Development of a Colorimetric Method for Functional Chloride Channel Assay

Abstract

Anion channels play significant physiological roles in humans and animals. However, the effort of screening for anion channel modulators was limited by the available assay technologies. This report discusses the development of a cell-based functional chloride channel assay using iodine as the chloride channel functional indicator. Iodine concentrations were measured with modified Sandell-Kolthoff reaction using colorimetric detection. The assay was rapid and quantitative. When WSS-1 cells were activated by γ-aminobutyric acid (GABA) in the condition that γ-aminobutyric acid type A receptor (GABAA receptor) conducted outwardly rectifying chloride channel function, the EC50 of GABA was 7.69 μM. IC50 swere 0.53 μM for bicuculline and 3.1 μM for picrotoxin, respectively, in the presence of 10 μM GABA. When Capan-1 cells were activated by forskolin, the EC50 was 0.14 μM. The assay can also be applied to inwardly rectifying anion channels as exemplified by GABAA channel with an EC50 of 294 μM. Thus, the assay is universal and reliable and can be used for anion channel high-throughput screening.