Effects of Human Type 1α Metabotropic Glutamate Receptor Expression Level on Phosphoinositide and Ca2+ Signalling in an Inducible Cell Expression System

    loading  Checking for direct PDF access through Ovid

Abstract

Abstract:

Stable expression of the human type 1α metabotropic glutamate (mGlu1α) receptor was achieved in Chinese hamster ovary cells using an isopropyl-β-D-thiogalactoside (IPTG)-repressible expression system. Treatment of the cells with IPTG resulted in a time- and concentration-dependent induction of receptor expression. Maximal expression was obtained after treatment of the cells with 100 μM IPTG for 20 h, leading to a marked increase in receptor immunoreactivity detected by western blot, >30-fold stimulation of 3 H-labelled inositol phosphate (3H-InsP) production, and a robust increase in intracellular calcium concentration in single cells after stimulation with 20 μM quisqualate. The basal level of 3H-InsP accumulation in cells induced with IPTG was increased by two- to threefold as compared with control cells; however, this basal activity was found to be dependent on glutamate released by the cells into the incubation medium. Following IPTG treatment, stable expression of the mGlu1α receptor was maintained for at least 1 week. Taken together, these results clearly indicate the advantages of working with an inducible expression system when studying the biochemical and pharmacological properties of the human mGlu1α receptor in transfected cells.

Related Topics

    loading  Loading Related Articles