Characterization of P2X 3 , P2Y 1 and P2Y 4 receptors in cultured HEK293-hP2X 3 cells and their inhibition byethanol and trichloroethanol

Membrane currents and changes in the intracellular Ca 2+ concentration ((Ca 2+ )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 a,b-meATPATP > ADP-b-S > UTP. A comparable rise in (Ca 2+ )i was observed after the slow superfusion of ATP, ADP- b-S and UTP; a,b-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 a,b-meATP is due to P2X3 receptor activation, while the ATP-induced rise in (Ca 2+ )i is evoked by P2Y1 and P2Y4 receptor activation. TCE depressed the a,b-meATP current in a manner compatible with a non-com- petitive antagonism. The ATP-induced increase of (Ca 2+ )i was much less sensitive to the inhibitory effect of TCE than the current response to a,b-meATP. The present study indicates that in HEK293-hP2X3 cells, TCE, but not ethanol, potently inhibits ligand-gated P2X3 receptors and, in addition, moder- ately 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