NAADP is a Physiologically-Relevant Calcium Mobilising Compound in Atrial Myocytes

Abstract

NAADP is a calcium mobilising compound acting via a two-pool mechanism (1). In both ventricular and atrial myocytes, NAADP increases cellular calcium transients through enhancement of SR calcium load (2,3).In ventricular myocytes Ned-19, a non-competitive antagonist of the NAADP receptor, abolishes the contractile response to NAADP(4). Furthermore, both bafilomycin and Ned-19 decrease whole-cell calcium transients and, after Ned-19 exposure, bafilomycin has no further effect (4).Tissue level of NAADP is raised after isoprenaline perfusion in whole heart, consistent with a role for NAADP in the beta-adrenergic response (2).This project aimed to measure the response of atrial myocytes to NAADP and investigate the relevance of the pathway to beta-adrenergic signalling.Enzymatically-isolated guinea pig atrial myocytes were loaded with fluo-5F calcium indicator dye and NPE-caged NAADP (5μM). Cells were stimulated during superfusion with physiological salt solution and NAADP released by UV-photolysis. Whole-cell calcium transients were measured with a photomultiplier system. All data are given mean±SEM with statistical significance considered as p<0.05.Flash-photolysis of NAADP increased atrial calcium transients by 67±15% (p<0.05, n=7), becoming significant 3min after photolysis and reaching a peak at 12min. This increase was prevented by 1μM patch-applied Ned-19.In field-stimulated atrial myocytes, bafilomycin (100nM) significantly reduced cellular calcium transients (by 19±5%, n=6) and inhibited enhancement of the calcium transient by isoprenaline (3nM), reducing it from a 63±8% (n=9) to a 36±9% (n=6) increase (p<0.05).These data support the two-pool model for NAADP actions in cardiac atrial myocytes, suggest the NAADP system is constitutively active and indicate a physiological role in beta-adrenergic signalling.1)Churchill and Galione EMBOJ (2001) 20: 2666-2671.2)Macgregor et al. Cell Calcium (2007) 41:537-546.3)Collins et al. Cell Calcium (2011) In press.4)Elson et al. Biophysical Journal (2011) 100, 81a-82a.