Cellular fluorescent high-throughput screening assays of the ATP-gated P2X7 receptor

The P2X₇ receptor (P2X₇R) is an important member of the P2X family of ligand-gated ion channels that respond to ATP as the endogenous agonist. Studies suggest that P2X₇R plays a potentially pivotal role in a variety of physiological functions, including peripheral and central neuronal transmission, smooth muscle contraction, and inflammation. Thus, P2X₇R may be a potential target for drug development. Here, we used a FlexStation to examine the function of recombinant P2X₇R stably expressed in human embryonic kidney 293 cells and to compare three high-throughput screening assays: a membrane potential assay, an ethidium bromide uptake assay, and a calcium influx assay. We found that all three assays were suitable for the analysis of P2X₇R, but the calcium influx assay was the most robust and is the best choice as a first high-throughput screening assay when embarking on a P2X₇R drug discovery project.