Premium
Increase of intracellular Ca 2+ by P2Y but not P2X receptors in cultured cortical multipolar neurons of the rat
Author(s) -
Fischer Wolfgang,
Nörenberg Wolfgang,
Franke Heike,
Schaefer Michael,
Illes Peter
Publication year - 2009
Publication title -
journal of comparative neurology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.855
H-Index - 209
eISSN - 1096-9861
pISSN - 0021-9967
DOI - 10.1002/cne.22079
Subject(s) - ppads , biology , p2y receptor , ionotropic effect , cyclopiazonic acid , p2 receptor , receptor , purinergic receptor , intracellular , microbiology and biotechnology , glutamate receptor , suramin , biochemistry , biophysics
The expression and functionality of P2X/P2Y receptor subtypes in multipolar nonpyramidal neurons of mixed cortical cell cultures were investigated by means of immunocytochemistry and fura‐2 microfluorimetry. The morphological studies revealed that most of the neurons are immunoreactive for GABA and express a range of P2X/P2Y receptors, predominantly of the P2X 2,4,6 and P2Y 1,2 subtypes. P2X 1 and P2X 7 receptor immunoreactivity (IR) was found on thin axon‐like processes and presynaptic structures, respectively. Application of ATP caused a small concentration‐dependent increase in intracellular Ca 2+ concentration ([Ca 2+ ] i ) in most investigated neurons, whereas only about the half of these cells responded to 2′,3′‐ O ‐(benzoyl‐4‐benzoyl)‐ATP (BzATP), ADPβS, 2MeSADP, or 2MeSATP and even fewer cells to UTP. In contrast, α,β‐meATP, UDP, and UDP‐glucose failed to produce any [Ca 2+ ] i signaling. The response to ATP itself was inhibited by pyridoxal‐5′‐phosphate‐6‐azophenyl‐2′,4′‐disulfonic acid (PPADS), Reactive Blue 2, 2′‐deoxy‐N 6 ‐methyl adenosine 3′,5′‐diphosphate (MRS2179), and suramin (300 μM) as well as by a cyclopiazonic acid‐induced depletion of intracellular Ca 2+ stores. A Ca 2+ ‐free external medium tended to decrease the ATP‐induced [Ca 2+ ] i transients, although this action did not reach statistical significance. Various blockers of voltage‐sensitive Ca 2+ channels and the gap junction inhibitor carbenoxolone did not interfere with the effect of ATP, whereas a combination of the ionotropic glutamate receptor antagonists D(–)‐2‐amino‐5‐phosphonopentanoic acid (AP5) and 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (CNQX) decreased it. Cross‐desensitization experiments between ADPβS or UTP and ATP suggested that ATP acts on the one hand via P2Y 1,2 receptors and on the other hand by additional signaling mechanisms. These mechanisms may involve the release of glutamate (which in consequence activates ionotropic glutamate receptors) and the entry of Ca 2+ via store‐operated Ca 2+ channels. Evidence for the presence of functional P2X receptors, in particular P2X 7 , remains elusive. J. Comp. Neurol. 516:343–359, 2009. © 2009 Wiley‐Liss, Inc.