Inhibition Mechanism of the Recombinant Rat P2X2 Receptor in Glial Cells by Suramin and TNP-ATP
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- 作者：Cleber A. Trujillo ; Arthur A. Nery ; Antonio Henrique B. Martins ; Paromita Majumder ; Fernando A. Gonzalez ; and Henning Ulrich
- 刊名：Biochemistry
- 出版年：2006
- 出版时间：January 10, 2006
- 年：2006
- 卷：45
- 期：1
- 页码：224 - 233
- 全文大小：128K
- 年卷期：v.45,no.1(January 10, 2006)
- ISSN：1520-4995
文摘
P2X receptors play an important role in communication between cells in the nervous system.Therefore, understanding the mechanisms of inhibition of these receptors is important for the developmentof new tools for drug discovery. Our objective has been to determine the pharmacological activity of theantagonist suramin, the most important antagonist of purinergic receptor function, as well as to demonstrateits noncompetitive inhibition and confirm a competitive mechanism between ATP and TNP-ATP in 1321N1glial cells stably transfected with the recombinant rat P2X2 receptor. A radioligand binding assay wasemployed to determine whether suramin, TNP-ATP, and ATP compete for the same binding site on thereceptor. TNP-ATP displaced [
-32P]ATP, whereas suramin did not interfere with [-32P]ATP-receptorbinding. To determine the inhibition mechanism relevant for channel opening, currents obtained in fastkinetic whole-cell recording experiments, following stimulation of cells by ATP in the presence of suramin,were compared to those obtained by ATP in the presence of TNP-ATP. Supported by a mathematicalmodel for receptor kinetics [Breitinger, H. G., Geetha, N., and Hess, G. P. (2001) Biochemistry 40, 8419-8429], the inhibition factors were plotted as functions of inhibitor or agonist concentrations. Analysis ofthe data indicated a competitive inhibition mechanism for TNP-ATP and a noncompetitive inhibition forsuramin. Taken together, both data support a noncompetitive inhibition mechanism of the rat recombinantP2X2 receptor by suramin, confirm the competitive inhibition by TNP-ATP, and allow the prediction ofa model for P2X2 receptor inhibition.
-32P]ATP, whereas suramin did not interfere with [-32P]ATP-receptorbinding. To determine the inhibition mechanism relevant for channel opening, currents obtained in fastkinetic whole-cell recording experiments, following stimulation of cells by ATP in the presence of suramin,were compared to those obtained by ATP in the presence of TNP-ATP. Supported by a mathematicalmodel for receptor kinetics [Breitinger, H. G., Geetha, N., and Hess, G. P. (2001) Biochemistry 40, 8419-8429], the inhibition factors were plotted as functions of inhibitor or agonist concentrations. Analysis ofthe data indicated a competitive inhibition mechanism for TNP-ATP and a noncompetitive inhibition forsuramin. Taken together, both data support a noncompetitive inhibition mechanism of the rat recombinantP2X2 receptor by suramin, confirm the competitive inhibition by TNP-ATP, and allow the prediction ofa model for P2X2 receptor inhibition.