Problems caused by high concentration of ATP on activation of the P2X(7) receptor in bone marrow cells loaded with the Ca2+ fluorophore fura-2

Problems caused by high concentration of ATP on activation of the P2X(7) receptor in bone marrow cells loaded with the Ca2+ fluorophore fura-2

Autor Paredes-Gamero, E. J. Google Scholar
Franca, J. P. Google Scholar
Moraes, AAFS Google Scholar
Aguilar, M. O. Google Scholar
Oshiro, M. E. Google Scholar
Ferreira, A. T. Google Scholar
Instituição Universidade Federal de São Paulo (UNIFESP)
Resumo Fura-2 is one of the most used fluorophore for measuring intracellular calcium concentration ([Ca2+](i)). in mouse bone marrow cell suspensions ATP produces a biphasic effect: till 1 mM, ATP produces increases in [Ca2+](i); from 1 mM on an increase is observed, that is followed by the decrease in the 340/380 nm ratio (R340/380). At high ATP (4 mM) concentration fura-2 leaked from loaded bone marrow cell suspensions. We observed that ATP decreases fluorescence in the absorption and excitation spectra of fura-2, consequently the emitted one is decreased including the isobestic point (360 nm). ATP analogs: BzATP, ATPgammaS and UTP, but not alphabetaATP, ADP or AMP, promote decrease of fluorescence in the isobestic point of fura-2. the physical/chemical process that reduces the absorption and excitation of fura-2 by ATP is unknown. the P2X(7) inhibitors, Mg2+ (5 mM), OxATP (300 muM) and Brilliant Blue (100 nM), blocked the efflux of fura-2 and ATP-induced R340/380 decrease. the J774 cell line and mononuclear cells with a higher expression of P2X(7) receptors show the same decrease in R340/380 as that induced by ATP. in the HL-60 cell line, myeloid cells and erythroblasts extracted from bone marrow, such effect does not occur. It is concluded that the use of the fluorescent Ca2+ indicator fura-2 does not allow the correct measurement of [Ca2+](i) in these cells in the presence of a higher concentration of ATP which activated the P2X(7) receptor.
Palavra-chave ATP
P2X(7) receptor
fura-2
bone marrow
Idioma Inglês
Data de publicação 2004-11-01
Publicado em Journal of Fluorescence. New York: Kluwer Academic/plenum Publ, v. 14, n. 6, p. 711-722, 2004.
ISSN 1053-0509 (Sherpa/Romeo, fator de impacto)
Publicador Kluwer Academic/plenum Publ
Extensão 711-722
Fonte http://dx.doi.org/10.1023/B:JOFL.0000047221.51493.e3
Direito de acesso Acesso restrito
Tipo Artigo
Web of Science WOS:000224920900006
Endereço permanente http://repositorio.unifesp.br/handle/11600/28013

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