title: Characterization of P2X3, P2Y1 and P2Y4 receptors in cultured HEK293-hP2X3 cells and their inhibition by ethanol and trichloroethanol.
abstract: Membrane currents and changes in the intracellular Ca2+ concentration ([Ca2+]i) were measured in HEK293 cells transfected with the human P2X3 receptor (HEK293-hP2X3). RT-PCR and immunocytochemistry indicated the additional presence of endogenous P2Y1 and to some extent P2Y4 receptors. P2 receptor agonists induced inward currents in HEK293-hP2X3 cells with the rank order of potency alpha,beta-meATP approximately ATP > ADP-beta-S > UTP. A comparable rise in [Ca2+]i was observed after the slow superfusion of ATP, ADP-beta-S and UTP; alpha,beta-meATP was ineffective. These data, in conjunction with results obtained by using the P2 receptor antagonists TNP-ATP, PPADS and MRS2179 indicate that the current response to alpha,beta-meATP is due to P2X3 receptor activation, while the ATP-induced rise in [Ca2+]i is evoked by P2Y1 and P2Y4 receptor activation. TCE depressed the alpha,beta-meATP current in a manner compatible with a non-competitive antagonism. The ATP-induced increase of [Ca2+]i was much less sensitive to the inhibitory effect of TCE than the current response to alpha,beta-meATP. The present study indicates that in HEK293-hP2X3 cells, TCE, but not ethanol, potently inhibits ligand-gated P2X3 receptors and, in addition, moderately interferes with G protein-coupled P2Y1 and P2Y4 receptors. Such an effect may be relevant for the interruption of pain transmission in dorsal root ganglion neurons following ingestion of chloral hydrate or trichloroethylene.
authors.affiliation: Rudolf-Boehm-Department of Pharmacology and Toxicology, Department of Forensic Medicine, University of Leipzig, Leipzig, Germany.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Fischer
authors.forename: Kerstin
authors.initials: K
authors.lastname: Wirkner
authors.forename: Marco
authors.initials: M
authors.lastname: Weber
authors.forename: Christoph
authors.initials: C
authors.lastname: Eberts
authors.forename: Laszlo
authors.initials: L
authors.lastname: Köles
authors.forename: Robert
authors.initials: R
authors.lastname: Reinhardt
authors.forename: Heike
authors.initials: H
authors.lastname: Franke
authors.forename: Clemens
authors.initials: C
authors.lastname: Allgaier
authors.forename: Clemens
authors.initials: C
authors.lastname: Gillen
authors.forename: Peter
authors.initials: P
authors.lastname: Illes
mesh_terms: D000244:Adenosine Diphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D000431:Ethanol / Q000494:pharmacology*; D005023:Ethylene Chlorohydrin / Q000031:analogs & derivatives* / Q000494:pharmacology*; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D018408:Patch-Clamp Techniques; D058919:Purinergic P2 Receptor Antagonists; D012333:RNA, Messenger / Q000378:metabolism; D018048:Receptors, Purinergic P2 / Q000235:genetics / Q000378:metabolism; D058477:Receptors, Purinergic P2X3; D058472:Receptors, Purinergic P2Y1; D013873:Thionucleotides / Q000494:pharmacology; D014162:Transfection; D014544:Uridine Triphosphate / Q000494:pharmacology
chemical_list: C546407:P2RX3 protein, human; C546382:P2RY1 protein, human; D058919:Purinergic P2 Receptor Antagonists; D012333:RNA, Messenger; D018048:Receptors, Purinergic P2; D058477:Receptors, Purinergic P2X3; D058472:Receptors, Purinergic P2Y1; D013873:Thionucleotides; C104749:purinoceptor P2Y4; C030812:adenosine 5'-O-(2-thiodiphosphate); D000431:Ethanol; D000244:Adenosine Diphosphate; D005023:Ethylene Chlorohydrin; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; C005849:2,2,2-trichloroethanol; C002630:alpha,beta-methyleneadenosine 5'-triphosphate; D002118:Calcium; D014544:Uridine Triphosphate