A comprehensive framework for evaluation of high pacing frequency and arrhythmic optical mapping signals.

Introduction: High pacing frequency or irregular activity due to arrhythmia produces complex optical mapping signals and challenges for processing. The objective is to establish an automated activation time-based analytical framework applicable to optical mapping images of complex electrical behavior. Methods: Optical mapping signals with varying complexity from sheep (N = 7) ventricular preparations were examined. Windows of activation centered on each action potential upstroke were derived using Hilbert transform phase. Upstroke morphology was evaluated for potential multiple activation components and peaks of upstroke signal derivatives defined activation time. Spatially and temporally clustered activation time points were grouped in to wave fronts for individual processing. Each activation time point was evaluated for corresponding repolarization times. Each wave front was subsequently classified based on repetitive or non-repetitive events. Wave fronts were evaluated for activation time minima defining sites of wave front origin. A visualization tool was further developed to probe dynamically the ensemble activation sequence. Results: Our framework facilitated activation time mapping during complex dynamic events including transitions to rotor-like reentry and ventricular fibrillation. We showed that using fixed AT windows to extract AT maps can impair interpretation of the activation sequence. However, the phase windowing of action potential upstrokes enabled accurate recapitulation of repetitive behavior, providing spatially coherent activation patterns. We further demonstrate that grouping the spatio-temporal distribution of AT points in to coherent wave fronts, facilitated interpretation of isolated conduction events, such as conduction slowing, and to derive dynamic changes in repolarization properties. Focal origins precisely detected sites of stimulation origin and breakthrough for individual wave fronts. Furthermore, a visualization tool to dynamically probe activation time windows during reentry revealed a critical single static line of conduction slowing associated with the rotation core. Conclusion: This comprehensive analytical framework enables detailed quantitative assessment and visualization of complex electrical behavior.

Tissue Preparation Techniques for Contrast-Enhanced Micro Computed Tomography Imaging of Large Mammalian Cardiac Models with Chronic Disease.

Structural remodeling is a common consequence of chronic pathological stresses imposed on the heart. Understanding the architectural and compositional properties of diseased tissue is critical to determine their interactions with arrhythmic behavior. Microscale tissue remodeling, below the clinical resolution, is emerging as an important source of lethal arrhythmia, with high prevalence in young adults. Challenges remain in obtaining high imaging contrast at sufficient microscale resolution for preclinical models, such as large mammalian whole hearts. Moreover, tissue composition-selective contrast enhancement for three-dimensional high-resolution imaging is still lacking. Non-destructive imaging using micro-computed tomography shows promise for high-resolution imaging. The objective was to alleviate sufferance from X-ray over attenuation in large biological samples. Hearts were extracted from healthy pigs (N = 2), and sheep (N = 2) with either induced chronic myocardial infarction and fibrotic scar formation or induced chronic atrial fibrillation. Excised hearts were perfused with: a saline solution supplemented with a calcium ion quenching agent and a vasodilator, ethanol in serial dehydration, and hexamethyldisilizane under vacuum. The latter reinforced the heart structure during air-drying for 1 week. Collagen-dominant tissue was selectively bound by an X-ray contrast-enhancing agent, phosphomolybdic acid. Tissue conformation was stable in air, permitting long-duration microcomputed tomography acquisitions to obtain high-resolution (isotropic 20.7 µm) images. Optimal contrast agent loading by diffusion showed selective contrast enhancement of the epithelial layer and sub-endocardial Purkinje fibers in healthy pig ventricles. Atrial fibrillation (AF) hearts showed enhanced contrast accumulation in the posterior walls and appendages of the atria, attributed to greater collagen content. Myocardial infarction hearts showed increased contrast selectively in regions of cardiac fibrosis, which enabled the identification of interweaving surviving myocardial muscle fibers. Contrast-enhanced air-dried tissue preparations enabled microscale imaging of the intact large mammalian heart and selective contrast enhancement of underlying disease constituents.

A novel system for mapping regional electrical properties and characterizing arrhythmia in isolated intact rat atria.

Detailed global maps of atrial electrical activity are needed to understand mechanisms of atrial rhythm disturbance in small animal models of heart disease. To date, optical mapping systems have not provided enough spatial resolution across sufficiently extensive regions of intact atrial preparations to achieve this goal. The aim of this study was to develop an integrated platform for quantifying regional electrical properties and analyzing reentrant arrhythmia in a biatrial preparation. Intact atria from 6/7-mo-old female spontaneously hypertensive rats (SHRs; n = 6) were isolated and secured in a constant flow superfusion chamber at 37°C. Optical mapping was performed with the membrane-voltage dye di-4-ANEPPS using LED excitation and a scientific complementary metal-oxide semiconductor (sCMOS) camera. Programmed stimulus trains were applied from right atrial (RA) and left atrial (LA) sites to assess rate-dependent electrical behavior and to induce atrial arrhythmia. Signal-to-noise ratio was improved by sequential processing steps that included spatial smoothing, temporal filtering, and, in stable rhythms, ensemble-averaging. Activation time, repolarization time, and action potential duration (APD) maps were constructed at high spatial resolution for a wide range of coupling intervals. These data were highly consistent within and between experiments. They confirmed preferential atrial conduction pathways and demonstrated distinct medial-to-lateral APD gradients. We also showed that reentrant arrhythmias induced in this preparation were explained by the spatial variation of these electrical properties. Our new methodology provides a robust means of 1) quantifying regional electrical properties in the intact rat atria at higher spatiotemporal resolution than previously reported, and 2) characterizing reentrant arrhythmia and analyzing mechanisms that give rise to it.NEW & NOTEWORTHY Despite wide-ranging optical mapping studies, detailed information on regional atrial electrical properties in small animal models of heart disease and how these contribute to reentrant arrhythmia remains limited. We have developed a novel experimental platform that enables both to be achieved in a geometrically intact isolated rat bi-atrial preparation.