Title: An Electrophysiology Assay for the Fast Characterization of GABAA Ion Channel Modulators
Abstract: The GABAA receptors belong to a family of ligand-gated ion channels mediating fast synaptic transmission. They are drawing great attention in the pharmaceutical industry due to their potential role in the development of new therapeutics affecting anxiety, sleep disorders, and muscle relaxation. However, ligand-gated ion channel screening has been hampered by the lack of suitable high throughput electrophysiology platforms. The existing electrophysiology screening platform (IonWorks) is unable to apply ligands rapidly during current recording, which can be a major drawback for inactivating channels. While some studies have shown that it is possible to record the GABA tail current and use that information during a screen, such methods have an inherently lower signal to noise ratio and cannot be used on faster desensitizers. Here we present the use of a novel electrophysiology screening platform integrating a microfluidics network for the study of GABAA receptor pharmacology. This platform features fast (<100ms) solution exchange coupled with simultaneous data recording. A novel assay could monitor GABA response in real time, and obtain a 3 point EC50 dose curve within 1 minute. The GABAA α1β3γ2 expressing HEK cells from Millipore were used for this study. The channel was targeted with agonists, including GABA and muscimol, inhibitors (picrotoxin, bicuculline, and gabazine), and positive modulators, including diazepam, zolpidem and chlordiazepoxide. The positive modulators produced concentration dependent augmentation of the GABA EC20 response. The pharmacology data determined using this method was consistent with the literature values obtained using other platforms. Statistical data for inter and intra-plate reproducibility, current stability, and Z-values, is used to validate this approach.