Characterizing ligand-gated ion channel receptors with genetically encoded Ca2++ sensors |
| |
Authors: | Yamauchi John G Nemecz Ákos Nguyen Quoc Thang Muller Arnaud Schroeder Lee F Talley Todd T Lindstrom Jon Kleinfeld David Taylor Palmer |
| |
Affiliation: | Department of Pharmacology, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California San Diego, La Jolla, California, United States of America. |
| |
Abstract: | We present a cell based system and experimental approach to characterize agonist and antagonist selectivity for ligand-gated ion channels (LGIC) by developing sensor cells stably expressing a Ca(2+) permeable LGIC and a genetically encoded F?rster (or fluorescence) resonance energy transfer (FRET)-based calcium sensor. In particular, we describe separate lines with human α7 and human α4β2 nicotinic acetylcholine receptors, mouse 5-HT(3A) serotonin receptors and a chimera of human α7/mouse 5-HT(3A) receptors. Complete concentration-response curves for agonists and Schild plots of antagonists were generated from these sensors and the results validate known pharmacology of the receptors tested. Concentration-response relations can be generated from either the initial rate or maximal amplitudes of FRET-signal. Although assaying at a medium throughput level, this pharmacological fluorescence detection technique employs a clonal line for stability and has versatility for screening laboratory generated congeners as agonists or antagonists on multiple subtypes of ligand-gated ion channels. The clonal sensor lines are also compatible with in vivo usage to measure indirectly receptor activation by endogenous neurotransmitters. |
| |
Keywords: | |
本文献已被 PubMed 等数据库收录! |
|