BACKGROUND:Activation recovery interval (ARI), extracted from unipolar electrograms, serves as a practical surrogate for repolarization during experimental studies in vivo. Far-field signal contamination and low spatial resolution obscure regional repolarization gradients and duration alternans detection using unipolar ARI. We hypothesized that the attenuation of far-field contamination with the principal component-referenced unipole will allow for a more accurate assessment of true local repolarization gradients and spatially assess action potential duration alternans. METHODS:Unipolar ARI and the novel method, RepolLoc, were validated for the detection of spatial and temporal repolarization changes using simultaneous optical and electrical mapping in a rabbit Langendorff model. Repolarization changes were created using global infusion of ibutilide or pinacidil, or topical application of lidocaine. Epicardial mapping was conducted in a porcine Langendorff model following the topical application of lidocaine to investigate the spatial resolution of each method. Generalized linear models of the two methods were used to compare with optical action potential duration (APD80). RESULTS:Following the infusion of antiarrhythmic drugs, the RepolLoc method (slope=0.90) had a slightly higher correlation to optical APD80 than the ARI method (slope=0.79). Following regional application of lidocaine, RepolLoc was better able to localize the site of drug administration with an average 26.12% reduction as compared with 18.66% reduction in unipolar ARI (P=0.0046). Additionally, temporal repolarization alternans and restitution changes assessed by RepolLoc method tracked optical APD80 quantified time domain changes. CONCLUSIONS:RepolLoc has higher sensitivity to local spatiotemporal repolarization heterogeneities and alternans than traditional ARI. Although ARI only correlates with uniformly distributed changes in repolarization in the entire myocardium, RepolLoc also provided accurate regional gradient assessment and duration alternans of repolarization. These findings suggest ARI has significant far-field contamination and RepolLoc may provide a better clinical mapping tool for spatiotemporal repolarization gradient mapping.

Background: We previously described the mortality associated with cardiac injury in patients with coronavirus disease 2019 (COVID-19). The activation of immune and thrombotic biomarkers at admission, and their ability to predict cardiac injury and mortality patterns in COVID-19, remains unclear. Methods: This retrospective cohort study included 170 patients with COVID-19 with cardiac injury at the time of admission to Tongji Hospital in Wuhan between January 29, 2020, and March 8, 2020. The temporal evolution of inflammatory cytokines, coagulation markers, clinical treatment, and mortality were analyzed. Continuous variables are expressed as median (interquartile range). The Mann-Whitney test was used for two-group comparisons, whereas the Kruskal-Wallis test was used for comparisons among three groups. Categorical variables are expressed as proportions and percentages, and Fisher’s exact test was used to compare differences. A multivariate regression model was used to predict in-hospital death. A simple linear regression analysis was applied to examine the correlation between baseline biomarkers and peak cTnI levels. Results: Of the 170 patients, 60 (35.3%) died early (<21 d), and 61 (35.9%) died after a prolonged stay. The admission laboratory findings correlating with early death were elevated interleukin 6 (IL-6) (P < 0.0001), tumor necrosis factor-α (P = 0.0025), and C-reactive protein (P < 0.0001). We observed the trajectory of biomarker changes in patients after admission hospitalization, and determined that early mortality was associated with a rapidly increasing D-dimer level, and gradually decreasing platelet and lymphocyte counts. Multivariate and simple linear regression models indicated that the risk of death was associated with immune and thrombotic pathway activation. Elevated admission cTnI levels were associated with elevated IL-6 (P = 0.03) and D-dimer (P = 0.0021) levels. Conclusion: In patients with COVID-19 with cardiac injury, IL-6 and D-dimer levels at admission predicted subsequently elevated cTnI levels and early death, thus highlighting the need for early inflammatory cytokine-based risk stratification in patients with cardiac injury.

Despite significant progress in pharmacotherapy, ventricular arrhythmia (VA) remains a challenge in heart failure (HF). Use of most conventional anti-arrhythmic drugs is limited by proarrhythmia and/or negative inotropic effects.

