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Journal of Biomolecular Screening
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Article

Ecdysone-Based System for Controlled Inducible Expression of Metabotropic Glutamate Receptor Subtypes 2, 5, and 8

Patrick M. Downey*, Gianluca Lozza, Roberta Petrò, Enrica Diodato, Chiara Foglia, Federica Bottazzoli, Rossella Brusa, Tatiana Asquini, Angelo Reggiani, Mariagrazia Grilli

Schering-Plough Research Institute, Neurobiology Research, San Raffaele Science Park, Milan, Italy.

* To whom correspondence should be addressed. E-mail: patrick.downey{at}spcorp.com.


  Abstract

Stable and inducible expression of human metabotropic glutamate receptor types 2, 5, and 8 was achieved in HEK293 cells using the ecdysone inducible system. Treatment of the respective cell lines with ponasterone A resulted in time- and concentration-dependent induction of receptor expression. In all cases, the functional activation of receptors was determined by measuring increases in intracellular calcium. The physiologically G{alpha}i-coupled receptors mGluR2 and mGluR8 were successfully coupled to phospholipase C activation using the chimeric G protein G{alpha}q/o. The pharmacological properties of recombinant receptors were characterized and proved to be similar to native receptors. Our data suggest that the ecdysone system has a number of characteristics that make it well suited for expressing mGluRs and that the combined use of this system and chimeric G proteins allows receptors to be characterized using a rapid and straightforward Ca2+ assay.

Key Words: metabotropic glutamate receptors, inducible expression, chimeric G proteins

First published on October 18, 2005, doi:10.1177/1087057105280285

Journal of Biomolecular Screening 2005;10:841.

A more recent version of this article appeared on December 1, 2005

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