIA trial demonstrated a significant decrease in cardiovascular events through vericiguat therapy. This study aimed to assess the potential mechanisms responsible for the reduction of cardiovascular events with vericiguat therapy in a rabbit model of myocardial infarction (MI). A chronic MI rabbit model was created through coronary artery ligation. Following 4 weeks, the hearts were harvested and Langendorff perfused. Subsequently, electrophysiological examinations and dual voltage-calcium optical mapping studies were conducted at baseline and after administration of vericiguat at a dose of 5 μmol/L. Acute vericiguat therapy demonstrated a significant reduction in premature ventricular beat burden and effectively suppressed ventricular arrhythmic inducibility. The electrophysiological influences of vericiguat therapy included an increased ventricular effective refractory period, prolonged action potential duration, and accelerated intracellular calcium (Cai) homeostasis, leading to the suppression of action potential and Cai alternans. The pacing-induced ventricular arrhythmias exhibited a reentrant pattern, attributed to fixed or functional conduction block in the peri-infarct zone. Vericiguat therapy effectively mitigated the formation of cardiac alternans as well as the development of reentrant impulses, providing additional anti-arrhythmic benefits. In the MI rabbit model, vericiguat therapy demonstrates anti-ventricular arrhythmia effects. The vericiguat therapy reduces ventricular ectopic beats, inhibiting the initiation of ventricular arrhythmias. Furthermore, the therapy successfully suppresses cardiac alternans, preventing conduction block and, consequently, the formation of reentry circuits.

Chronic Expression of Interleukin-10 Transgene Modulates Cardiac Sympathetic Ganglion Resulting in Reduced Ventricular Arrhythmia.

The cardiac autonomic nervous system (CANS) is intimately connected to the regulation of electrophysiology and arrhythmogenesis in cardiac systems. This work aimed at investigating whether interleukin-10 (IL-10) could effectively modulate CANS and suppress ischemia-induced ventricular arrhythmia (VA) through chronically acting on the cardiac sympathetic ganglion (CSG). Using an adeno-associated virus (AAV), we achieved local chronic overproduction of IL-10 in the CSG, left stellate ganglion (LSG). As a result, in the IL-10 group, we observed a decreased number of tyrosine hydroxylase-positive (TH+) cells in the LSG. IL-10 markedly downregulated the nerve growth factor, synaptophysin, as well as growth-associated protein 43 expression. In vivo, results from ambulatory electrocardiography showed that IL-10 overexpression significantly inhibited the cardiac sympathetic nervous system activity and improved heart rate variability. Meanwhile, we observed decreased LSG function as well as prolonged ventricular effective refractory period and suppressed VA after myocardial infarction (MI) in the IL-10 group. In addition, IL-10 overexpression attenuated inflammation and decreased norepinephrine levels in the myocardium after acute MI. In conclusion, our data suggest that chronic IL-10 overexpression modulates cardiac sympathetic nerve remodeling and suppresses VA induced by MI. Neuromodulation through AAV-mediated IL-10 overexpression may have the characteristics of and advantages as a potential neuroimmunotherapy for preventing MI-induced VAs.

Analysis of cardiohemodynamic and electrophysiological effects of morphine along with its toxicokinetic profile using the halothane-anesthetized dogs.

Although morphine has been used for treatment-resistant dyspnea in end-stage heart failure patients, information on its cardiovascular safety profile remains limited. Morphine was intravenously administered to halothane-anesthetized dogs (n=4) in doses of 0.1, 1 and 10 mg/kg/10 min with 20 min of observation period. The low and middle doses attained therapeutic (0.13 µg/mL) and supratherapeutic (0.97 µg/mL) plasma concentrations, respectively. The low dose hardly altered any of the cardiovascular variables except that the QT interval was prolonged for 10-15 min after its start of infusion. The middle dose reduced the preload and afterload to the left ventricle for 5-15 min, then decreased the left ventricular contractility and mean blood pressure for 10-30 min, and finally suppressed the heart rate for 15-30 min. Moreover, the middle dose gradually but progressively prolonged the atrioventricular conduction time, QT interval/QTcV, ventricular late repolarization period and ventricular effective refractory period without altering the intraventricular conduction time, ventricular early repolarization period or terminal repolarization period. A reverse-frequency-dependent delay of ventricular repolarization was confirmed. The high dose induced cardiohemodynamic collapse mainly due to vasodilation in the initial 2 animals by 1.9 and 3.3 min after its start of infusion, respectively, which needed circulatory support to treat. The high dose was not tested further in the remaining 2 animals. Thus, intravenously administered morphine exerts a rapidly appearing vasodilator action followed by slowly developing cardiosuppressive effects. Morphine can delay the ventricular repolarization possibly through IKr inhibition in vivo, but its potential to develop torsade de pointes will be small.

