Ionic effects on human recombinant P2X(7) receptor function

The actions of monovalent and divalent ions on the P2X(7) receptor have bee n assessed by measuring their effect on responses to the P2 receptor agonis t, 2'- and 3'-O-(4-benzoyl-benzoyl)-ATP (DbATP), in HEK293 cells expressing the human recombinant P2X(7) receptor. Tn these cells, DbATP increased the cellular accumulation of the DNA binding, fluorescent dye, YO-PRO-1. The p otency of DbATP to elicit this effect was decreased by both calcium and mag nesium ions. In addition, when the pH was increased above 8 or reduced belo w 6.5, the potency of DbATP was less than obtained at pH 7.5. Monovalent io ns also affected the P2X(7) receptor such that the potency of DbATP was 19- fold higher in NaCl-free buffer containing 280 mM sucrose (pEC(50)=6.48) th an in 140 mM NaCl containing buffer (pEC(50)=5.19). Monovalent cations diff erentially affected the potency of DbATP. Thus, when the chloride concentra tion was maintained at 140 mM, PEC50 values for DbATP were 6.14, 5.87 and 5 .19 when the counter cation was 140 mM choline, potassium or sodium, respec tively. Monovalent anions also differentially affected the potency of DbATP and in the presence of 140 mM sodium ions, PEC50 values for DbATP were 6.1 4, 6.07, 5.19 and 4.53, respectively, when the counter anion was 140 mM asp artate, glutamate, chloride or iodide. The inhibitory effect of monovalent anions on P2X(7) receptor function was also observed in electrophysiologica l studies. Thus in sodium glutamate containing buffer the potency of DbATP (pEC(50)=5.55) was approximately 22-fold higher than in NaCl containing buf fer (pEC(50)=4.20). This study has demonstrated that P2X(7) receptor function can be markedly a ffected by a wide range of ions and that physiological concentrations of so dium and chloride ions, as well as divalent cations, contribute to the low potency of ATP as an agonist at this receptor.