Development and validation of a high-throughput calcium mobilization assay for the orphan receptor GPR88 View Full Text


Ontology type: schema:ScholarlyArticle      Open Access: True


Article Info

DATE

2017-03-27

AUTHORS

Ann M. Decker, Elaine A. Gay, Kelly M. Mathews, Taylor C. Rosa, Tiffany L. Langston, Rangan Maitra, Chunyang Jin

ABSTRACT

BackgroundGPR88 is an orphan G protein-coupled receptor highly expressed in the striatum and is implicated in basal ganglia-associated disorders. However, the receptor functions of GPR88 are still largely unknown due to the lack of potent and selective ligands appropriate for central nervous system investigation. Development of a high-throughput screening assay for GPR88 should facilitate the discovery of novel ligands to probe GPR88 functions.MethodsIn this paper, we describe the development of a CHO-Gαqi5-GPR88 cell-based calcium mobilization assay. The assay takes advantage of functional coupling of GPR88 with the promiscuous Gαqi5 protein and consequent mobilization of intracellular calcium, which can be measured in a 384-well format with a Fluorescent Imaging Plate Reader.ResultsThe CHO-Gαqi5-GPR88 cell-based calcium mobilization assay was validated by the structure-activity relationship study of known GPR88 agonist (1R,2R)-2-PCCA analogues. The assay was automated and miniaturized to a 384-well format, and was deemed robust and reproducible with a Z’-factor of 0.72 and tolerated dimethyl sulfoxide to a final concentration of 2%. Screening a pilot neurotransmitter library consisting of 228 compounds yielded 10 hits, but none of the hits were confirmed as GPR88 agonists in follow-up assays.ConclusionsWe have developed a high-throughput calcium mobilization assay for the orphan receptor GPR88. This calcium mobilization assay can be used to identify several different types of GPR88 ligands including agonists, competitive and noncompetitive antagonists, inverse agonists, and allosteric modulators. These ligands will serve as valuable tools to probe signaling mechanisms and in vivo functions of GPR88, and could expedite development of novel therapies for diseases potentially mediated by GPR88. More... »

PAGES

23

References to SciGraph publications

Identifiers

URI

http://scigraph.springernature.com/pub.10.1186/s12929-017-0330-3

DOI

http://dx.doi.org/10.1186/s12929-017-0330-3

DIMENSIONS

https://app.dimensions.ai/details/publication/pub.1084251125

PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/28347302


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