Targeted ablation of the left middle cervical ganglion prevents ventricular arrhythmias and cardiac injury induced by AMI.

Cardiac sympathetic overactivation is a critical driver in the progression of acute myocardial infarction (AMI). The left middle cervical ganglion (LMCG) is an important extracardiac sympathetic ganglion. However, the regulatory effects of LMCG on AMI have not yet been fully documented. In the present study, we detected that the LMCG was innervated by abundant sympathetic components and exerted an excitatory effect on the cardiac sympathetic nervous system in response to stimulation. In canine models of AMI, targeted ablation of LMCG reduced the sympathetic indexes of heart rate variability and serum norepinephrine, resulting in suppressed cardiac sympathetic activity. Moreover, LMCG ablation could improve ventricular electrophysiological stability, evidenced by the prolonged ventricular effective refractory period, elevated action potential duration, increased ventricular fibrillation threshold, and enhanced connexin43 expression, consequently showing antiarrhythmic effects. Additionally, compared with the control group, myocardial infarction size, circulating cardiac troponin I, and myocardial apoptosis were significantly reduced, accompanied by preserved cardiac function in canines subjected to LMCG ablation. Finally, we performed the left stellate ganglion (LSG) ablation and compared its effects with LMCG destruction. The results indicated that LMCG ablation prevented ventricular electrophysiological instability, cardiac sympathetic activation, and AMI-induced ventricular arrhythmias with similar efficiency as LSG denervation. In conclusion, this study demonstrated that LMCG ablation suppressed cardiac sympathetic activity, stabilized ventricular electrophysiological properties and mitigated cardiomyocyte death, resultantly preventing ischemia-induced ventricular arrhythmias, myocardial injury, and cardiac dysfunction. Neuromodulation therapytargeting LMCG represented a promising strategy for the treatment of AMI.

Effect of Flecainide and Ibutilide Alone and in Combination to Terminate and Prevent Recurrence of Atrial Fibrillation.

There is a need for improved approaches to rhythm control therapy of atrial fibrillation (AF). The effectiveness of flecainide (1.5 µmol/L) and ibutilide (20 nmol/L), alone and in combination, to cardiovert and prevent AF recurrence was studied in canine-isolated coronary-perfused right atrioventricular preparations. We also examined the safety of the combination of flecainide (1.5 µmol/L) and ibutilide (50 nmol/L) using canine left ventricular wedge preparations. Sustained AF (>1 hour) was inducible in 100%, 60%, 20%, and 0% of atria in the presence of acetylcholine alone, acetylcholine+ibutilide, acetylcholine+flecainide, and acetylcholine+ibutilide+flecainide, respectively. When used alone, flecainide and ibutilide cardioverted sustained AF in 40% and 20% of atria, respectively, but in 100% of atria when used in combination. Ibutilide prolonged atrial and ventricular effective refractory period by 15% and 8%, respectively, at a cycle length of 500 ms (P<0.05 for both). Flecainide increased the effective refractory period in atria by 27% (P<0.01) but by only 2% in the ventricles. The combination of the 2 drugs lengthened the effective refractory period by 42% in atria (P<0.01) but by only 7% (P<0.05) in the ventricles. In left ventricular wedges, ibutilide prolonged QT and Tpeak-Tend intervals by 25 and 55%, respectively (P<0.05 for both; cycle length, 2000 ms). The addition of flecainide (1.5 µmol/L) partially reversed these effects (P<0.05 for both parameters versus ibutilide alone). Torsades de Pointes score was relatively high with ibutilide alone and low with the drug combination. In our experimental model, a combination of flecainide and ibutilide significantly improves cardioversion and prevents the recurrence of AF compared with monotherapies with little to no risk for the development of long-QT-mediated ventricular proarrhythmia.

