IMMORTALIZED HYPOTHALAMIC NEURONS EXPRESS METABOTROPIC GLUTAMATE RECEPTORS POSITIVELY COUPLED TO CYCLIC-AMP FORMATION

We have characterized the expression pattern and pharmacological profi le of activation of metabotropic glutamate receptors (mGluRs) in immor talized, gonadotropin releasing hormone (GnRH)-secreting GT1-7 cells, which represent a homogeneous cellular population of hypothalamic orig in. These cells are known to respond to the mGluR agonist (1S,3R)-cycl opentanedicarboxylic acid (1S,3R-ACPD) with increased GnRH release. To establish which specific mGluR subtypes are expressed by GT1-7 cells, we used polyclonal antibodies raised against non-conserved regions of the carboxy-terminal domains of individual subtypes. The selectivity of these antibodies was tested in HEK 293 cells transiently transfecte d with each mGluR subtype. GT1-7 cells stained positively for the subt ypes mGluR1a, -1b and -5 (belonging to group I mGluRs), mGluR2/3 (grou p II) and mGluR7 (group III). Agonists of group I mGluRs, including 1S ,3R-ACPD, activated phosphoinositide hydrolysis in GT1-7 cells. This e ffect, however, was manifested only when cell density was low, and it disappeared when cells reached confluence. Stimulation of phosphoinosi tide hydrolysis could not therefore have been related to hormone secre tion because 1S,3R-ACPD effectively released GnRH in confluent culture s. We then focused on group II and ill mGluRs, which in transfected ce lls are negatively linked to adenylate cyclase activity. Unexpectedly, however, agonists which preferentially activate group II and III mGlu Rs increased both basal and forskolin-stimulated cAMP accumulation in GT1-7 cells. Stimulation of cAMP accumulation by mGluR agonists was no t prevented by enzymatic depletion of endogenous adenosine, but was ob literated when cells were incubated with agonists of receptors positiv ely coupled to adenylate cyclase, such as beta-adrenergic and prostagl andin E(2) receptors. These results suggest that GT1-7 cells express a novel mGluR subtype positively coupled to adenylate cyclase, which sh ares the same transduction pathway of other classical receptors couple d with a G(s)-type of GTP-binding protein.