Journal
CELL CALCIUM
Volume 39, Issue 2, Pages 143-153Publisher
ELSEVIER SCI LTD
DOI: 10.1016/j.ceca.2005.10.001
Keywords
Fura-2; cyclopiazonic acid; ionomycin; calcium stores; endoplasmic reticulum
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Nicotinic acid adenine dinucleotide phosphate (NAADP) has been shown to release intracellular Ca2+ in several types of cells. We have used Ca2+-sensitive fluorescent dyes (Fura-2, Fluo-4) to measure intracellular Ca2+ in astrocytes in culture and in situ. Bath-applied NAADP elicited a reversible and concentration-dependent Ca2+ rise in up to 90% of astrocytes in culture (EC50 = 7 mu M). The NAADP-evoked Ca2+ rise was maintained in the absence of extracellular Ca2+, but was suppressed after depleting the Ca2+ stores of the ER with ATP (20 mu M), with cyclopiazonic acid (10 mu M) or with ionomycin (5 mu M). P-2 receptor antagonist pyridoxalphosphate-6-azophenyl-2'4'-disulfonic acid (PPADS, 100 mu M), IP3 receptor blocker 2-aminoethoxydiphenyl borate (2-APB, 100 mu M) and PLC inhibitor U73122 (10 mu M) also reduced or suppressed the NAADP-evoked Ca2+ rise. NAADP still evoked a Ca2+ response after application Of glyCyl-L-phenylalanine-beta-naphthylamide (GPN, 200 mu M), which permeabilizes lysosomes, or preincubation with H+-ATPase inhibitor bafilomycin A1 (4 mu M) and of p-trifluoromethoxy carbonyl cyanide phenylhydrazone (FCCP, 2 mu M), that impairs mitochondrial Ca2+ handling. In acute brain slices, NAADP (10 mu M) evoked Ca2+ transients in cerebellar Bergmann glial cells and in hippocampal astrocytes. Our results suggest that NAADP recruits Ca2+ from mositol 1,4,5-trisphosphate-sensitive Ca2+ stores in mammalian astrocytes, at least partly by activating metabotropic P2Y receptors. (C) 2005 Elsevier B.V. All rights reserved.
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