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Influence of Extracellular Monovalent Cations on Pore and Gating Properties of P2X₇ Receptor-Operated Single-Channel Currents

T. Riedel ^*, G. Schmalzing  and F. Markwardt ^*

^* Julius-Bernstein-Institute for Physiology, Martin Luther University Halle, Halle/Saale, Germany; and RWTH Aachen University, Department of Molecular Pharmacology, Aachen, Germany

Correspondence: Address reprint requests to F. Markwardt, Julius-Bernstein Institute for Physiology, Martin Luther University Halle, Magdeburger Str. 6, D-06097 Halle/Saale, Germany. Tel.: 49-345-557-1390; Fax.: 49-345-552-7899; E-mail: fritz.markwardt{at}medizin.uni-halle.de.

Using the patch-clamp method, we studied the influence of external alkali and organic monovalent cations on the single-channel properties of the adenosine triphosphate (ATP)-activated recombinant human P2X₇ receptor. The slope conductance of the hP2X₇ channel decreased and the reversal potential was shifted to more negative values as the ionic diameter of the organic test cations increased. From the relationship between single-channel conductance and the dimensions of the inward current carrier, the narrowest portion of the pore was estimated to have a mean diameter of 8.5 Å. Single-channel kinetics and permeation properties remained unchanged during receptor activation by up to 1 mM ATP^4– for >1 min, arguing against a molecular correlate of pore dilation at the single P2X₇ channel level. Substitution of extracellular Na⁺ by any other alkali or organic cation drastically increased the open probability of the channels by prolonging the mean open time. This effect seems to be mediated allosterically through an extracellular voltage-dependent Na⁺ binding site with a K_d of 5 mM Na⁺ at a membrane potential of –120 mV. The modulation of the ATP-induced hP2X₇ receptor gating by extracellular Na⁺ could be well described by altering the rate constant from the open to the neighboring closed state in a C-C-C-O kinetic receptor model. We suggest that P2X₇ receptor-induced depolarization and associated K⁺-efflux may reduce Na⁺ occupancy of the regulatory Na⁺ binding site and thus increase the efficacy of ATP^4– in a feed-forward manner in P2X₇ receptor-expressing cells.