Background We recently demonstrated that acute administration of ibrutinib, a Bruton's tyrosine kinase inhibitor used in chemotherapy for blood malignancies, increases ventricular arrhythmia (VA) vulnerability. A pathway of ibrutinib‐induced vulnerability to VA that can be modulated for cardioprotection remains unclear. Methods and Results The effects of ibrutinib on cardiac electrical activity and Ca 2+ dynamics were investigated in Langendorff‐perfused hearts using optical mapping. We also conducted Western blotting analysis to evaluate the impact of ibrutinib on various regulatory and Ca 2+ ‐handling proteins in rat cardiac tissues. Treatment with ibrutinib (10 mg/kg per day) for 4 weeks was associated with an increased VA inducibility (72.2%±6.3% versus 38.9±7.0% in controls, P <0.002) and shorter action potential durations during pacing at various frequencies ( P <0.05). Ibrutinib also decreased heart rate thresholds for beat–to–beat duration alternans of the cardiac action potential ( P <0.05). Significant changes in myocardial Ca 2+ transients included lower amplitude alternans ratios ( P <0.05), longer times‐to‐peak ( P <0.05), and greater spontaneous intracellular Ca 2+ elevations ( P <0.01). We also found lower abundance and phosphorylation of myocardial AMPK (5′‐adenosine monophosphate‐activated protein kinase), indicating reduced AMPK activity in hearts after ibrutinib treatment. An acute treatment with the AMPK activator 5‐aminoimidazole‐4‐carboxamide‐1‐β‐D‐ribofuranoside ameliorated abnormalities in action potential and Ca 2+ dynamics, and significantly reduced VA inducibility (37.1%±13.4% versus 72.2%±6.3% in the absence of 5‐aminoimidazole‐4‐carboxamide‐1‐β‐D‐ribofuranoside, P <0.05) in hearts from ibrutinib‐treated rats. Conclusions VA vulnerability inflicted by ibrutinib may be mediated in part by an impairment of myocardial AMPK activity. Pharmacological activation of AMPK may be a protective strategy against ibrutinib‐induced cardiotoxicity.

Cardiac arrhythmias remain a significant concern with Ibrutinib (IBR), a first-generation Bruton’s tyrosine kinase inhibitor (BTKi). Acalabrutinib (ABR), a next-generation BTKi, is associated with reduced atrial arrhythmia events. However, the role of ABR in ventricular arrhythmia (VA) has not been adequately evaluated. Our study aimed to investigate VA vulnerability and ventricular electrophysiology following chronic ABR therapy in male Sprague–Dawley rats utilizing epicardial optical mapping for ventricular voltage and Ca2+ dynamics and VA induction by electrical stimulation in ex-vivo perfused hearts. Ventricular tissues were snap-frozen for protein analysis for sarcoplasmic Ca2+ and metabolic regulatory proteins. The results show that both ABR and IBR treatments increased VA vulnerability, with ABR showing higher VA regularity index (RI). IBR, but not ABR, is associated with the abbreviation of action potential duration (APD) and APD alternans. Both IBR and ABR increased diastolic Ca2+ leak and Ca2+ alternans, reduced conduction velocity (CV), and increased CV dispersion. Decreased SERCA2a expression and AMPK phosphorylation were observed with both treatments. Our results suggest that ABR treatment also increases the risk of VA by inducing proarrhythmic changes in Ca2+ signaling and membrane electrophysiology, as seen with IBR. However, the different impacts of these two BTKi on ventricular electrophysiology may contribute to differences in VA vulnerability and distinct VA characteristics.

The current antiarrhythmic paradigm is mainly centered around modulating membrane voltage. However, abnormal cytosolic calcium (Ca2+) signaling, which plays an important role in driving membrane voltage, has not been targeted for therapeutic purposes in arrhythmogenesis. There is clear evidence for bidirectional coupling between membrane voltage and intracellular Ca2+. Cytosolic Ca2+ regulates membrane voltage through Ca2+-sensitive membrane currents. As a component of Ca2+-sensitive currents, Ca2+-activated nonspecific cationic current through the TRPM4 (transient receptor potential melastatin 4) channel plays a significant role in Ca2+-driven changes in membrane electrophysiology. In myopathic and ischemic ventricles, upregulation and/or enhanced activity of this current is associated with the generation of afterdepolarization (both early and delayed), reduction of repolarization reserve, and increased propensity to ventricular arrhythmias. In this review, we describe a novel concept for the management of ventricular arrhythmias in the remodeled ventricle based on mechanistic concepts from experimental studies, by uncoupling the Ca2+-induced changes in membrane voltage by inhibition of this TRPM4-mediated current.

