Huda Asfour

Bisphenol A exposure and cardiac electrical conduction in excised rat hearts.

Background: Bisphenol A (BPA) is used to produce polycarbonate plastics and epoxy resins that are widely used in everyday products, such as food and beverage containers, toys, and medical devices. Human biomonitoring studies have suggested that a large proportion of the population may be exposed to BPA. Recent epidemiological studies have reported correlations between increased urinary BPA concentrations and cardiovascular disease, yet the direct effects of BPA on the heart are unknown. Objectives: The goal of our study was to measure the effect of BPA (0.1–100 μM) on cardiac impulse propagation ex vivo using excised whole hearts from adult female rats. Methods: We measured atrial and ventricular activation times during sinus and paced rhythms using epicardial electrodes and optical mapping of transmembrane potential in excised rat hearts exposed to BPA via perfusate media. Atrioventricular activation intervals and epicardial conduction velocities were computed using recorded activation times. Results: Cardiac BPA exposure resulted in prolonged PR segment and decreased epicardial conduction velocity (0.1–100 μM BPA), prolonged action potential duration (1–100 μM BPA), and delayed atrioventricular conduction (10–100 μM BPA). These effects were observed after acute exposure (≤ 15 min), underscoring the potential detrimental effects of continuous BPA exposure. The highest BPA concentration used (100 μM) resulted in prolonged QRS intervals and dropped ventricular beats, and eventually resulted in complete heart block. Conclusions: Our results show that acute BPA exposure slowed electrical conduction in excised hearts from female rats. These findings emphasize the importance of examining BPA’s effect on heart electrophysiology and determining whether chronic in vivo exposure can cause or exacerbate conduction abnormalities in patients with preexisting heart conditions and in other high-risk populations. Citation: Posnack NG, Jaimes R III, Asfour H, Swift LM, Wengrowski AM, Sarvazyan N, Kay MW. 2014. Bisphenol A exposure and cardiac electrical conduction in excised rat hearts. Environ Health Perspect 122:384–390; http://dx.doi.org/10.1289/ehp.1206157

Signal Decomposition of Transmembrane Voltage-Sensitive Dye Fluorescence Using a Multiresolution Wavelet Analysis

Fluorescence imaging of transmembrane voltage-sensitive dyes is used to study electrical activation in cardiac tissue. However, the fluorescence signals, typically, have low SNRs and may be contaminated with motion artifact. In this report, we introduce a new processing approach for fluoresced transmembrane potentials (fTmps) that is based upon a discrete wavelet transform. We show how fTmp signals can be decomposed and reconstructed to form three subsignals that contain signal noise (noise signal), the early depolarization phase of the action potential (rTmp signal), and motion artifact (rMA signal). A coiflet4 wavelet is used for fTmp decomposition and reconstruction of these subsignals. Results using fTmp signals that are contaminated with motion artifact indicate that the approach is a useful processing step to remove baseline drift, reduce noise, and reveal wavefronts. It streamlines the preprocessing of fTmps for the subsequent measurement of activation times and conduction velocities. It is a promising approach for studying wavefronts without aggressive mechanical tissue constraint or electromechanical uncoupling agents and is, useful for single-camera systems that do not provide for ratiometric imaging.

NADH Fluorescence Imaging of Isolated Biventricular Working Rabbit Hearts

Since its inception by Langendorff1, the isolated perfused heart remains a prominent tool for studying cardiac physiology2. However, it is not well-suited for studies of cardiac metabolism, which require the heart to perform work within the context of physiologic preload and afterload pressures. Neely introduced modifications to the Langendorff technique to establish appropriate left ventricular (LV) preload and afterload pressures3. The model is known as the isolated LV working heart model and has been used extensively to study LV performance and metabolism4-6. This model, however, does not provide a properly loaded right ventricle (RV). Demmy et al. first reported a biventricular model as a modification of the LV working heart model7, 8. They found that stroke volume, cardiac output, and pressure development improved in hearts converted from working LV mode to biventricular working mode8. A properly loaded RV also diminishes abnormal pressure gradients across the septum to improve septal function. Biventricular working hearts have been shown to maintain aortic output, pulmonary flow, mean aortic pressure, heart rate, and myocardial ATP levels for up to 3 hours8. When studying the metabolic effects of myocardial injury, such as ischemia, it is often necessary to identify the location of the affected tissue. This can be done by imaging the fluorescence of NADH (the reduced form of nicotinamide adenine dinucleotide)9-11, a coenzyme found in large quantities in the mitochondria. NADH fluorescence (fNADH) displays a near linearly inverse relationship with local oxygen concentration12 and provides a measure of mitochondrial redox state13. fNADH imaging during hypoxic and ischemic conditions has been used as a dye-free method to identify hypoxic regions14, 15 and to monitor the progression of hypoxic conditions over time10. The objective of the method is to monitor the mitochondrial redox state of biventricular working hearts during protocols that alter the rate of myocyte metabolism or induce hypoxia or create a combination of the two. Hearts from New Zealand white rabbits were connected to a biventricular working heart system (Hugo Sachs Elektronik) and perfused with modified Krebs-Henseleit solution16 at 37 °C. Aortic, LV, pulmonary artery, and left & right atrial pressures were recorded. Electrical activity was measured using a monophasic action potential electrode. To image fNADH, light from a mercury lamp was filtered (350±25 nm) and used to illuminate the epicardium. Emitted light was filtered (460±20 nm) and imaged using a CCD camera. Changes in the epicardial fNADH of biventricular working hearts during different pacing rates are presented. The combination of the heart model and fNADH imaging provides a new and valuable experimental tool for studying acute cardiac pathologies within the context of realistic physiological conditions.

