Cardiac arrhythmias are a leading cause of death in humans and occur in diverse conditions. The proposed research seeks to identify and characterize fundamental mechanisms that underlie fatal cardiac arrhythmias. Specific cellular and molecular events that trigger arrhythmias will be examined to test the hypothesis that changes in subcellular calcium signaling contribute to arrhythmogenesis. Animal models of altered electrical activity in the heart will be studied at the single cells level using whole patch clamp methods and confocal calcium imaging. Isolated cardiac myocytes from control and transgenic animals and cells expressing specific constructs will be used in the planned work. Preliminary results have demonstrated calcium-dependent links between altered electrical behavior and the expression of specific cellular proteins that are being examined in Project 1 (Russo), Project 2 (Marks) and Project 3 (Kass). The proteins of particular interest include beta1AR, beta2AR, RyR2, FKB12, FKBP12.6, SCN5A and mutations of these proteins. The proposed work examines how expression of the target proteins affects intracellular [Ca2+]i and also Ca2+-dependent membrane currents. This examination will explore the importance of the action potential shape and duration on [Ca2+]i signaling in the proposed experimental models. Additionally the relationship between SR Ca2+ content and Ca2+ release (as measured by Ca2+ sparks and the global (Ca2+) transient) will be examined with these molecular models. The experiments carried out in this project should thus provide fundamental new information on the arrhythmogenic roles played by the betaAR signaling system, sarcolemmal ion channels and the intracellular Ca2+ release channels.