The essential role of arterial pulse in venous return.

The close proximity of arteries and veins is a well-known anatomical finding documented in the extremities of all vertebrates. However, the physiological consequences of this arrangement are rarely given proper consideration nor are they covered in the textbook list of mechanisms that aid blood flow. We hypothesized that arterial pressure pulsations can significantly increase blood flow in the adjacent valve-containing vein segments. To demonstrate the existence of this mechanism, 10- to 15-cm sections of the bovine forelimb neurovascular bundle were isolated. The proximal and distal ends of the median artery and adjacent veins were cannulated, their tributaries were tied off, and the dissected bundle was then inserted into an airtight enclosure to mimic in vivo encasement by surrounding muscle. Pulsatile pressure was subsequently applied to the arterial segment while recording venous flow. At pressure settings mimicking physiological scenarios, arterial pulsations caused a highly significant increase in venous return. The amplitude of this effect was dependent on the arterial pulsation rate, stroke volume, and pressure gradient across the vein segment.

Human ES-cell-derived cardiomyocytes electrically couple and suppress arrhythmias in injured hearts.

Transplantation studies in mice and rats have shown that human embryonic-stem-cell-derived cardiomyocytes (hESC-CMs) can improve the function of infarcted hearts, but two critical issues related to their electrophysiological behaviour in vivo remain unresolved. First, the risk of arrhythmias following hESC-CM transplantation in injured hearts has not been determined. Second, the electromechanical integration of hESC-CMs in injured hearts has not been demonstrated, so it is unclear whether these cells improve contractile function directly through addition of new force-generating units. Here we use a guinea-pig model to show that hESC-CM grafts in injured hearts protect against arrhythmias and can contract synchronously with host muscle. Injured hearts with hESC-CM grafts show improved mechanical function and a significantly reduced incidence of both spontaneous and induced ventricular tachycardia. To assess the activity of hESC-CM grafts in vivo, we transplanted hESC-CMs expressing the genetically encoded calcium sensor, GCaMP3 (refs 4, 5). By correlating the GCaMP3 fluorescent signal with the host ECG, we found that grafts in uninjured hearts have consistent 1:1 host­graft coupling. Grafts in injured hearts are more heterogeneous and typically include both coupled and uncoupled regions. Thus, human myocardial grafts meet physiological criteria for true heart regeneration, providing support for the continued development of hESC-based cardiac therapies for both mechanical and electrical repair.

Physiological response of cardiac tissue to bisphenol A: alterations in ventricular pressure and contractility.

Biomonitoring studies have indicated that humans are routinely exposed to bisphenol A (BPA), a chemical that is commonly used in the production of polycarbonate plastics and epoxy resins. Epidemiological studies have shown that BPA exposure in humans is associated with cardiovascular disease; however, the direct effects of BPA on cardiac physiology are largely unknown. Previously, we have shown that BPA exposure slows atrioventricular electrical conduction, decreases epicardial conduction velocity, and prolongs action potential duration in excised rat hearts. In the present study, we tested if BPA exposure also adversely affects cardiac contractile performance. We examined the impact of BPA exposure level, sex, and pacing rate on cardiac contractile function in excised rat hearts. Hearts were retrogradely perfused at constant pressure and exposed to 10(-9)-10(-4) M BPA. Left ventricular developed pressure and contractility were measured during sinus rhythm and during pacing (5, 6.5, and 9 Hz). Ca(2+) transients were imaged from whole hearts and from neonatal rat cardiomyocyte layers. During sinus rhythm in female hearts, BPA exposure decreased left ventricular developed pressure and inotropy in a dose-dependent manner. The reduced contractile performance was exacerbated at higher pacing rates. BPA-induced effects on contractile performance were also observed in male hearts, albeit to a lesser extent. Exposure to BPA altered Ca(2+) handling within whole hearts (reduced diastolic and systolic Ca(2+) transient potentiation) and neonatal cardiomyocytes (reduced Ca(2+) transient amplitude and prolonged Ca(2+) transient release time). In conclusion, BPA exposure significantly impaired cardiac performance in a dose-dependent manner, having a major negative impact upon electrical conduction, intracellular Ca(2+) handing, and ventricular contractility.

Changes in attachment and metabolic activity of rat neonatal cardiomyocytes and nonmyocytes caused by Macrovipera lebetina obtusa venom.

