DIFFERENTIAL INTRACELLULAR SIGNALING OF THE GALR1 AND GALR2 GALANIN RECEPTOR SUBTYPES

The diverse physiological functions exerted by the neuropeptide galani n may be regulated by multiple G protein-coupled receptor subtypes and intracellular signaling pathways. Three galanin receptor subtypes (Ga lRs) have been recently cloned, but the G protein coupling profiles of these receptors are not completely understood. We have generated GalR 1- and GalR2-expressing Chinese hamster ovary (CHO) cell lines and sys tematically examined the potential for these two receptors to couple t o the Gs, Gi, Go, and Gq proteins. Galanin did not stimulate an increa se in cAMP levels in GalR1/CHO or GalR2/CHO cells, suggesting an inabi lity of either receptor to couple to Gs. Galanin inhibited forskolin-s timulated cAMP production in GalR1/CHO cells by 70% and in GalR2/CHO c ells by 30%, suggesting a strong coupling of GalR1 to Gi and a more mo dest coupling between GalR2 and Gi, GalR1 and GalR2 both mediated pert ussis toxin-sensitive MAPK activity (2-3-fold). The stimulation mediat ed by GalR1 was inhibited by expression of the C-terminus of beta-adre nergic receptor kinase (beta ARKct), which specifically inhibits G bet a gamma signaling, but was not affected by the protein kinase C (PKC) inhibitor, bis[indolylmaleimide], or cellular depletion of PKC. In con trast, GalR2-mediated MAPK activation was not affected by beta ARKct e xpression but was abolished by inhibition of PKC activity. The data de monstrate that GalR1 is coupled to a Gi beta gamma signaling pathway t o mediate MAPK activation. In contrast, GalR2 utilizes a distinct sign aling pathway to mediate MAPK activation, which is consistent with Go- mediated MAPK activation in CHO cells. Galanin was unable to stimulate inositol phosphate (IP) accumulation in CHO or COS-7 cells expressing GalR1. In contrast, galanin stimulated a 7-fold increase in IP produc tion in CHO or COS-7 cells expressing GalR2. The GalR2-mediated LP pro duction was not affected by pertussis toxin, suggesting a linkage of G alR2 with Gq/G11. Thus, the GalR1 receptor appears to activate only th e Gi pathway. By contrast, GalR2 is capable of stimulating signaling w hich is consistent with activation of Go, Gq/G11, and Gi. The differen tial signaling profiles and the tissue distribution patterns of GalR1 and GalR2 may underlie the functional spectra of galanin action mediat ed by these galanin receptors and regulate the diverse physiological f unctions of galanin.