Establishment of an assay for P2X(7) receptor-mediated cell death

Abstract

The P2X(7) receptor, an ATP-gated cation channel, induces cell death in immune cells and is involved in neurodegenerative diseases. Although the receptor plays various roles in these diseases, the cellular mechanisms involved are poorly understood and antagonists are limited. Here, the development of a cell-based assay for human P2X(7) receptor is reported. We established permanent lines of HEK 293 cells expressing a high level of hP2X(7) receptor. Functional activity of the hP2X(7) receptor was confirmed by whole-cell patch recording of ATP-induced ion currents. Prolonged exposure to ATP resulted in death of the hP2X(7)-expressing HEK 293 cells and this cell death could be quantified. Two known P2X(7) antagonists, PPADS and KN-62, blocked ATP-induced death in a concentration-dependent manner. Thus, this assay can be used to screen for new antagonists of hP2X(7) receptors.