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.

Seeing the Invisible: Revealing Atrial Ablation Lesions Using Hyperspectral Imaging Approach

Background Currently, there are limited means for high-resolution monitoring of tissue injury during radiofrequency ablation procedures. Objective To develop the next generation of visualization catheters that can reveal irreversible atrial muscle damage caused by ablation and identify viability gaps between the lesions. Methods Radiofrequency lesions were placed on the endocardial surfaces of excised human and bovine atria and left ventricles of blood perfused rat hearts. Tissue was illuminated with 365nm light and a series of images were acquired from individual spectral bands within 420-720nm range. By extracting spectral profiles of individual pixels and spectral unmixing, the relative contribution of ablated and unablated spectra to each pixel was then displayed. Results of spectral unmixing were compared to lesion pathology. Results RF ablation caused significant changes in the tissue autofluorescence profile. The magnitude of these spectral changes in human left atrium was relatively small (< 10% of peak fluorescence value), yet highly significant. Spectral unmixing of hyperspectral datasets enabled high spatial resolution, in-situ delineation of radiofrequency lesion boundaries without the need for exogenous markers. Lesion dimensions derived from hyperspectral imaging approach strongly correlated with histological outcomes. Presence of blood within the myocardium decreased the amplitude of the autofluorescence spectra while having minimal effect on their overall shapes. As a result, the ability of hyperspectral imaging to delineate ablation lesions in vivo was not affected. Conclusions Hyperspectral imaging greatly increases the contrast between ablated and unablated tissue enabling visualization of viability gaps at clinically relevant locations. Data supports the possibility for developing percutaneous hyperspectral catheters for high-resolution ablation guidance.

Preprocessing of fluoresced transmembrane potential signals for cardiac optical mapping

Fluorescence imaging of transmembrane voltage-sensitive dyes is used to study electrical activation in cardiac tissue. However, fluorescence signals typically have a low signal to noise ratio that can be contaminated with motion artifacts. We describe an alternative processing approach for fluoresced transmembrane potentials (fTmps) using the wavelet multiresolution analysis. We show that fTmp signals can be decomposed and reconstructed to form three sub-signals that contain signal noise (noise signal), the early depolarization phase of the action potential (rTmp signal), and motion artifact (rMA signal). Discrete wavelet transform is used with coiflet 4 scaling and wavelet functions for fTmp decomposition and reconstruction of these sub-signals. Our results show that this type of analysis can be used to remove baseline drift, reduce noise, and reveal wavefronts. It streamlines the preprocessing of fTmps for subsequent measurement of activation times and conduction velocities. The approach is promising for studying wave fronts without aggressive mechanical tissue constraint or electromechanical uncoupling agents, and it is particularly useful for single camera systems that do not provide for ratiometric imaging.

Lesion detection for cardiac ablation from auto-fluorescence hyperspectral images

