Functional coupling of human metabotropic glutamate receptor hmGlu1d: comparison to splice variants hmGlu1a and -1b

Functional coupling of the human mGlu1 splice variants was examined by heterologous expression. In cells stably (CHO) or transiently (A9) expressing the hmGlu1d receptor. agonists elevated intracellular calcium with a rank order of potency typical of a group I mGlu receptor (quisqualate > L-glutamate > (S)-dihydroxyphenylglycine > (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-ACPD)). These responses were reduced by the antagonist (+)-alpha-methyl-4-carboxyphenylglycine (MCPG), by pretreatment with pertussis toxin and phorbol ester, and by removal of extracellular calcium. In transiently transfected HEK293 cells, the hmGlu1b and -1d receptors increased inositol monophosphate (IP) production only in the presence of glutamate, whereas hmGlu1a coupled even in the absence of agonist. This was not due to differences in receptor expression levels as assessed by immunoblotting. Adenylate cyclase activity in HEK293 cells expressing the hmGlu1 variants was neither stimulated nor inhibited by glutamate. In A9 cells hmGlu1a-mediated calcium/fluo-3 fluorescence was sensitive to depletion of intracellular calcium stores by thapsigargin, but the hmGlu1d response was resistant. Thus, hmGlu1d receptors can be distinguished from hmGlu1a by their lack of agonist-independent coupling and their dependence on extracellular calcium.