Development and validation of ligand-gated ion channel assays using the Qube 384 automated electrophysiology platform

著者: Abigail Marklew, Juha Kammonen, Emma Richardson and Gary Clark

Ligand-gated ion channels are of particular interest to the pharmaceutical industry for the treatment of diseases from a variety of therapeutic areas including CNS disorders, respiratory disease and chronic pain. Ligand-gated ion channels have historically been investigated using fluorescence-based and low throughput patch-clamp techniques. However, with the development of the Qube 384 automated patch-clamp system, the rapid exchange of liquid and direct measurement of ion channel currents on a millisecond timescale is now possible at a greater throughput than previously possible. Here, we have used the Qube platform to develop assays against two ligand-gated families: 1) the P2X receptor and 2) the GABAA receptor families. The P2X family is comprised of 7 family members, which are cation permeable and gated by the binding of extracellular ATP. We have assessed both agonist and antagonist pharmacology of 4 members of the P2X family, P2X1, P2X2, P2X3 and P2X4, as well as two species homologs, rP2X3 and gpP2X3. The GABAA α1β3γ2 receptor is a chloride permeable ion channel gated by the binding of GABA. We utilized stacked liquid addition to assess the open state kinetics of the channel and to investigate the effects of a positive allosteric modulator on channel function. As such, we have successfully characterized and developed assays for both the P2X receptor and GABAA receptor families and present EC/IC50 data for antagonists and positive allosteric modulators.