Characterization of P2X3, P2Y1 and P2Y4 receptors in cultured HEK293-hP2X3 cells and their inhibition by ethanol and trichloroethanol.
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.
Fischer
Wolfgang
W
Rudolf-Boehm-Department of Pharmacology and Toxicology, Department of Forensic Medicine, University of Leipzig, Leipzig, Germany.
Wirkner
Kerstin
K
Weber
Marco
M
Eberts
Christoph
C
Köles
Laszlo
L
Reinhardt
Robert
R
Franke
Heike
H
Allgaier
Clemens
C
Gillen
Clemens
C
Illes
Peter
P
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
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