Abstract Funding Acknowledgements Type of funding sources: None. Background In addition to variety of force time indices, electrophysiologist use peak to peak voltage (Vpp) and or S wave on unipolar electrogram to assess RF lesions they deliver. This strategy ignores cardiac conduction speed, direction, and wave curvature, all of which are affected by delivered lesion. Electrode array catheters such as the Advisor HD Grid allow individual cardiac wavefronts from sites to be characterized simultaneously in two dimensions from which vectorcardiograms can be derived. This underappreciated attribute of electrograms when integrated into activation mapping may provide new insights into the electrophysiologic state of tissue including wavefront amplitude, direction and velocity. Objective We tested the hypothesis that intracardiac vectorcardiogram loops contain aspects of properties that may better detect lesions compared to Vpp reduction and that a novel multi-modal approach will provide additional insights on wavefront interaction with substrate. Methods In 5 ex-vivo Langendorff perfused swine hearts, 56-pole (7 x 8 configuration) electrode arrays were sutured to the anterior epicardial left ventricle. Unipolar electrograms were acquired with UHN mapping system during programmed stimulation from all cardinal directions before and after creating RF ablation lesions. Vector loop maps were derived from orthogonal bipole pairs to study wavefront organization and voltage. Isochrone maps were generated from unipolar local activation times. Vector loop eccentricity was defined for each 4-electrode group as a function of loop’s major and minor axis lengths. Low values denote less eccentricity (more circular). Results An illustrative example of vector, loop and isochrone maps before and after ablation is shown in figures 1 and 2. Pre-ablation conditions show activation from a point stimulation on the left side with slender vector loops indicative of a relatively homogeneous linear conduction. In contrast, post-ablation vector loops are much less eccentric near the ablated area and smaller reflecting attenuated amplitude. Interestingly, near border zone locations large round loops are observed, suggesting curved or nonlinear wave propagation. Figure 2 shows that vector loops recorded from scar areas were diminished in size following ablation with a mean Vpp of 3.76 vs 2.18mV (p=0.0001) and exhibited less mean eccentricity, 0.925 vs 0.848 (p<0.0001), with more round loops (eccentricity < 0.7) found after ablation (12% of sites vs 4% pre-ablation, p < 0.0001). Conclusions Significant changes in loop characteristics were observed even when ablation induced voltage attenuation was minimal. This suggests that vectorial loop eccentricity may provide information in addition to omnipolar Vpp. Wavefront propagation disruption near ablation border zones is thus potentially valuable to detect lesion gaps. This information along with wave amplitude and direction can now be integrated into a novel multi-modal map.

To better understand sodium channel (SCN5A)-related cardiomyopathies, we generated ventricular cardiomyocytes from induced pluripotent stem cells obtained from a dilated cardiomyopathy patient harbouring the R222Q mutation, which is only expressed in adult SCN5A isoforms. Because the adult SCN5A isoform was poorly expressed, without functional differences between R222Q and control in both embryoid bodies and cell sheet preparations (cultured for 29-35 days), we created heart-on-a-chip biowires which promote myocardial maturation. Indeed, biowires expressed primarily adult SCN5A with R222Q preparations displaying (arrhythmogenic) short action potentials, altered Na+ channel biophysical properties and lower contractility compared to corrected controls. Comprehensive RNA sequencing revealed differential gene regulation between R222Q and control biowires in cellular pathways related to sarcoplasmic reticulum and dystroglycan complex as well as biological processes related to calcium ion regulation and action potential. Additionally, R222Q biowires had marked reductions in actin expression accompanied by profound sarcoplasmic disarray, without differences in cell composition (fibroblast, endothelial cells, and cardiomyocytes) compared to corrected biowires. In conclusion, we demonstrate that in addition to altering cardiac electrophysiology and Na+ current, the R222Q mutation also causes profound sarcomere disruptions and mechanical destabilization. Possible mechanisms for these observations are discussed.