The Caucasian viper Macrovipera lebetina obtusa (MLO) is one of the most prevalent and venomous snakes in the Caucasus and the surrounding regions, yet the effects of MLO venom on cardiac function remain largely unknown. We examined the influence of MLO venom (crude and with inhibited metalloproteinases and phospholipase A2) on attachment and metabolic activity of rat neonatal cardiomyocytes (CM) and nonmyocytes (nCM), assessed at 1 and 24 h. After exposing both CM and nCM to varying concentrations of MLO venom, we observed immediate cytotoxic effects at a concentration of 100 µg/ml, causing detachment from the culture substrate. At lower MLO venom concentrations both cell types detached in a dose-dependent manner. Inhibition of MLO venom metalloproteinases significantly improved CM and nCM attachment after 1-hour exposure. At 24-hour exposure to metalloproteinases inhibited venom statistically significant enhancement was observed only in nCM attachment. However, metabolic activity of CM and nCM did not decrease upon exposure to the lower dose of the venom. Moreover, we demonstrated that metalloproteinases and phospholipases A2 are not the components of the MLO venom that change metabolic activity of both CM and nCM. These results provide a valuable platform to study the impact of MLO venom on prey cardiac function. They also call for further exploration of individual venom components for pharmaceutical purposes.

Properties of blebbistatin for cardiac optical mapping and other imaging applications.

Blebbistatin is a recently discovered myosin II inhibitor. It is rapidly becoming a compound of choice to reduce motion artifacts during cardiac optical mapping, as well as to study cell motility and cell invasion. Although blebbistatin has a number of advantages over other electromechanical uncouplers, many of its properties have yet to be addressed. Here we describe several methodological issues associated with the use of blebbistatin, including its spectral properties, reversibility, and its effect on tissue metabolic state. We show that if precautions are not taken, perfusion with blebbistatin may result in blebbistatin precipitate that accumulates in the vasculature. Although such precipitate is fluorescent, it is not detectable within wavelength bands that are typically used for transmembrane voltage fluorescence imaging (i.e., emission wavelengths >600 nm). Therefore, blockage of the microcirculation by blebbistatin may cause data misinterpretation in studies that use voltage-sensitive dyes. Blebbistatin may also impact imaging of green fluorophores due to the spectral shift it causes in endogenous tissue fluorescence. 3D excitation-emission matrices of blebbistatin in precipitate form and in various solutions (DMSO, water, and 1 % aqueous albumin) revealed significant changes in the fluorescence of this molecule in different environments. Finally, we examined the reversibility of blebbistatin's uncoupling effect on cardiac contraction. Our findings provide important new information about the properties of this myosin II inhibitor, which will aid in the proper design and interpretation of studies that use this compound.

Use of endogenous NADH fluorescence for real-time in situ visualization of epicardial radiofrequency ablation lesions and gaps.

Radiofrequency ablation (RFA) aims to produce lesions that interrupt reentrant circuits or block the spread of electrical activation from sites of abnormal activity. Today, there are limited means for real-time visualization of cardiac muscle tissue injury during RFA procedures. We hypothesized that the fluorescence of endogenous NADH could be used as a marker of cardiac muscle injury during epicardial RFA procedures. Studies were conducted in blood-free and blood-perfused hearts from healthy adult Sprague-Dawley rats and New Zealand rabbits. Radiofrequency was applied to the epicardial surface of the heart using a 4-mm standard blazer ablation catheter. A dual camera optical mapping system was used to monitor NADH fluorescence upon ultraviolet illumination of the epicardial surface and to record optical action potentials using the voltage-sensitive probe RH237. Epicardial lesions were seen as areas of low NADH fluorescence. The lesions appeared immediately after ablation and remained stable for several hours. Real-time monitoring of NADH fluorescence allowed visualization of viable tissue between the RFA lesions. Dual recordings of NADH and epicardial electrical activity linked the gaps between lesions to postablation reentries. We found that the fluorescence of endogenous NADH aids the visualization of injured epicardial tissue caused by RFA. This was true for both blood-free and blood-perfused preparations. Gaps between NADH-negative regions revealed unablated tissue, which may promote postablation reentry or provide pathways for the conduction of abnormal electrical activity.

Building Valveless Impedance Pumps From Biological Components: Progress and Challenges.

