Rapid internalization and surface expression of a functional, fluorescently tagged G-protein-coupled glutamate receptor

L-Glutamate is the principal excitatory neurotransmitter in the vertebrate central nervous system, where it mediates many of its actions via G-protein -coupled metabotropic glutamate (mGlu) receptors. Since little is known abo ut the dynamics of mGlu receptors at the plasma membrane, we have construct ed a fusion protein comprising the mGlu receptor subtype 1 alpha (mGlu(1 al pha)) and green fluorescent protein (GFP). Using imaging of Ca2+ release fr om intracellular stores as a functional assay, the agonist pharmacology of this fluorescently tagged receptor was found to be similar to that of the w ild-type receptor when expressed in HEK-293 cells. Receptor movement and fu nction were measured simultaneously by combined imaging of Ca2+, using fura -red, and GFP fluorescence in single cells. Exposure to agonist induced a r apid loss of up to 30% of membrane-associated fluorescence, with a correspo nding decrease in the functional response. Following removal of the agonist there was recovery of both the membrane fluorescence and the functional re sponse. These data suggest that the surface expression of G-protein-coupled glutamate receptors might be rapidly regulated in response to agonist acti vation.