Identification and characterization of an endogenous P2X 7 (P2Z) receptor in CHO‐K1 cells

1 CHO‐K1 cells were examined for their cellular responses to the P2 receptor agonist, 2′‐ and 3′‐O‐(4‐benzoylbenzoyl)‐ATP (DbATP), and for the presence of mRNA for P2X receptors. 2 Reverse transcriptase‐polymerase chain reactions, using primers directed against the rat P2X subunits, detected the presence of P2X 7 but not P2X 1 ‐P2X 6 subunits. 3 DbATP (EC 50 ∼100 μ m ) evoked non‐desensitizing inward currents which reversed at ∼0mV, suggesting activation of a non‐selective cation channel. ATP also evoked inward currents but was less potent than DbATP. 4 DbATP also stimulated the accumulation of 45 calcium ( 45 Ca 2+ ) and the DNA binding dye, YO‐PRO‐1, in CHO‐K1 cells. Both responses were inhibited by NaCl and MgCl 2 . In 280 m m sucrose buffer, 45 Ca 2+ accumulation was measurable within 10–20 s of agonist addition, whereas YO‐PRO‐1 accumulation was only detectable after 8 min. ATP and ATPγS were also agonists but were less potent than DbATP, while UTP, 2‐methylthio ATP, ADP and αβmethylene ATP were inactive at concentrations up to 100 μ m . 5 DbATP increased lactate dehydrogenase release from CHO‐K1 cells, suggesting cell lysis, although this effect was only pronounced after 60–90 min. 6 These data suggest that CHO‐K1 cells express an endogenous P2X 7 receptor which can be activated by DbATP to cause a rapid inward current and accumulation of 45 Ca 2+ . Prolonged receptor activation results in a delayed, increased permeability to larger molecules such as YO‐PRO‐1 and ultimately leads to cell lysis. Importantly, the presence of an endogenous P2X 7 receptor should be considered when these cells are used to study recombinant P2X receptors.British Journal of Pharmacology (1998) 125 , 1194–1201; doi: 10.1038/sj.bjp.0702205