Valveless pumping based on Liebau mechanism entails asymmetrical positioning of the compression site relative to the attachment sites of the pump's elastic segment to the rest of the circuit. Liebau pumping is believed to play a key role during heart development and be involved in several other physiological processes. Until now studies of Liebau pump have been limited to numerical analyses, in silico modeling, experiments using non-biological elements, and a few indirect in vivo measurements. This review aims to stimulate experimental efforts to build Liebau pumps using biologically compatible materials in order to encourage further exploration of the fundamental mechanisms behind valveless pumping and its role in organ physiology. The covered topics include the biological occurrence of Liebau pumps, the main differences between them and the peristaltic flow, and the potential uses and body sites that can benefit from implantable valveless pumps based on Liebau principle. We then provide an overview of currently available tools to build such pumps and touch upon limitations imposed by the use of biological components. We also talk about the many variables that can impact Liebau pump performance, including the concept of resonant frequencies, the shape of the flowrate-frequency relationship, the flow velocity profiles, and the Womersley numbers. Lastly, the choices of materials to build valveless impedance pumps and possible modifications to increase their flow output are briefly discussed.

Output of a valveless Liebau pump with biologically relevant vessel properties and compression frequencies.

Liebau pump is a tubular, non-peristaltic, pulsatile pump capable of creating unidirectional flow in the absence of valves. It requires asymmetrical positioning of the pincher relative to the attachment sites of its elastic segment to the rest of the circuit. Biological feasibility of such valveless pumps remains a hotly debated topic. To test the feasibility of the Liebau-based pumping in vessels with biologically relevant properties we quantified the output of Liebau pumps with their  compliant segments made of a silicone rubber that mimicked the Young modulus of soft tissues. The lengths, the inner diameters, thicknesses of the tested compliant segments ranged from 1 to 5 cm, 3 to 8 mm and 0.3 to 1 mm, respectively. The compliant segment of the setup was compressed at 0.5-2.5 Hz frequencies using a 3.5-mm-wide rectangular piston. A nearest-neighbor tracking algorithm was used to track movements of 0.5-mm carbon particles within the system. The viscosity of the aqueous solution was varied by increased percentage of glycerin. Measurements yielded quantitative relationships between viscosity, frequency of compression and the net flowrate. The use of the Liebau principle of valveless pumping in conjunction with physiologically sized vessel and contraction frequencies yields flowrates comparable to peristaltic pumps of the same dimensions. We conclude that the data confirm physiological feasibility of Liebau-based pumping and warrant further testing of its mechanism using excised biological conduits or tissue engineered components. Such biomimetic pumps can serve as energy-efficient flow generators in microdevices or to study the function of embryonic heart during its normal development or in diseased states.

Key factors behind autofluorescence changes caused by ablation of cardiac tissue.

Radiofrequency ablation is a commonly used clinical procedure that destroys arrhythmogenic sources in patients suffering from atrial fibrillation and other types of cardiac arrhythmias. To improve the success of this procedure, new approaches for real-time visualization of ablation sites are being developed. One of these promising methods is hyperspectral imaging, an approach that detects lesions based on changes in the endogenous tissue autofluorescence profile. To facilitate the clinical implementation of this approach, we examined the key variables that can influence ablation-induced spectral changes, including the drop in myocardial NADH levels, the release of lipofuscin-like pigments, and the increase in diffuse reflectance of the cardiac muscle beneath the endocardial layer. Insights from these experiments suggested simpler algorithms that can be used to acquire and post-process the spectral information required to reveal the lesion sites. Our study is relevant to a growing number of multilayered clinical targets to which spectral approaches are being applied.

A Percutaneous Catheter for In Vivo Hyperspectral Imaging of Cardiac Tissue: Challenges, Solutions and Future Directions.

PURPOSE: Multiple studies have shown that spectral analysis of tissue autofluorescence can be used as a live indicator for various pathophysiological states of cardiac tissue, including ischemia, ablation-induced damage, or scar formation. Yet today there are no percutaneous devices that can detect autofluorescence signals from inside a beating heart. Our aim was to develop a prototype catheter to demonstrate the feasibility of doing so. METHODS AND RESULTS: Here we summarize technical solutions leading to the development of a percutaneous catheter capable of multispectral imaging of intracardiac surfaces. The process included several iterations of light sources, optical filtering, and image acquisition techniques. The developed system included a compliant balloon, 355 nm laser irradiance, a high-sensitivity CCD, bandpass filtering, and image acquisition synchronized with the cardiac cycle. It enabled us to capture autofluorescence images from multiple spectral bands within the visible range while illuminating the endocardial surface with ultraviolet light. Principal component analysis and other spectral unmixing post-processing algorithms were then used to reveal target tissue. CONCLUSION: Based on the success of our prototype system, we are confident that the development of ever more sensitive cameras, recent advances in tunable filters, fiber bundles, and other optical and computational components makes it possible to create percutaneous catheters capable of acquiring hyper or multispectral hypercubes, including those based on autofluorescence, in real-time. This opens the door for widespread use of this methodology for high-resolution intraoperative imaging of internal tissues and organs-including cardiovascular applications.

