title: Regulation of the P2X7 receptor permeability to large molecules by extracellular Cl- and Na+.
abstract: Upon continuous stimulation, the pore of the monovalent cation-selective P2X7 receptor (P2X7R) expands to accommodate large molecules such as N-methyl-D-glucamine (NMDG+). How the change in P2X7R permeability is regulated is not known. Here we report that extracellular Cl- (Cl-(o)) regulates the outward current, whereas extracellular Na+ (Na+(o)) regulates the inward current of large molecules by P2X7Rs. The P2X7R-mediated current was measured in parotid acinar and duct cells of wild type and P2X7R-/- mice and in HEK293 cells expressing the human or mouse P2X7R isoforms. In symmetrical NaCl, triethylammonium chloride, and NMDG+ chloride solutions, the P2X7R current followed a linear current/voltage relationship. In symmetrical NaCl, removal of Cl-(o) reduced the inward Na+ current by approximately 35% and the outward Na+ current by only 10%. By contrast, in the absence of Na+(i) and the presence of Na+(o) or NMDG+(o), the removal of Cl-(o) reduced the inward Na+ or NMDG+ currents by 35% but the outward NMDG+ current by >95%. The effect of Cl-(o) was half-maximal at approximately 60 mm. Reducing Cl-(i) from 150 to 10 mm did not reproduce the effects of Cl-(o). All currents were eliminated in P2X7R-/- cells and reproduced by expressing the P2X7Rs in HEK cells. These findings suggest that Cl-(o) primarily regulates the outward P2X7R current of large molecules. When cells dialyzed with NMDG+ were stimulated in the presence of Na+(o), subsequent removal of Na+(o) resulted in a strongly outward rectifying NMDG+ current, indicating maintained high selectivity for Na+ over NMDG+. During continuous incubation in Na+-free medium, the permeability of the P2X7Rs to NMDG+ gradually increased. On the other hand, when the cells were incubated in symmetrical NMDG+ and only then stimulated with ATP, the NMDG+ current by P2X7Rs followed a linear current/voltage relationship and did not change with time. These findings suggest that the P2X7R has a "Na+(o) memory" and that Na+(o) regulates the inward permeability of P2X7Rs to large molecules. The novel regulation of P2X7R outward and inward permeability to large molecules by Cl-(o) and Na+(o), respectively, may have an important protective function, particularly in secretory epithelial cells.
authors.affiliation: Department of Physiology, University of Texas Southwestern Medical Center, Dallas, Texas 75390-9040, USA.
authors.forename: Qin
authors.initials: Q
authors.lastname: Li
authors.forename: Xiang
authors.initials: X
authors.lastname: Luo
authors.forename: Shmuel
authors.initials: S
authors.lastname: Muallem
mesh_terms: D000255:Adenosine Triphosphate; D000818:Animals; D002460:Cell Line; D002713:Chlorine / Q000378:metabolism*; D004594:Electrophysiology; D004847:Epithelial Cells / Q000502:physiology; D006801:Humans; D007477:Ions / Q000378:metabolism; D008536:Meglumine / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D010306:Parotid Gland / Q000166:cytology / Q000502:physiology; D010539:Permeability; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D058486:Receptors, Purinergic P2X7; D012964:Sodium / Q000378:metabolism*
chemical_list: D007477:Ions; C546468:P2RX7 protein, human; C546469:P2rx7 protein, mouse; D018048:Receptors, Purinergic P2; D058486:Receptors, Purinergic P2X7; D002713:Chlorine; D008536:Meglumine; D000255:Adenosine Triphosphate; D012964:Sodium