The benefit of a double ventricular extra stimuli protocol to improve functional substrate mapping during ablation of ventricular tachycardia. - The S3-VT-FREE project

Abstract Introduction Ablation of ventricular tachycardia (VT) based on substrate-based mapping has been proposed to itemize areas of interest identifying areas of low-voltage, late potentials (LPs), and deceleration zones (DZs). However, partial delineation of targets for ablation is likely responsible for a suboptimal long-term success rate. The abnormal electrograms (EGMs), featuring low-voltage near-field components may be hidden within the high-amplitude far-field signal. We hypothesized that the acquisition of functional mapping after double ventricular extrastimuli (S3 protocol) would uncover relevant areas of functional substrate that could be missed during sinus or S1 map. We aim to evaluate the potential benefit and the feasibility of functional substrate mapping using an S3 protocol for VT ablation and to assess its colocalization with detected corridors using late gadolinium enhancement magnetic resonance imaging (LGE-MRI). Methods We included consecutively 29 patients with structural cardiomypathy who underwent a VT ablation. The S3 protocol included: (1) assessment of the ventricular effective refractory period (ERP), (2) pacing protocol from the apex of the right ventricle with S1 x 6 (cycle length: 600ms) followed by S2 (ERP+30ms) and S3 (ERP+50ms). Electroanatomical mapping was acquired with the EnsiteX system (Abbott, IL) using HD Grid catheter and omnipolar signals. ILAMs were used to identify deceleration zones and areas of late potentials were identified for all maps. A non-invasive substrate assessment was done with LGE-MRI and post-processing with a commercially available system (ADAS3D, Spain) with automated identification of corridors. Results The S3 protocol was completed in 25 of the 29 procedures (86.2%). The repeated induction of ventricular arrhythmias did not allow for full protocol of the S3 protocol in 4 patients (13.8%). The functional substrate identified during the S3 activation mapping was significantly more extensive than the one identified using S1 and S2 activation mappings (p&amp;lt;0.05) (Table). The S3 protocol unmasked a higher number of DZs (p&amp;lt;0.01), a wider area of LPs (p&amp;lt;0.01), and a longer duration of the DZs (p=0.01). All DZ maps were compared with preprocedural cardiac magnetic resonance. The number of DZs matching with conduction corridors identified by LGE-MRI was higher in the activation mapping of S3 compared with the S1 and S2 ones (p=0.02). After VT ablation based on an S3 protocol, 88% of patients have been VT free during a median follow-up of 9 months. Conclusions This novel substrate-based protocol to assess the ventricular functional substrate is feasible in the majority of patients (86.2%) and allows for a better identification of targets for ablation and could facilitate VT ablation, and improve the prognosis of the patients after ablation.

Simultaneous assessment of mechanical and electrical function in Langendorff-perfused ex-vivo mouse hearts.

The Langendorff-perfused ex-vivo isolated heart model has been extensively used to study cardiac function for many years. However, electrical and mechanical function are often studied separately-despite growing proof of a complex electro-mechanical interaction in cardiac physiology and pathology. Therefore, we developed an isolated mouse heart perfusion system that allows simultaneous recording of electrical and mechanical function. Isolated mouse hearts were mounted on a Langendorff setup and electrical function was assessed via a pseudo-ECG and an octapolar catheter inserted in the right atrium and ventricle. Mechanical function was simultaneously assessed via a balloon inserted into the left ventricle coupled with pressure determination. Hearts were then submitted to an ischemia-reperfusion protocol. At baseline, heart rate, PR and QT intervals, intra-atrial and intra-ventricular conduction times, as well as ventricular effective refractory period, could be measured as parameters of cardiac electrical function. Left ventricular developed pressure (DP), left ventricular work (DP-heart rate product) and maximal velocities of contraction and relaxation were used to assess cardiac mechanical function. Cardiac arrhythmias were observed with episodes of bigeminy during which DP was significantly increased compared to that of sinus rhythm episodes. In addition, the extrasystole-triggered contraction was only 50% of that of sinus rhythm, recapitulating the "pulse deficit" phenomenon observed in bigeminy patients. After ischemia, the mechanical function significantly decreased and slowly recovered during reperfusion while most of the electrical parameters remained unchanged. Finally, the same electro-mechanical interaction during episodes of bigeminy at baseline was observed during reperfusion. Our modified Langendorff setup allows simultaneous recording of electrical and mechanical function on a beat-to-beat scale and can be used to study electro-mechanical interaction in isolated mouse hearts.

Evidence for the cardiodepressive effects of the plasticizer di-2-ethylhexyl phthalate.