Autofluorescence properties of balloon polymers used in medical applications.

SIGNIFICANCE: For use in medical balloons and related clinical applications, polymers are usually designed for transparency under illumination with white-light sources. However, when illuminated with ultraviolet (UV) or blue light, most of these materials autofluoresce in the visible range, which can be a concern for modalities that rely on tissue autofluorescence for diagnostic or therapeutic purposes. AIM: A search for published information on spectral properties of polymers that can be used for medical balloon manufacturing revealed a scarcity of published information on this subject. The aim of these studies was to address this gap. APPROACH: The autofluorescence properties of polymers used in medical balloon manufacturing were examined for their suitability for hyperspectral imaging and related applications. Excitation-emission matrices of different balloon materials were acquired within the 320- to 620-nm spectral range. In parallel, autofluorescence profiles from the 420- to 620-nm range were extracted from hyperspectral datasets of the same samples illuminated with UV light. The list of tested polymers included polyurethanes, nylon, polyethylene terephthalate (PET), polyether block amide (PEBAX), vulcanized silicone, thermoplastic elastomers with and without talc, and cyclic olefin copolymers, known by their trade name TOPAS. RESULTS: Each type of polymer exhibited a specific pattern of autofluorescence. Polyurethanes, PET, and thermoplastic elastomers containing talc had the highest autofluorescence values, while sheets made of nylon, PEBAX, and TOPAS exhibited negligible autofluorescence. Hyperspectral imaging was used to illustrate how the choice of specific balloon material can impact the ability of principal component analysis to reveal the ablated cardiac tissue. CONCLUSIONS: The data revealed significant differences between autofluorescence profiles of the polymers and pointed to the most promising balloon materials for clinical implementation of approaches that depend on tissue autofluorescence.

Optical mapping of human embryonic stem cell-derived cardiomyocyte graft electrical activity in injured hearts.

BACKGROUND: Human embryonic stem cell-derived cardiomyocytes (hESC-CMs) show tremendous promise for cardiac regeneration, but the successful development of hESC-CM-based therapies requires improved tools to investigate their electrical behavior in recipient hearts. While optical voltage mapping is a powerful technique for studying myocardial electrical activity ex vivo, we have previously shown that intra-cardiac hESC-CM grafts are not labeled by conventional voltage-sensitive fluorescent dyes. We hypothesized that the water-soluble voltage-sensitive dye di-2-ANEPEQ would label engrafted hESC-CMs and thereby facilitate characterization of graft electrical function and integration. METHODS: We developed and validated a novel optical voltage mapping strategy based on the simultaneous imaging of the calcium-sensitive fluorescent protein GCaMP3, a graft-autonomous reporter of graft activation, and optical action potentials (oAPs) derived from di-2-ANEPEQ, which labels both graft and host myocardium. Cardiomyocytes from three different GCaMP3+ hESC lines (H7, RUES2, or ESI-17) were transplanted into guinea pig models of subacute and chronic infarction, followed by optical mapping at 2 weeks post-transplantation. RESULTS: Use of a water-soluble voltage-sensitive dye revealed pro-arrhythmic properties of GCaMP3+ hESC-CM grafts from all three lines including slow conduction velocity, incomplete host-graft coupling, and spatially heterogeneous patterns of activation that varied beat-to-beat. GCaMP3+ hESC-CMs from the RUES2 and ESI-17 lines both showed prolonged oAP durations both in vitro and in vivo. Although hESC-CMs partially remuscularize the injured hearts, histological evaluation revealed immature graft structure and impaired gap junction expression at this early timepoint. CONCLUSION: Simultaneous imaging of GCaMP3 and di-2-ANEPEQ allowed us to acquire the first unambiguously graft-derived oAPs from hESC-CM-engrafted hearts and yielded critical insights into their arrhythmogenic potential and line-to-line variation.

Clinically relevant concentrations of di (2-ethylhexyl) phthalate (DEHP) uncouple cardiac syncytium.

