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Establishing the role of Purkinje fibres in the development of arrhythmias for the prevention of Sudden Cardiac Death: Insights from a combined in vivo and ex vivo study

Final Report Summary - PSCD (Establishing the role of Purkinje fibres in the development of arrhythmias for the prevention of Sudden Cardiac Death: Insights from a combined in vivo and ex vivo study)

Cardiovascular disease is the first cause of mortality worldwide accounting for 29% of deaths. In Europe and the USA, 80% of 350,000 Sudden cardiac deaths (SCD) occurring each year are due to VF. France alone accounts for 50,000 SCDs, which is equivalent to the cumulative mortality of breast, lung and colon cancers, and AIDS. Currently, the majority of individuals dying suddenly cannot be recognized pre-emptively. Sudden cardiac death (SCD) following ventricular fibrillation (VF) is a major cause of mortality in the industrialised world. VF is characterised by chaotic electrical activity in the ventricles resulting in severe cardiac dysfunction. Thus far, the underlying mechanisms for the initiation and maintenance of such arrhythmias have not yet been conclusively identified and may depend on the specific condition of the patient. Significant emphasis has been placed on a specialized cell type, the Purkinje fibers, which are responsible for conduction to occur between the atria to the ventricles. Due to challenges to indentify accurately the macro structure of the Purkinje network, where the Purkinje interact with the myocardium, properties of Purkinje fibers and myocardium at junctional sites or to characterise electrophysiological properties at such sites, much remains unknown regarding the role of Purkinje fibers in arrhythmia formation or maintenance. The current project sought to resolve some of these issues.
Electrophysiological gradients of electrical activity underlie electrical propagation and repolarization patterns in the heart and moreover, determine the potential for arrhythmigeneisis and dynamics of electrical abnormalities. Therefore, an early phase of the project was dedicated to understanding this fundemental aspect. Specifically, the impact of electrical coupling on repolarization gradients within the myocardium as a result of differences in activation sequence, action potential morphology and tissue geometry. It was observed that modulation of the repolarization gradients due to electrical coupling was especially pronounced in small hearts with murine-like action potentials and both tissue architecture and action potential morphology played an important role. These results were published as a stand alone research article in Frontiers in Physiology [1].

Gradients of repolarization are observed both in base-apex and transmural planes. These gradients play a role in normal cardiac function and help synchronizing ventricular contraction. However, abnormal repolarization gradients, especially transmurally, have been linked to conduction abnormalities and the onset of arrhythmias. Studies thus far during more than 30 years of debate have only considered different myocardial phenotypes to impact the transmural gradient of repolarization. Furthermore, very little is currently known regarding the impact of repolarization gradients in heart failure of humans. In this study we included a model of human heart failure to directly compare with non-failing myocardium. Our computational results theorize that

Purkinje also have the potential to determine repolarization gradients in the myocardium and therefore we proposed a novel source for repolarization heterogeneity in the ventricles of non-failing and failing myocardium. For the first time, the impact of Purkinje fibers on transmural gradients of heterogeneity were verified experimentally using optical mapping in sheep left ventricles during the current reporting period. It was found that at sites of origin of activation when stimulating through the Purkinje network, action potential duration was longer when the Purkinje muscle-junction resided within the optical integration volume (the volume of tissue beneath the imaged surface that contributes to the optical signal). Such regions of prolonged action potential duration were consistent with observations of islands of long action potential durations thought previously only to be attributed to a distinct myocardial cell type. These results have since been published in Cardiovascular Research [2].

Recordings of electrical activity from the heart without causing damage to tissue is limited to surface recordings. Many efforts have been made to improve estimations of eletrical activation time using various electrical mapping systems. Such systems use activcation times measured from all recording nodes and find the best solution for a global activation map. This drastically limits the imapct of noise and allows good approximations even signal quality is poor. A similar approach was developed for use with optical recordings. A global activation map approachfor optical signals was presented and published as a conference proceedings at the 35th Annual International conference of the IEEE: EMBC, Osaka, Japan [3]. This publication formed the first step towards correlating in vivo elctrical data to ex vivo optical recordings. This approach will inevitably pave the way towards determing the 3D organization of normal and arrhythmic electrical activity in vivo.