Abstract Funding Acknowledgements Type of funding sources: None. Background In addition to wave propagation, tissue anisotropy is thought to determine repolarization (repol). This may explain PVC-induced vulnerability to re-entry, based on change of activation setting up arrhythmogenic repol gradients. However, differential repolarization effects produced by waves propagating in various directions has not been studied. Though this effect could be measured with optical mapping it is unknown whether egm-based methods can detect these differences. Purpose The objective of this study was to determine the effect of wave direction on cardiac repol as measured by optical mapping. We recently proposed a novel method of assessing repol with the use of equi-spaced array catheters that allow integration of orthogonal bipolar egms and we sought to compare the performance of unipolar-based ARI methods and orthogonal bipolar egms in detecting the changes shown in optical mapping. Methods Simultaneous optical mapping and epicardial mapping with equi-spaced array catheters (Optrell and HD Grid) was performed in 6 rabbit Langendorff experiments. Unipolar egms from 4 electrodes forming a square in the middle of the array were recorded. A compound egm, called rEGM was created from orthogonal bipolar egms derived from the unipolar egms. Optical mapping was performed with a sampling rate of 3333 frames/s. Epicardial waves propagating in different directions were produced by point stimulation (CL = 200ms) at various location respective to the electrode array: Left (anterior LV, Pace B), right (lateral LV, Pace A), catheter distal (apex, Pace D) and catheter proximal (anterior base, Pace C). An endocardial source located transmurally across the electrode array was also evaluated. APD80 from optical data was measured from an algorithm described before. An APD estimate from rEGM, APDc was measured from the onset of QRS to baseline return of rEGM. For each method (optical, APDc and ARI) a statistical analysis evaluating the effect of wave propagation was performed. Results a) Gold Standard, optical mapping: Left column on figure shows the APD80 measurements respective to wave direction for two of the experiments. Kruskal-Wallis analysis showed a significant effect of wave direction on APD80 (p < 0.01 for both experiments 3 & 4). Multiple comparisons showed most notable effect from Pace A (lateral side). b) APDc shown in middle column successfully detected the changes (p < 0.01). c) Unipolar ARI: Right column shows the ARI measurements from the same experiments that failed to detect a directional repolarization difference (Exp3 p=0.10; Exp4 p =0.62). Conclusions Changes in direction of wave front propagation can change repol timing of as much as 30msec. These changes were not detectable by ARI method. However subtle changes of repolarization were successfully detected by orthogonal bipolar approach using equi-spaced array catheters, enabling a reliable method to assess minute repolarization changes locally.

AbstractAimsElectroanatomical maps using automated conduction velocity (CV) algorithms are now being calculated using two-dimensional (2D) mapping tools. We studied the accuracy of mapping surface 2D CV, compared to the three-dimensional (3D) vectors, and the influence of mapping resolution in non-scarred animal and human heart models.Methods and resultsTwo models were used: a healthy porcine Langendorff model with transmural needle electrodes and a computer stimulation model of the ventricles built from an MRI-segmented, excised human heart. Local activation times (LATs) within the 3D volume of the mesh were used to calculate true 3D CVs (direction and velocity) for different pixel resolutions ranging between 500 μm and 4 mm (3D CVs). CV was also calculated for endocardial surface-only LATs (2D CV). In the experimental model, surface (2D) CV was faster on the epicardium (0.509 m/s) compared to the endocardium (0.262 m/s). In stimulation models, 2D CV significantly exceeded 3D CVs across all mapping resolutions and increased as resolution decreased. Three-dimensional and 2D left ventricle CV at 500 μm resolution increased from 429.2 ± 189.3 to 527.7 ± 253.8 mm/s (P < 0.01), respectively, with modest correlation (R = 0.64). Decreasing the resolution to 4 mm significantly increased 2D CV and weakened the correlation (R = 0.46). The majority of CV vectors were not parallel (<30°) to the mapping surface providing a potential mechanistic explanation for erroneous LAT-based CV over-estimation.ConclusionVentricular CV is overestimated when using 2D LAT-based CV calculation of the mapping surface and significantly compounded by mapping resolution. Three-dimensional electric field-based approaches are needed in mapping true CV on mapping surfaces.