Di-2-ethylhexyl phthalate (DEHP) is commonly used in the manufacturing of plastic materials, including intravenous bags, blood storage bags, and medical-grade tubing. DEHP can leach from plastic medical products, which can result in inadvertent patient exposure. DEHP concentrations were measured in red blood cell units stored between 7 and 42 days (17-119 μg/ml). Using these concentrations as a guide, Langendorff-perfused rat heart preparations were acutely exposed to DEHP. Sinus activity remained stable with lower doses of DEHP (25-50 μg/ml), but sinus rate declined by 43% and sinus node recovery time (SNRT) prolonged by 56.5% following 30-min exposure to 100 μg/ml DEHP. DEHP exposure also exerted a negative dromotropic response, as indicated by a 69.4% longer PR interval, 108.5% longer Wenckebach cycle length (WBCL), and increased incidence of atrioventricular (AV) uncoupling (60-min exposure). Pretreatment with doxycycline partially rescued the effects of DEHP on sinus activity, but did not ameliorate the effects on AV conduction. DEHP exposure also prolonged the ventricular action potential and effective refractory period, but had no measurable effect on intracellular calcium transient duration. Follow-up studies using human-induced pluripotent stem cell-derived cardiomyocytes confirmed that DEHP slows electrical conduction in a time (15 min-3 h) and dose-dependent manner (10-100 μg/ml). Previous studies have suggested that phthalate toxicity is specifically attributed to metabolites of DEHP, including mono-2-ethylhexylphthalate. This study demonstrates that DEHP exposure also contributes to cardiac dysfunction in a dose- and time-dependent manner. Future work is warranted to investigate the impact of DEHP (and its metabolites) on human health, with special consideration for clinical procedures that employ plastic materials.

Association of Sinus Wavefront Activation and Ventricular Extrastimuli Mapping With Ventricular Tachycardia Re-Entrant Circuits

BackgroundSubstrate-based ablation targets areas of delayed and fractionated electrograms during sinus rhythm, which are sensitive for identifying the ventricular tachycardia (VT) isthmus but is influenced by the activation wavefront direction and decremental pacing. ObjectivesThe aim of this study was to correlate the areas of latest activation during varying wavefront activation mapping and decremental pacing mapping with sites critical to the VT isthmus. MethodsThree high-density electroanatomical substrate maps were created in patients presenting for ablation of monomorphic VT: 1) native sinus rhythm; 2) right ventricular (RV) apical pacing; and 3) an RV apical S2 map following the S1 drive train at 20 ms above the ventricular effective refractory period. Areas corresponding to the latest activation were compared with the VT isthmus identified by conventional mapping. ResultsTwenty patients with structural heart disease with a mean age of 55.6 ± 16.9 years were included. The majority of the cohort consisted of patients with ischemic heart disease (50%) and arrhythmogenic RV cardiomyopathy (35%). Epicardial ablation was performed in 45% of patients. The concordance of the site of latest activation in sinus rhythm with the VT isthmus was 75%. The location of the latest activation during RV apical pacing corresponded with the VT isthmus in 85% of cases. However, in 95% of cases, the site of the latest activation following the S2 stimulus colocalized to the VT isthmus. ConclusionsIn a mix of underlying myocardial substrates, regions of conduction slowing during decremental pacing colocalize with the VT isthmus more frequently than sinus rhythm activation mapping and may have a role in substrate-based ablation where VT induction is undesirable.

Transient cardiac electrophysiological changes in a rat model of subarachnoid haemorrhage: a brain-heart interaction.

Subarachnoid haemorrhage (SAH) is one of the causes of sudden cardiac death (SCD). However, the time course of ventricular arrhythmias and potential mechanisms responsible for this effect after SAH remain unknown. This study aims to investigate the effect of SAH on ventricular electrophysiological changes and its potential mechanisms in long-term phase. We examined the ventricular electrophysiological remodelling and potential mechanisms in a Sprague Dawley rat model of SAH at six time points (baseline, and Days 1, 3, 7, 14 and 28) and explored the potential mechanisms. We measured the ventricular effective refractory period (ERP), ventricular fibrillation threshold (VFT) and left stellate ganglion (LSG) activity at different time points before and after SAH. We also detected neuropeptide Y (NPY) levels in plasma and myocardial tissues by enzyme-linked immunosorbent assay, and quantified NPY 1 receptor (NPY1R) protein and mRNA expression levels by western blotting and quantitative real-time reverse transcription-polymerase chain reaction, respectively. Subarachnoid haemorrhage gradually prolonged QTc intervals, shortened ventricular ERP and reduced VFT during the acute phase, peaking at Day 3. However, no significant changes were observed from Days 14 to 28 compared to Day 0. Subarachnoid haemorrhage gradually increased LSG activity, increased NPY concentrations and up-regulated NPY1R expression in the acute phase of SAH, peaking at Day 3. However, no significant variations were found from Days 14 to 28 compared to Day 0. Subarachnoid haemorrhage increases the transient susceptibility of VAs in the acute phase, and the underlying mechanisms for this response included increased sympathetic activity and up-regulated NPY1R expression.