The Young Scientist's, using expertise in optical recordings of electrical activity in the heart, sought to develop novel optical approaches to reconstruct electrical activity in 3D throughout the ventricular free wall. Different optical imaging modalities were explored along with assessing the optical spectra to optimise signals for a novel dual wavelength – dual modality optical imaging technique. This mulitmodal approach enabled sequential acquisition of multiple volumes within the myocardium for comparison. This method revealed transmural delays of activation when stimulating one surface and initiating impulses to propagate towards the opposing surface. Furthermore, the data shows the extent of transmural wave front propagation at high resolution (<1 mm) across a large field of view. Therefore, a major outcome of this project has been the development of the first non-destructive approach to reconstruct electrical activity in 3D in ex vivo issue of large animal hearts. This technique was presented and published at the 40th International Congress of Electrocardiology, International Society of Electrocardiology, 2013, Glasgow, UK[4] where the Young Scientist was awarded 3rd prize for the Young Investigator's award.

A major mechanism for ventricular arrhythmias is by re-entry-induced ventricular tachycardia. The Young Scientist has, during the course of the project, identified a novel pathway that can exist to sustain ventricular tachycardia in normal healthy myocardium of sheep. For the first time, continuous conduction between the septum and right ventricular free wall was identified along a muscular free-running band thought originally to moderate ventricular distension, termed the moderator band. This band has since been found to contain a major fascicle of the Purkinje network. However, the relatively slower conducting muscular layer forms the substrate for sustained re-entrant ventricular tachycardia that can be initiated by premature ventricular contractions originating from the moderator band. This important discovery should call for re-evaluation of the target sites for ablation therapy in the clinic.

The current project has provided a novel insight in to electrophysiological heterogeneities in the ventricles due to electrical coupling of the myocardium to Purkinje fibers. This will impact additively to our understanding of the mechanisms of arrhythmias initiated by electrophysiological gradients. The novel approach for 3D reconstruction of electrical activity will significantly impact on our understanding of the 3D dynamics of electrical propagation. Particularly in cases of disease as the approach is versatile and transferabe between different experimental models, species and electrophysiological activation patterns for other research studies. As such, research to better characterize diseases based on understanding of the electrical activity at all depths of the myocardium shall be achieved, instead of jugements for treatments based solely on surface recordings. Particularly ablation therapy, the most effective anti-arrhythmic treatment, shall benefit. Ulitimately, ablation accuracy, surgery time and success rates shall see dramatic improvements with appropriate studies using the novel technology developed during this project.

1. Walton RD, Benson AP, Hardy ME, White E, Bernus O. 2013, Electrophysiological and structural determinants of electrotonic modulation of repolarization by the activation sequence; Frontiers in Physiology; 4:281. doi: 10.3389/fphys.2013.00281.
2. Richard D. Walton, PhD, Marine E. Martinez, Msc, Martin S. Bishop, PhD, Meleze Hocini, MD, Michel Haissaguerre, MD, Gernot Planck, PhD, Olivier Bernus, PhD and Edward J. Vigmond, 2014; Influence of the Purkinje-muscle junction on transmural repolarization heterogeneity; Cardiovascular Research; 103(4): 629-640. Doi: 10.1093/cvr/cvu165.
3. Walton RD, Bernus O, Dubois R. 2013; A novel approach for deriving global activation maps from non-averaged cardiac optical signals. Engineering in Medicine and Biology (EMBC) 2013 35th Annual International conference of the IEEE. Osaka, Japan. doi: 10.1109/EMBC.2013.6609864
4. Walton RD, Bernus O, 2013; Dual excitation wavelength optical imaging of transmural electrophysiological heterogeneity in pig ventricles; Journal of Electrocardiology; 46(4), e35.