Di(2-ethylhexyl) phthalate (DEHP) is a widely used plasticizer found in a variety of polyvinyl chloride (PVC) medical products. The results of studies in experimental animals suggest that DEHP leached from flexible PVC tubing may cause health problems in some patient populations. While the cancerogenic and reproductive effects of DEHP are well recognized, little is known about the potential adverse impact of phthalates on the heart. This study examined the effects of clinically relevant concentrations of DEHP on neonatal rat cardiomyocytes. It was found that application of DEHP to a confluent, synchronously beating cardiac cell network, leads to a marked, concentration-dependent decrease in conduction velocity and asynchronous cell beating. The mechanism behind these changes was a loss of gap junctional connexin-43, documented using Western blot analysis, dye-transfer assay and immunofluorescence. In addition to its effect on electrical coupling, DEHP treatment also affected the mechanical movement of myocyte layers. The latter was linked to the decreased stiffness of the underlying fibroblasts, as the amount of triton-insoluble vimentin was significantly decreased in DEHP-treated samples. The data indicate that DEHP, in clinically relevant concentrations, can impair the electrical and mechanical behavior of a cardiac cell network. Applicability of these findings to human patients remains to be established.

Use of GelMA for 3D printing of cardiac myocytes and fibroblasts.

AIM: To 3D print heart tissue, one must understand how the main two types of cardiac cells are affected by the printing process. MATERIALS & METHODS: Effects of gelatin methacryloyl (GelMA) concentration, extruder pressure and duration of UV exposure on survival of cardiac myocytes and fibroblasts were examined using lactate dehydrogenase and LIVE/DEAD assays, bioluminescence imaging and morphological assessment. RESULTS & CONCLUSION: Cell survival within 3D printed cardiomyocyte-laden GelMA constructs was more sensitive to extruder pressure and GelMA concentrations than within 3D fibroblast-laden GelMA constructs. Cells within both types of constructs were adversely impacted by the UV curing step. Use of mixed cell populations and enrichment of bioink formulation with fibronectin led to an improvement of cardiomyocyte survival and spreading.

Generation and escape of local waves from the boundary of uncoupled cardiac tissue.

We aim to understand the formation of abnormal waves of activity from myocardial regions with diminished cell-to-cell coupling. En route to this goal, we studied the behavior of a heterogeneous myocyte network in which a sharp coupling gradient was placed under conditions of increasing network automaticity. Experiments were conducted in monolayers of neonatal rat cardiomyocytes using heptanol and isoproterenol as means of altering cell-to-cell coupling and automaticity, respectively. Experimental findings were explained and expanded using a modified Beeler-Reuter numerical model. The data suggest that the combination of a heterogeneous substrate, a gradient of coupling, and an increase in oscillatory activity of individual cells creates a rich set of behaviors associated with self-generated spiral waves and ectopic sources. Spiral waves feature a flattened shape and a pin-unpin drift type of tip motion. These intercellular waves are action-potential based and can be visualized with either voltage or calcium transient measurements. A source/load mismatch on the interface between the boundary and well-coupled layers can lock wavefronts emanating from both ectopic sources and rotating waves within the inner layers of the coupling gradient. A numerical approach allowed us to explore how 1), the spatial distribution of cells, 2), the amplitude and dispersion of cell automaticity, and 3), the speed at which the coupling gradient moves in space affect wave behavior, including its escape into well-coupled tissue.

Formation of cardiac fibers in Matrigel matrix.

We report a simple in vitro model of cardiac tissue that mimics three-dimensional (3-D) environment and mechanical load conditions and, as such, may serve as a convenient method to study stem cell engraftment or address developmental questions such as cytoskeleton or intercalated disk maturation. To create in vitro cardiac fibers we used Matrigel, a commercially available basement membrane preparation. A semisolid pillow from concentrated Matrigel was overlaid with a suspension of rat neonatal cardiomyocytes in a diluted Matrigel solution. This created an environment in which the multicellular fibers continuously contracted against a mechanical load. The described approach allows continuous structural and functional monitoring of 20-300-micron-thick cardiac fibers and provides easy access to epitopes for immunostaining purposes.

Controlled regional hypoperfusion in Langendorff heart preparations.