Site-specific effects of dobutamine on cardiac conduction and refractoriness.

Isoproterenol, a non-specific beta agonist, is commonly used during electrophysiology studies (EPS). However, with the significant increase in the price of isoproterenol in 2015 and the increasing number of catheter ablations performed, the cost implications cannot be ignored. Dobutamine is a less expensive synthetic compound developed from isoproterenol with a similar mechanism to enhance cardiac conduction and shorten refractoriness, thus making it a feasible substitute with a lower cost. However, the use of dobutamine for EPS has not been well-reported in the literature. To determine the site-specific effects of various doses of dobutamine on cardiac conduction and refractoriness and assess its safety during EPS. From February 2020 to October 2020, 40 non-consecutive patients scheduled for elective EPS, supraventricular tachycardia, atrial fibrillation, and premature ventricular contraction ablations at a single center were consented and prospectively enrolled to assess the effect of dobutamine on the cardiac conduction system. At the end of each ablation procedure, measures of cardiac conduction and refractoriness were recorded at baseline and with incremental doses of dobutamine at 5, 10, 15, and 20 mcg/kg/min. For the primary analysis, the change per dose of dobutamine from baseline to each dosing level of dobutamine received by the patients, comparing atrioventricular node block cycle length (AVNBCL), ventricular atrial block cycle length (VABCL) and sinus cycle length (SCL), was tested using mixed-effect regression. For the secondary analysis, dobutamine dose level was tested for association with relative changes from baseline of each electrophysiologic parameter (SCL, AVNBCL, VABCL, atrioventricular node effective refractory period (AVNERP), AH, QRS, QT, QTc, atrial effective refractory period (AERP), ventricular effective refractory period (VERP), using mixed-effect regression. Changes in systolic and diastolic blood pressures were also assessed. The Holm-Bonferroni method was used to adjust for multiple testing. For the primary analysis there was no statistically significant change of AVNBCL and VABCL relative to SCL from baseline to each dose level of dobutamine. The SCL, AVNBCL, VABCL, AVNERP, AERP, VERP and the AH, and QT intervals all demonstrated a statistically significant decrease from baseline to at least one dose level with incremental dobutamine dosing. Two patients (5%) developed hypotension during the study and one patient (2.5%) received a vasopressor. Two patients (5%) had induced arrhythmias but otherwise no major adverse events were noted. In this study, there was no statistically significant change of AVNBCL and VABCL relative to SCL from baseline to any dose level of dobutamine. As expected, the AH and QT intervals, and the VABCL, VERP, AERP and AVNERP all significantly decreased from baseline to at least one dose level with an escalation in dobutamine dose. Dobutamine was well-tolerated and safe to use during EPS.

Optimisation of decrementing evoked potential mapping for functional substrate identification in ischaemic ventricular tachycardia ablation

