title: [Mg(2+) inhibits ATP-activated current mediated by rat P2X4 receptors expressed in Xenopus oocytes].
abstract: To investigate the modulation of Mg(2+) on rat P2X4 receptors and its underlying mechanism, we transcribed cDNA coding for wild-type and mutant P2X4 receptors to cRNA in vitro, injected the cRNA to oocytes of Xenopus laevis using the microinjection technique and revealed the effect of Mg(2+) on ATP-activated currents (I(ATP)) mediated by P2X4 receptors using the two-electrode whole-cell voltage clamp technique. The effects of extracellular Mg(2+) on I(ATP) were as follows: (1) In oocytes expressing P2X4 receptors, Mg(2+) with concentration ranging from 0.5-10 mmol/L inhibited the amplitude of I(ATP) in a concentration-dependent and reversible manner, with a 50% inhibitory concentration value (IC(50)) of (1.24 ± 0.07) mmol/L for current activated by 100 μmol/L ATP. (2) Mg(2+) (1 mmol/L) shifted the dose-response curve for I(ATP) right-downward without changing the EC(50), but reduced the maximal current (E(max)) by (42.0 ± 2.1)%. (3) After being preincubated with Mg(2+) for 80 s, the inhibitory effect of the Mg(2+) on I(ATP) reached the maximum. (4) The inhibition of Mg(2+) on I(ATP) was independent of membrane potential from -120 mV to +60 mV. (5) Compared with the current activated by 100 μmol/L ATP in the wild-type P2X4 receptors, mutant P2X4 D280Q responded to the application of 100 μmol/L ATP with a smaller current. The peak current was only (4.12 ± 0.15)% of that seen in wild-type receptors. Mutant P2X4 D280E responded to ATP stimulation with a current similar to that observed in cells expressing wild-type receptors. (6) When Asp280 was removed from P2X4,  the current amplitude of I(ATP) was increased almost one-fold, and Mg(2+) with concentration ranging from 0.5-10 mmol/L did not affect the I(ATP) significantly. The results suggest that Mg(2+) inhibits I(ATP) mediated by P2X4 receptors non-competitively, reversibly, concentration-dependently, time-dependently and voltage-independently. The inhibitory effect of Mg(2+) might be realized by acting on the site Asp280 of the P2X4 receptors.
authors.affiliation: Department of Physiology, Medical College, Wuhan University of Science and Technology, Wuhan 430065, China.
authors.forename: Fang
authors.initials: F
authors.lastname: Peng
authors.forename: Yu-Qin
authors.initials: YQ
authors.lastname: Zhang
authors.forename: Yan
authors.initials: Y
authors.lastname: Zeng
authors.forename: Yan-Ling
authors.initials: YL
authors.lastname: Zhou
mesh_terms: D000255:Adenosine Triphosphate / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D000818:Animals; D005260:Female; D008274:Magnesium / Q000494:pharmacology*; D008564:Membrane Potentials / Q000187:drug effects; D009865:Oocytes / Q000378:metabolism* / Q000502:physiology; D018408:Patch-Clamp Techniques; D051381:Rats; D058848:Receptors, Purinergic P2X4 / Q000235:genetics / Q000502:physiology*; D014982:Xenopus laevis
chemical_list: D058848:Receptors, Purinergic P2X4; D000255:Adenosine Triphosphate; D008274:Magnesium