Development of a functional cell-based HTS assay for the identification mGluR4 modulators.
- Author:
Yaling ZHANG
1
;
Yanqiu BAI
Author Information
1. Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 101300, China. zhangyl@big.ac.cn
- Publication Type:Journal Article
- MeSH:
Aminobutyrates;
pharmacology;
Cell Line;
DNA, Complementary;
genetics;
Drug Evaluation, Preclinical;
Humans;
Kidney;
cytology;
embryology;
Phosphoserine;
pharmacology;
Plasmids;
genetics;
Receptors, Metabotropic Glutamate;
agonists;
antagonists & inhibitors;
genetics;
Transfection
- From:
Chinese Journal of Biotechnology
2009;25(3):457-463
- CountryChina
- Language:Chinese
-
Abstract:
To identify metabotropic glutamate receptor 4 (mGluR4) modulators by Ca2+ influx assay, we developed the functional cell-based high throughput-screening (HTS) assay. The human mGluR4 cDNA was transfected into HEK-293 stably expressing promiscuous G-protein (Ga alpha15) cells. Recombinant stable mGluR4 cell line was selected under Zeocin and validated by Ca2+ influx assay. The assay was optimized on loading time of Fluo Calcium Indicator, Dimethyl sulfoxide (DMSO) tolerance and sodium hydroxide (NaOH) tolerance using agonist (L-Glutamic acid (L-Glu)) of mGluR4. The rank order of the agonist potency for the stable human mGluR4 cell line was L-(+)-2-Amino-4-phosphonobutyric acid (L-AP4) > L-Serine-O-phosphate (L-SOP) > L-Glu, and of the antagonist potency was (RS)-alpha-Methylserine-O-phosphate (MSOP) > (RS)-alpha-Methyl-4-phosphonophenylglycine (MPPG). Z' factor value of the cell line in 96- and 384-well plate format was 0.80 and 0.65. Our data indicate a successful development of functional human mGluR4 recombinant stable cell line that was suitable for high throughput screening to identify mGluR4 agonist/antagonist.