Abstract Funding Acknowledgements Type of funding sources: None. Background Decrement evoked potential (DeEP) mapping has been shown to identify the functional substrate critical to the Ventricular Tachycardia (VT) circuit with a higher specificity than late potential mapping.[1] However, there is limited data on methodological optimisation of this technique which could identify more relevant regions of conduction delay and functional block that occurs during clinical VT by activating the substrate at coupling intervals closer to the VT cycle length (CL). Purpose The goal of this study was to investigate a range of short coupled extra-stimuli and decrement thresholds to improve DeEP mapping target identification in patients with ischaemic cardiomyopathy. Methods Thirteen patients (70±12 years) underwent VT ablation. Mapping was performed with the Advisor HD Grid multipolar catheter. Maps were generated using omnipolar electrograms (EGMs). DeEP mapping was performed using 3 different extra-stimuli (S2) sequentially in each case: (1) 400 ms (DeEP-400); (2) VT cycle length (VTCL); (3) ventricular effective refractory period (VERP) + 20 ms. For each of the DeEP-400, DeEP-VTCL and DeEP-VERP maps, the definition of a DeEP was altered from the standard 10 ms decrement to multiple thresholds of 10 ms to 50 ms in 10 ms increments (figure 1). DeEP maps were compared to the VT exit site identified with VT activation maps. All patients underwent DeEP-400 focused ablation. Results All patients had 1 clinical VT (VTCL = 353 ± 48 ms; VERP + 20 ms = 300 ± 20 ms). Mean left ventricular ejection fraction was 30 ± 12 %. Sensitivity and specificity analyses (figure 2) show the optimal threshold for defining a DeEP is with an EGM prolongation of ≥ 20 ms for the DeEP-400 (sens. 77%, spec. 79%, ROC AUC 0.84), DeEP-VTCL (sens. 77%, spec. 79%, ROC AUC 0.83) and DeEP-VERP (sens. 69%, spec. 74%, ROC AUC 0.76) maps. Proportional to total mapped area, the DeEP-400 map area was significantly smaller than DeEP-VERP map area (19.0 ± 7.6 % vs 26.4 ± 14.3%; p=0.02). Delivering the S2 at the conventional VERP + 20 ms using a 10 ms decrement definition yielded a sensitivity of 84.6% and a specificity of 62.0% with a target ablation area of 33.9 cm2. With the optimised protocol of delivering the S2 at 400 ms using a 20 ms decrement definition, the sensitivity remains high at 76.9% with an improved specificity of 79.0%, thus reducing the ablation target area by 50% to 16.8 cm2. 92% patients were free of any VT after DeEP-400 guided ablation at 6-month median follow-up. Conclusion In this prospective, mechanistic study utilising omnipolar technology to explore a range of S2 extra-stimuli and decrement thresholds, DeEPs were optimally defined by a conduction delay of ≥ 20 ms with an S2 extra-stimulus coupling interval of 400 ms or VTCL to more accurately localise the VT exit site. This optimised DeEP mapping protocol further refines identification of the critical myocardial sites involved in the initiation and maintenance of VT.

PO-01-148 COMBINATION OF FLECAINIDE AND IBUTILIDE FOR EFFECTIVE AND SAFE MANAGEMENT OF ATRIAL FIBRILLATION

There is a need for improved approaches to rhythm control therapy of atrial fibrillation (AF). To investigate a novel pharmacological approach for rhythm control management of AF. We studied the effectiveness of clinically relevant concentrations of flecainide (1.5 μM) and ibutilide (20 nM), alone and in combination, to prevent the induction of AF and to cardiovert persistent AF, defined here as AF lasting > 1 hr, using canine isolated coronary-perfused right atrial preparations. In addition, we examined the safety of the flecainide and ibutilide (50 nM) combination using canine left ventricular (LV) wedge preparations. Persistent AF could be induced in 100% of preparations following exposure to acetylcholine (0.5 μM; n=5). Persistent AF remained inducible in 60% of atria in the presence of ibutilide alone and in 20% of preparations in the presence of flecainide alone, but in 0% of preparations in the presence of the combination of ibutilide and flecainide (n=5 per group). When used alone, flecainide and ibutilide cardioverted persistent AF in 40% and 20% of atria, respectively, but in 100% of atria when used in combination (n=5 per group). Ibutilide prolonged both atrial and ventricular effective refractory period (ERP; by 15 and 8%, respectively; p<0.05 for both; Cycle length [CL] = 500 ms; n=5 for each). Flecainide significantly prolonged ERP in atria (by 27%; p<0.001) but not in ventricles (2% increase) due to atrial predominant induction of post-repolarization refractoriness. The combination of the two drugs prolonged ERP by 42% in atria (p<0.001 vs. control and 0<0.05 vs. flecainide alone) but by only 7% in ventricles (p<0.05; n=5, CL=500 ms). In separate experiments involving LV wedge preparations, risk factors for Torsade de Pointes were accentuated by ibutilide (50 nM) and slowed pacing (CL= 2000 ms; n=5). Under these conditions, ibutilide induced significant prolongation of QT and Tpeak /Tend intervals (by 25 and 55%, respectively; p<0.01 for both; n=5). Further addition of flecainide (1.5 μM) partially reversed these effects leading to an increase of these parameters by only 15 and 19%, respectively; p<0.05, combination vs. ibutilide; n=5). Torsade de Pointes was never observed with the drug combination. Our experimental data indicate that a combination of flecainide and ibutilide significantly improves pharmacological cardioversion of AF as well as prevention of AF with very little or no risk of long QT-mediated ventricular proarrhythmia.