We describe a new approach that combines several techniques to allow abnormal electrical and calcium activity to be visualized within hypoperfused myocardial tissue. A flexible microcannula was inserted into the left anterior descending artery of Langendorff perfused rat hearts, an air-tight seal between the coronary artery and the cannula was created, and an HPLC pump was used to deliver a specified flowrate through the microcannula. High resolution optical mapping of NADH/calcium, NADH/voltage or calcium/voltage was then conducted using a dual camera system. The ECG was acquired using surface electrodes. This perfusion technique is superior to occluding a vessel by either a tie or a clamp because it allows precise control of the composition and amount of flow to a defined ischemic bed. Another advantage is that flow can be stopped and resumed remotely, without touching the heart. This allows ectopic beats, or other arrhythmogenic activity, such as alternans, to be recorded immediately after changes in flow are imposed. Altogether, the described method provides a powerful new tool to assess how coronary flow rate affects the degree of local ischemia by the ability to record abnormal patterns of electrical activity and associated intracellular calcium transients with high spatiotemporal resolution from epicardial areas as small as 100 x 100 microm.

Application of unsupervised learning to hyperspectral imaging of cardiac ablation lesions.

Atrial fibrillation is the most common cardiac arrhythmia. It is being effectively treated using the radiofrequency ablation (RFA) procedure, which destroys culprit tissue and creates scars that prevent the spread of abnormal electrical activity. Long-term success of RFA could be improved further if ablation lesions can be directly visualized during the surgery. We have shown that autofluorescence-based hyperspectral imaging (aHSI) can help to identify lesions based on spectral unmixing. We show that use of k -means clustering, an unsupervised learning method, is capable of detecting RFA lesions without a priori knowledge of the lesions' spectral characteristics. We also show that the number of spectral bands required for successful lesion identification can be significantly reduced, enabling the use of increased spectral bandwidth. Together, these findings can help with clinical implementation of a percutaneous aHSI catheter, since by reducing the number of spectral bands one can reduce hypercube acquisition and processing times, and by increasing the spectral width of individual bands one can collect more photons. The latter is of critical importance in low-light applications such as intracardiac aHSI. The ultimate goal of our studies is to help improve clinical outcomes for atrial fibrillation patients.

Optimization of wavelength selection for multispectral image acquisition: a case study of atrial ablation lesions.

In vivo autofluorescence hyperspectral imaging of moving objects can be challenging due to motion artifacts and to the limited amount of acquired photons. To address both limitations, we selectively reduced the number of spectral bands while maintaining accurate target identification. Several downsampling approaches were applied to data obtained from the atrial tissue of adult pigs with sites of radiofrequency ablation lesions. Standard image qualifiers such as the mean square error, the peak signal-to-noise ratio, the structural similarity index map, and an accuracy index of lesion component images were used to quantify the effects of spectral binning, an increased spectral distance between individual bands, as well as random combinations of spectral bands. Results point to several quantitative strategies for deriving combinations of a small number of spectral bands that can successfully detect target tissue. Insights from our studies can be applied to a wide range of applications.

Hyperspectral imaging for label-free in vivo identification of myocardial scars and sites of radiofrequency ablation lesions.

BACKGROUND: Treatment of cardiac arrhythmias often involves ablating viable muscle tissue within or near islands of scarred myocardium. Yet, today there are limited means by which the boundaries of such scars can be visualized during surgery and distinguished from the sites of acute injury caused by radiofrequency (RF) ablation. OBJECTIVE: We sought to explore a hyperspectral imaging (HSI) methodology to delineate and distinguish scar tissue from tissue injury caused by RF ablation. METHODS: RF ablation of the ventricular surface of live rats that underwent thoracotomy was followed by a 2-month animal recovery period. During a second surgery, new RF lesions were placed next to the scarred tissue from the previous ablation procedure. The myocardial infarction model was used as an alternative way to create scar tissue. RESULTS: Excitation-emission matrices acquired from the sites of RF lesions, scar region, and the surrounding unablated tissue revealed multiple spectral changes. These findings justified HSI of the heart surface using illumination with 365 nm UV light while acquiring spectral images within the visible range. Autofluorescence-based HSI enabled to distinguish sites of RF lesions from scar or unablated myocardium in open-chest rats. A pilot version of a percutaneous HSI catheter was used to demonstrate the feasibility of RF lesion visualization in atrial tissue of live pigs. CONCLUSION: HSI based on changes in tissue autofluorescence is a highly effective tool for revealing-in vivo and with high spatial resolution-surface boundaries of myocardial scar and discriminating it from areas of acute necrosis caused by RF ablation.