Abstract 13235: Enhanced Receptor-independent Nucleoside Diphosphate Kinase Mediated G s Protein Signaling Contributes to Arrhythmogenic Calcium Mishandling in Atrial Fibrillation

Abstract

Background: Enhanced G-protein related cyclic AMP (cAMP) signaling and abnormal Ca 2+ handling due to dysfunctional ryanodine receptor type-2 (RyR2) are implicated in atrial fibrillation (AF). Nucleoside diphosphate kinase (NDPK) forms complexes with G s proteins, increasing cAMP signaling via receptor-independent G protein activation. Here, we studied the possible proarrhythmic role of NDPKs in clinical (chronic AF, cAF) and experimental AF. Methods: NDPK and G protein expression were assessed by immunoblotting in atria from sinus rhythm (Ctl) and cAF patients; dogs undergoing AF-mimicking tachycardia remodeling (ATR, 400 bpm, 7 days) or AF-promoting chronic atrial ischemia; and canine atrial cardiomyocytes (CMs) paced in vitro for 24-48 hrs at 1 (P1) or 3 Hz (P3). cAMP was measured by immunoassay. Immunostaining was used for protein visualization. Spontaneous sarcoplasmic reticulum (SR) Ca 2+ release events (SCaEs) in CMs overexpressing NDPKs or EGFP-control (adenoviral gene-transfer) were recorded with epifluorescence (Indo-1). Results: NDPK-B, NDPK-C, Gα s and Gβ 1 were upregulated in cAF patient atria (Fig A). ATR and chronic atrial ischemia increased NDPK-B expression in canine atria (Fig B). In vitro tachypacing reproduced AF effects on NDPK/Gα s in CMs (Fig C). Accordingly, cAMP levels were higher in cAF patients and P3-paced CMs (Fig D-E). Adenoviral NDPK-B and NDPK-C overexpression increased cAMP levels (Fig F). NDPK-B, NDPK-C and Gα s co-localized at the plasma membrane when upregulated by tachypacing canine CMs. SCaEs, a proarrhythmic consequence of diastolic SR Ca 2+ leak, were enhanced in NDPK-B or NDPK-C overexpressing CMs, and the latency from last regular SR Ca 2+ release was reduced (Fig G). Conclusions: Receptor-independent NDPK-mediated G s -protein/cAMP signaling is upregulated in clinical and experimental AF, causes arrhythmogenic SCaEs, and is likely a novel contributor to ectopy-inducing atrial triggered activity due to SR Ca 2+ leak.