Pulmonary Artery Denervation Inhibits Left Stellate Ganglion Stimulation-Induced Ventricular Arrhythmias Originating From the RVOT

BackgroundElectrical stimulation of the left stellate ganglion (LSG) can evoke ventricular arrhythmias (VAs) that originate from the right ventricular outflow tract (RVOT). The involvement of pulmonary artery innervation is unclear. ObjectivesThis study investigated the effects of selective pulmonary artery denervation (PADN) on blood pressure (BP), sympathetic activity, ventricular effective refractory period (ERP), and the incidence of VAs induced by LSG stimulation in canines. MethodsRadiofrequency ablation with basic anesthetic monitoring was used to induce PADN in canines. In Protocol 1 (n = 11), heart rate variability, serum norepinephrine and angiotensin-II levels, BP changes and ventricular ERP in response to LSG stimulation were measured before and after PADN. In Protocol 2 (n = 8), the incidence of VAs induced by LSG stimulation was calculated before and after PADN in a canine model of complete atrioventricular block. In addition, sympathetic nerves in the excised pulmonary arteries were immunohistochemically stained with tyrosine hydroxylase. ResultsThe low-frequency components of heart rate variability, serum norepinephrine and angiotensin-II levels were remarkably decreased post-PADN. Systolic BP elevation and RVOT ERP shortening induced by LSG stimulation were mitigated by PADN. The number of RVOT-premature ventricular contractions as well as RVOT tachycardia episodes and duration induced by LSG stimulation were significantly reduced after PADN. In addition, a large number of tyrosine hydroxylase-immunoreactive nerve fibers were located in the anterior wall of the pulmonary artery. ConclusionsPADN ameliorated RVOT ERP shortening, and RVOT-VAs induced by LSG stimulation by inhibiting cardiac sympathetic nerve activity.

Temporal and Spatial Changes of Proarrhythmic Substrate in Premature Ventricular Contraction–Induced Cardiomyopathy

BackgroundThe changes in proarrhythmic substrates and malignant ventricular arrhythmia mechanisms caused by premature ventricular contraction–induced cardiomyopathy (PVCCM) remain unclear. ObjectivesThe goal of this study was to establish the electrophysiological mechanism of how high-load PVC causes malignant arrhythmia. MethodsThirteen swine were exposed to 50% paced PVC from the right ventricular apex for 12 weeks (PVCCM, n = 6) and no pacing for 12 weeks (control, n = 7). Cardiac function was quantified biweekly with echocardiography. Computed tomography scans and electrophysiological examinations were performed monthly to dynamically evaluate the changes in the cardiac structure and the arrhythmogenic substrate. ResultsThe decreases in the cardiac function and ventricular enlargement in the PVCCM group were significant after 12 weeks of PVC stimulation compared with the control group (P < 0.001). Electrophysiological examination found that the ventricular effective refractory period dispersion (0.071 ± 0.008), area of the low-voltage zone (9.41 ± 1.55 cm2), and malignant ventricular arrhythmia inducibility (33.3%) of the PVCCM group increased significantly at week 8 after pacing (P < 0.001 vs the control group); these changes slowed down after 8 weeks. Moreover, the distribution of the low-voltage zone presented obvious spatial heterogeneity, especially in the anterior wall of the right ventricle, accompanied by delayed activation in the sinus rhythm (67 ± 13 milliseconds). Consistently, the proportion of ventricular fibrosis– and expression-related proteins were significantly increased in the PVCCM group (P < 0.001), especially in the right ventricle. Moreover, proteomic analysis confirmed the spatial profile of these fibrotic changes in the PVCCM group. ConclusionsHigh-burden PVC can cause significant temporal and spatial heterogeneity changes in proarrhythmic substrates, which are potentially related to the upregulation of calcium signaling caused by asynchronous activation.

LL-VNS attenuates SK2 expression and incidence of arrhythmias following acute myocardial infarction in rats.

Our previous study has demonstrated that low-level vagus nerve stimulation (LL-VNS) protects the heart against ventricular arrhythmias (VAs) induced by acute myocardial infarction (AMI). However, the potential mechanisms by which it influences ventricular electrophysiology remain unknown. Forty-five rats were divided into three groups: a Control group (sham AMI followed by sham LL-VNS, n = 15), an AMI group (AMI followed by sham LL-VSN for 60 mins, n = 15), and an AMI + LL-VNS group (AMI followed by LL-VSN for 60 mins, n = 15). Heart rate variability (HRV), ventricular effective refractory period (ERP), ventricular fibrillation threshold (VFT), and left stellate ganglion (LSG) activity were measured at baseline and during AMI. Finally, myocardial tissues were collected for tissue analysis. AMI directly induced hyperactivity in the LSG and reduced vagal tone as indexed by HRV. AMI also decreased VFT, and shortened ERP but increased ERP dispersion. AMI resulted in an increase in expression of ventricular small-conductance Ca2+-activated K+ (SK2). However, LL-VNS significantly mitigated or eliminated the effects of AMI. LL-VNS altered the electrophysiological properties of the ventricles through inhibition of cardiac sympathetic nervous activity and reduction in SK2 expression.

