During a cardiac ventricular action potential (AP), a small Ca flux across the transverse tubule (TT) membrane activates ryanodine receptor (RyR) channels on the sarcoplasmic reticululum (SR), a process called Ca-induced Ca release (CICR). During diastole, single RyRs can also spontaneously open (albeit infrequently). When one does, the SR Ca released may trigger a localized bout of inter-RyR CICR (a Ca spark). Abnormally frequent or large sparks can evoke propagating Ca waves. By driving surface electrogenic Na-Ca exchange, waves may generate delayed afterdepolarizations (DADs). DADs can be life-threatening arrhythmogenic events and are observed in both catecholaminergic polymorphic ventricular tachycardia (CPVT) and heart failure (HF). A RyR DAD control nexus is diastolic inter-RyR CICR initiation, which is driven by an individual spontaneous RyR opening. The decisive opening produces a local cytosolic Ca signal that either evokes inter-RyR CICR or not. Most diastolic openings do not. But, the likelihood that one will increases during HF and CPVT, elevating DAD propensity Carvedilol is a commonly used FDA-approved β-blocker. We recently (2011) made a non-β-blocking carvedilol derivative (VK-II-86) that (like carvedilol) reduces RyR open time (OT) and consequently eliminated waves, DADs and CPVT in model mice (50). We removed -blocking because it dose-limited the drug’s RyR- targeted action. We have made/screened 100+ carvedilol derivatives and identified some new very promising agents that can dosed to provide an optimal RyR anti-DAD action, independent of β-block need. Here, we will define the RyR DAD control nexus, determine how CPVT/HF distort operation of this nexus and identify RyR-targeted drugs that normalize the DAD-driving nexus output. We will test the hypothesis that distortion of the spatiotemporal cytosolic Ca signal created by an individual spontaneous diastolic RyR opening is a key pathogenically shared DAD control nexus that can be therapeutically normalized using novel RyR- targeted agents. This multi-PI proposal combines an almost unprecedented combination of single RyR recording, intracellular Ca imaging and muscle cell biology expertise to address our mechanism-to-therapeutic intervention hypothesis. The specific aims are 1) Determine how CPVT and HF distorts the DAD control nexus and whether those distortions are nexus control factor-specific, disease-dependent and/or part of a continuum of RyR dysfunction and 2) Identify RyR-targeted anti-DAD drugs that normalize pathologically distorted single RyR cytosolic Ca signals and thus limit threat of life-threatening arrhythmias in HF and CPVT. This study will provides a strong mechanistic foundation for understanding DAD origination and its involvement in cardiac disease and thus represents a needed step toward rational design of better therapeutic interventions.

Project Narrative Intracellular calcium signals drive a myriad of cellular phenomena. In the heart, calcium release from intracellular storage sites drives contraction. This calcium release usually does not occur in resting healthy hearts. Malfunction of calcium release control in resting hearts is pathogenic and may trigger arrhythmias and contribute to heart failure. This project defines how calcium release control is distorted by disease and identifies new agents that normalize those distortions. Consequently, this study may lead to the development of a new and promising class of anti-arrhythmic agents and have direct implications for the prevention and treatment of cardiac arrhythmias.