The development of advanced technologies for electrophysiological mapping of cardiac propagation has been a defining factor in recent successes both in basic understanding of arrhythmia mechanisms and clinical applications such as mapping-guided radiofrequency ablation. However, one of the major challenges both in basic and clinical cardiac electrophysiology remains the mapping of abnormally conducting arrhythmogenic regions hidden inside the myocardial wall. The ultimate goal of this proposal is to develop a minimally invasive 3D tomographic optical mapping technique utilizing novel near-infrared (NIR) voltage-sensitive dyes, which will significantly enhance one's ability to study electrical excitation inside the myocardial wall. During the previous period of support, we have conducted extensive modeling and experimental studies that indicate the feasibility of our method. The next step will be to implement, validate, and assess the potential of the new technique for 3D mapping of normal myocardium and physiologically relevant arrhythmogenic ischemic substrates. The Specific Aims of this proposal are: 1) Build a fast optical tomographic system for the time-resolved 3D imaging of intra-mural electrical waves in isolated coronary-perfused ventricular wall preparations. 2) Perform the 3D reconstructions of intramural electrical excitation wave in Tyrode-perfused and blood-perfused pig ventricular wall preparations. 3) Test the accuracy of the reconstructions under conditions simulating non- uniform staining patterns characteristic of arrhythmogenic substrates (ischemic border, infarction scar). 4) Utilize the new techniques to determine the role of 3D geometry of perfusion territories in arrhythmogenesis during acute regional ischemia. We will test the hypothesis that the anatomy of the perfusion territory of a given coronary artery defines the geometry of the ischemic region and is a major factor that determines the pattern of electrical propagation emerging after occlusion of that artery. Successful completion of the proposed Specific Aims will produce, validate, and test both under normal and clinically relevant conditions a fast new tomographic method for the 3D optical imaging of intra-myocardial excitation. It will also provide new mechanistic information regarding the links between the anatomy of perfusion territories and ischemia-related conduction abnormalities. PUBLIC HEALTH RELEVANCE: The goal of the proposed studies is to develop an optical imaging technology for detecting sources of arrhythmia hidden deep inside the myocardial wall. Such sources emerge during the blockage of arteries feeding cardiac muscle and often cause sudden cardiac death. The new technology will help to understand the mechanisms of initiation of such sources, which is the key to developing effective anti-arrhythmic therapies and prevention of sudden cardiac death.