Background:Post-defibrillation myocardial contractile dysfunction adversely affects the survival of patients after cardiac arrest. Attenuation of diastolic calcium (Ca2+) overload by stabilization of the cardiac ryanodine receptor (RyR2) is found to reduce refibrillation after long-duration ventricular fibrillation (LDVF). Objective:In the present study, we explored the effects of RyR2 stabilization by azumolene on systolic Ca2+ release synchrony and myocardial contractility. Methods:After completion of baseline optical mapping, Langendorff-perfused rabbit hearts were subjected to global ischemia followed by reperfusion with azumolene or deionized distilled water (vehicle). Following reperfusion, LDVF was induced with burst pacing. In the first series of experiments (n = 16), epicardial Ca2+ transient was analyzed for Ca2+ transient amplitude alternans and dispersion of Ca2+ transient amplitude alternans index (CAAI). In the second series of experiments following the same protocol (n = 12), ventricular contractility was assessed by measuring the left ventricular pressure. Results:Ischemic LDVF led to greater CAAI (0.06 ± 0.02 at baseline vs 0.12 ± 0.02 post-LDVF, P < .01) and magnitude of dispersion of CAAI (0.04 ± 0.01 vs 0.09 ± 0.01, P < .01) in control hearts. In azumolene-treated hearts, no significant changes in CAAI (0.05 ± 0.01 vs 0.05 ± 0.01, P = .84) and dispersion of CAAI (0.04 ± 0.01 vs 0.04 ± 0.01, P = .99) were noted following ischemic LDVF. Ischemic LDVF was associated with reduction in left ventricular developed pressure (100% vs 36.8% ± 6.1%, P = .002) and dP/dtmax (100% vs 45.3% ± 6.5%, P = .003) in control hearts, but these reductions were mitigated (left ventricular developed pressure: 100% vs 74.0% ± 8.1%, P = .052, dP/dtmax: 100% vs 80.8% ± 7.9%, P = .09) in azumolene-treated hearts. Conclusion:Treatment with azumolene is associated with improvement of systolic Ca2+ release synchrony and myocardial contractility following ischemic LDVF.

Traditionally, LAT maps depicting earliest activation, derived star burst vector depictions, and QS patterns on unipoles have been used to detect breakout sites. In the ventricle such strategies may be limited due to indeterminate LAT annotation, myocardial thickness and tangential activation to the mapping surface.

Background: Conventional mapping of focal ventricular arrhythmias relies on unipolar electrogram characteristics and early local activation times. Deep intramural foci are common and associated with high recurrence rates following catheter-based radiofrequency ablation. We assessed the accuracy of unipolar morphological patterns and mapping surface indices to predict the site and depth of ventricular arrhythmogenic focal sources. Methods: An experimental beating-heart model used Langendorff-perfused, healthy swine hearts. A custom 56-pole electrode array catheter was positioned on the left ventricle. A plunge needle was placed perpendicular in the center of the grid to simulate arrhythmic foci at variable depths. Unipolar electrograms and local activation times were generated. Simulation models from 2 human hearts were also included with grids positioned simultaneously on the endocardium-epicardium from multiple left ventricular, septal, and outflow tract sites. Results: A unipolar Q or QS complex lacks specificity for superficial arrhythmic foci, as this morphology pattern occupies a large surface area and is the predominant pattern as intramural depth increases without developing a R component. There is progressive displacement from the arrhythmic focus to the surface exit as intramural focus depth increases. A shorter total activation time over the overlying electrode array, larger surface area within initial 20 ms activation, and a dual surface breakout pattern all indicate a deep focus. Conclusions: Displacement from the focal intramural origin to the exit site on the mapping surface could lead to erroneous lesion delivery strategies. Traditional unipolar electrogram features lack specificity to predict the intramural arrhythmic source; however, novel endocardial-epicardial mapping surface indices can be used to determine the depth of arrhythmic foci.