Direct visualization of the ablated region in the left atrium during radiofrequency ablation (RFA) surgery for treating atrial fibrillation (AF) can improve therapy success rates. Our visualization approach is auto-fluorescence hyperspectral imaging (aHSI), which constructs each hypercube containing 31 auto-fluorescence images of the tissue. We wish to use the spectral information to characterize ablated lesions as being successful or not. In this paper, we reshaped one hypercube to a 2D matrix. Each row (sample) in the matrix represents a pixel in the spatial dimension, and the matrix has 31 columns corresponding to 31 spectral features. Then, we applied k-means clustering to detect ablated regions without a priori knowledge about the lesion. We introduced an accuracy index to evaluate the results of k-means by comparing with the reference truth images quantitatively. To speed-up the detection process, we implemented a grouping procedure to decrease the number of features. The 31 features were divided into four contiguous disjoint groups. In each group, the summation of values yielded a new feature. By the same evaluation method, we found the best 4-feature groups to adequately detect the lesions from all possible combinations. The average accuracy for detection by k-means (k=10) using 31 features was about 74% of reference truth images. And, for using the best grouped 4 features, it was about 95% of that using 31 features. The time cost of 4-feature clustering is about 41% of the 31-feature clustering. We expect that the reduction of time for both acquisition and processing will make possible intraoperative real-time display of ablation status.

Anatomical and Optical Properties of Atrial Tissue: Search for a Suitable Animal Model.

The purpose of this study was to evaluate structural and optical properties of atrial tissue from common animal models and to compare it with human atria. We aimed to do this in a format that will be useful for development of better ablation tools and/or new means for visualizing atrial lesions. Human atrial tissue from clinically relevant age group was compared and contrasted with atrial tissue of large animal models commonly available for research purposes. These included pigs, sheep, dogs and cows. The presented data include area measurements of smooth atrial surface available for ablation and estimates of thickness of collagen and muscle for five different species. We also described methods to quantify presence of collagen and overall thickness of atrial wall. Provided information enables placement of atrial lesions to locations with clinically relevant atrial wall thickness and macroscopic structure ultimately helping investigators to develop better ablation and imaging tools. It also highlights the impact of collagen thickness on optical measurements and lesion visualization.

Decomposition of Fluoresced Transmembrane Potentials Using Multiresolution Wavelet Analysis

Fluorescence imaging of transmembrane voltage- sensitive dyes is used to study electrical activation in cardiac tissue. However, fluorescence signals suffer from sensitivity to motion due to loss of registration between the target and the imaging device. We introduce a new processing approach for fluoresced transmembrane potentials (fTmps) that is based upon the discreet wavelet transform. We show how fTmp signals can be decomposed and reconstructed to form three sub-signals that contain signal noise (noise signal), the early depolarization phase of the action potential (rTmp signal), and motion artifact (rMA signal). Rat hearts stained with RH237 were used to obtain fTmp signals contaminated with motion artifact. fTmp signals acquired from the epicardial surface of the heart were decomposed and reconstructed using coiflet4 wavelet. Results indicate that the approach is a useful processing step to remove baseline drift, reduce noise, and reveal wave fronts. In addition, local motion artifact amplitudes can be measured using rMA signals. Multiresolution wavelet analysis can be used to study wave fronts without aggressive mechanical tissue constraint or electromechanical uncoupling agents and is particularly useful for single camera systems that do not provide for ratiometric imaging.

Chemical Ablation Of Purkinje Fibers Diminishes Spontaneous Activity In A Rat Model Of Regional Ischemia And Reperfusion

Spontaneous activity and arrhythmias associated with acute local ischemia and reperfusion were studied in isolated Langendorff-perfused hearts from healthy Sprague-Dawley rats (n=16). Epicardial fluorescence imaging of transmembrane potential and NADH were used to relate sources of electrical activity to changes in mitochondrial redox state caused by local ischemia. The left anterior descending coronary artery was cannulated and the flow of perfusate to the cannula was controlled by a high-pressure/low-flow HPLC pump. Studies were conducted using a local ischemia/reperfusion protocol that consisted of 10 min of normal flow, 20 min of regional LV ischemia, followed by 20 minutes of reduced flow reperfusion, and then 20 min of normal flow reperfusion. Control hearts (n=9) were compared with hearts in which the endocardium (containing the Purkinje fibers) was chemically ablated by applying a Lugols iodine solution to the ventricular cavities (n=7). The ablation significantly reduced spontaneous activity in each phase of the protocol. Specifically, during acute regional ischemia, spontaneous activity was reduced by 80% (p<0.005); by 70% during low-flow reperfusion (p<0.005); and by 85% during full-flow reperfusion (p<0.001). Omission of blebbistatin, an electro-mechanical uncoupling agent, did not change the diminishing effect of the ablation on spontaneous activity (n=3). Epicardial imaging showed that spontaneous ectopic beats were manifested as concentric epicardial breakthrough patterns, located near spatiotemporal gradients of NADH fluorescence. These data strongly suggest that in un-paced hearts from healthy rats that are perfused with Tyrodes solution, the main mechanism of spontaneous ectopic activity associated with either ischemia, low-flow or full-flow reperfusion is activation of local Purkinje fibers.