PROJECT SUMMARY: Atrial fibrillation (AF) is the most common cardiac arrhythmia (affecting ~1-2% of the general population), resulting in markedly reduced quality of life and increased mortality, due to a combination of altered hemodynamics, progressive atrial and ventricular dysfunction, and embolic stroke. Many diseases and conditions, like heart failure, are known to contribute to pathological changes leading to AF. Limitations in current therapy allow AF paroxysms to progress to persistent and chronic AF, as a result of extensive atrial structural and electrical changes that facilitate AF maintenance (“AF begets AF”). The development of urgently needed new strategies for AF treatment hinges upon improved understanding of how abnormalities in cellular function trigger and sustain arrhythmia in atrial tissue. At the cellular level, a hallmark structural change of many chronic cardiac diseases is degradation of the intricate membrane architecture that couples cardiac electrical excitation to intracellular Ca2+ release and myocardial contraction (EC coupling) – i.e., the transverse tubule (TT) structures, which project orthogonally from the cell surface to its interior and thereby synchronize EC coupling throughout the cell. Degradation of the TT architecture is generally associated with arrhythmia, but it is not yet clear whether TT loss is a direct contributor to arrhythmia, a compensatory maladaptation, or an epiphenomenon. This is even less clear in atria, as atrial myocytes exhibit a vastly variable range of TT architectures, with prominent axial tubules. Further, TT degradation induced by the process of isolating atrial myocytes (vs. denser TTs in intact tissues) and challenges in experimentally detubulating intact cardiac tissue has so far limited the design of mechanistic myocyte and tissue studies. As a result, the literature surrounding the role of subcellular structural (ultrastructural) remodeling in AF has remained fractured, and currently we know relatively little about its role in contributing to AF pathophysiology. The overarching goal of this proposal is to discriminate the role of changes in atrial myocyte ultrastructure from other disease-associated sequelae by combining detailed multi-level experimental analyses of rabbit atrial myocytes and rabbit and human atrial tissues with extensive quantitative multi-scale computational modeling. The project will develop and validate a suite of modeling tools used to investigate the mechanisms by which: (1) naturally occurring variations in atrial TTs influence EC coupling and membrane stability in isolated atrial myocytes; (2) tissue gradients in TT organization influence tissue-level electrophysiological and EC coupling outcomes; (3) ultrastructural remodeling synergizes with ionic remodeling to favor atrial arrhythmogenesis in atrial cardiomyopathy. We contend that quantifying the role of atrial ultrastructure in AF pathology may shed new mechanistic insight into AF management. Each a...