Production of NAADP and its role in Ca super(2+) mobilization associated with lysosomes in coronary arterial myocytes

• Published in: American journal of physiology. Heart and circulatory physiology 2006-07, Vol.291 (1), p.H274-H282
• Main Authors: Zhang, F, Zhang, G, Zhang, A Y, Koeberl, MJ, Wallander, E, Li, P-L
• Format: Article
• Language: English
• ISSN: 0363-6135
• DOI: 10.1152/ajpheart.01064.2005

The present study was designed to determine the production of nicotinic acid adenine dinucleotide phosphate (NAADP) and its role associated with lysosomes in mediating endothelin-1 (ET-1)-induced vasoconstriction in coronary arteries. HPLC assay showed that NAADP was produced in coronary arterial smooth muscle cells (CASMCs) via endogenous ADP-ribosyl cyclase. Fluorescence microscopic analysis of intracellular Ca super(2+) concentration ([Ca super(2+)] sub(i)) in CASMCs revealed that exogenous 100 nM NAADP increased [Car super(+)] sub(i) by 711 plus or minus 47 nM. Lipid bilayer experiments, however, demonstrated that NAADP did not directly activate ryanodine (Rya) receptor Ca super(2+) release channels on the sarcoplasmic reticulum. In CASMCs pretreated with 100 nM bafilomycin A1 (Baf), an inhibitor of lysosomal Ca super(2+) release and vacuolar proton pump function, NAADP-induced [Ca super(2+)] sub(i) increase was significantly abolished. Moreover, ET-1 significantly increased NAADP formation in CASMCs and resulted in the rise of [Ca super(2+)] sub(i) in these cells with a large increase in global Ca super(2+) level of 1,815 plus or minus 84 nM. Interestingly, before this large Ca super(2+) increase, a small Ca super(2+) spike with an increase in [Ca super(2+)] sub(i) of 529 plus or minus 32 nM was observed. In the presence of Baf (100 nM), this ET-1-induced two-phase [Ca super(2+)] sub(i) response was completely abolished, whereas Rya (50 mu M) only markedly blocked the ET-1-induced large global Ca super(2+) increase. Functional studies showed that 100 nM Baf significantly attenuated ET-1-induced maximal constriction from 82.26 plus or minus 4.42% to 51.80 plus or minus 4.36%. Our results suggest that a lysosome-mediated Ca super(2+) regulatory mechanism via NAADP contributes to ET-1-induced Ca super(2+) mobilization in CASMCs and consequent vasoconstriction of coronary arteries.