EFFECTS OF HUMAN TYPE 1-ALPHA METABOTROPIC GLUTAMATE-RECEPTOR EXPRESSION LEVEL ON PHOSPHOINOSITIDE AND CA2+ SIGNALING IN AN INDUCIBLE CELL EXPRESSION SYSTEM

Stable expression of the human type 1 alpha metabotropic glutamate (mG lu1 alpha) receptor was achieved in Chinese hamster ovary cells using an isopropyl-beta-D-thiogalactoside (IPTG)-repressible expression syst em. Treatment of the cells with IPTG resulted in a time-and concentrat ion-dependent induction of receptor expression. Maximal expression was obtained after treatment of the cells with 100 mu M IPTG for 20 h, le ading to a marked increase in receptor immunoreactivity detected by we stern blot, >30-fold stimulation of H-3-labelled inositol phosphate (H -3-InsP) production, and a robust increase in intracellular calcium co ncentration in single cells after stimulation with 20 mu M quisqualate . The basal level of H-3-InsP accumulation in cells induced with IPTG was increased by two-to threefold as compared with control cells; howe ver, this basal activity was found to be dependent on glutamate releas ed by the cells into the incubation medium. Following IPTG treatment, stable expression of the mGlu1 alpha receptor was maintained for at le ast 1 week. Taken together, these results clearly indicate the advanta ges of working with an inducible expression system when studying the b iochemical and pharmacological properties of the human mGlu1 alpha rec eptor in transfected cells.