Median nerve stimulation elevates ventricular fibrillation threshold via the cholinergic anti-inflammatory pathway in myocardial infarction canine model.

Median nerve stimulation (MNS) diminishes regional myocardial ischemia and ventricular arrhythmia; however, the underlying mechanism has not been elucidated. In this study, we randomly categorized 22 adult mongrel dogs into a control group, MNS group 1, and MNS group 2. After a 4-week experimental myocardial infarction (MI), ventricular electrophysiology was measured in the MNS group 1 before and after 30 min of MNS. The same measurements were performed in the MNS group 2 dogs via bilateral vagotomy. Venous blood and ventricular tissue were collected to detect molecular indicators related to inflammation and cholinergic pathways by enzyme-linked immunosorbent assay (ELISA), immunohistochemistry (IHC), and Western blot (WB). No significant changes were reported in the ventricular effective refractory period (ERP) in the MNS group 1 and MNS group 2 dogs before and after MNS. The ventricular fibrillation threshold (VFT) in the MNS group 1 was significantly higher than that in the MNS group 2 (20.3 ± 3.7 V vs. 8.7 ± 2.9 V, P < 0.01). The levels of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and nuclear transcription factor-κB (NF-κB) were lower (P < 0.01), whereas the levels of Ach were higher in the peri-infarct zone tissues in the MNS group 1 dogs than those in the MNS group 2 dogs (P < 0.01). This study demonstrated that MNS increases VFT in a canine model with MI. The effects of MNS on VFT are potentially associated with the cholinergic anti-inflammatory pathway.

Evaluation of the direct protective effects of Canagliflozin on the Isoproterenol-induced cell injury in rat cardiomyocytes

Abstract Background Sodium-glucose cotransporter-2 (SGLT2) inhibitors are agents that act by inhibiting glucose and sodium reabsorption in the proximal renal tubule which promotes urinary glucose excretion. More recently, significant benefit data of SGLT2 inhibitors in patients with heart failure, independent of the presence of type 2 diabetes has been reported. We have previously demonstrated that Canagliflozin (Cana), a SGLT2 inhibitor, reduced the ventricular effective refractory period in isoproterenol (ISP)-induced myocardial injury rat model accompanied with the suppression of reactive oxygen species and the elevation of ketone bodies, suggesting the effect of Cana on electrical cardiac remodeling. The direct effect of Cana to the cardiomyocytes and its underlying molecular mechanism was remained to be clarified. We therefore established an ISP-induced neonatal rat ventricular cardiomyocyte (NRVCM) in vitro model, pretreated with Cana and/or ketone bodies. Methods Primary NRVCM were isolated from Wistar rats, were pretreated by Cana with or without βOHB (the most abundant ketone body in circulation), followed by a stimulation of ISP (10μM). Cells without drug or ketone body pretreatment were used as control. We then analyzed its effect on cell viability, apoptosis, and mitochondrial membrane potential using MTT assay, TUNEL assay, and mitochondrial membrane potential assay, respectively. MTT assay was also performed with or without PI3k inhibitor, LY294002. The end-labeling of DNA fragmentation were labelled with FITC, followed by the nuclei counterstain with DAPI and were observed with confocal microscope. The apoptotic index was defined as the percentage of TUNEL positive cells / total nuclei. Results Cana rescued the reduction of NRVCM cell viability induced by ISP stimulation for 24 hours which was inhibited by LY294002 compared to cells without pretreatment. Interestingly, pretreatment of βOHB with or without Cana improved also the NRCVM cell viability whereas there was no significant difference between these two conditions or with cells treated with Cana only, suggesting the direct protective effect of Cana. In 48 hours of ISP stimulation, the apoptotic index intends to decrease in Cana and/or βOHB compared to cells without pretreatment (Figure 1). Although the mitochondrial function was maintained in Cana-pretreated cells compared to cells without pretreatment, there was no significant difference in βOHB-pretreated cells. Conclusions Cana has a direct protective effect on cardiomyocytes cell viability, apoptosis as well as the mitochondrial function impaired by ISP through the cell survival signaling PI3K/Akt pathway. This brings a new insight to the therapeutic target of cardiovascular disease. Funding Acknowledgement Type of funding sources: None.