Receptor subtype and density determine the coupling repertoire of the 5-HT2 receptor subfamily

Abstract

The 5-Hydroxytryptamine (5-HT)(2C) receptor (originally known as the 5-HT(1C) receptor) is a member of the 5-HT2 subfamily of G protein coupled receptors, which is known to couple to phospholipase C. Within the 5-HT2 subfamily, only the 5-HT(2C) receptor also coupled to inhibition of forskolin-stimulated cAMP production when expressed at high density (12 pmol/mg membrane protein) in stably transformed AV12 cells. The 5-HT(2C) receptor coupled with high efficacy to both phospholipase C as measured by IP3 (inositol 1,4,5-trisphosphate) production and to inhibition of forskolin-stimulated cAMP production (EC50 = 2.98 nM ± 0.9 and IC50 = 47.99 nM ± 10.25 respectively). The 5-HT(2A) and 5-HT(2B) receptors, while coupling to phospholipase C with high affinity EC50s of 19.24 nM ± 6.44 and 1.24 nM ± 0.136 respectively), did not decrease adenylyl cyclase activity. The 5-HT(2C) receptor actions in both systems showed the expected pharmacology for the 5-HT(2C) receptor, e.g., mesulergine antagonized the effects of 5-HT and spiperone did not. Preincubation of cells with PTX showed that the G protein coupling of the 5-HT(2C) receptor to phospholipase C is PTX insensitive, while the G protein coupling to inhibition of adenylyl cyclase is PTX sensitive, even to concentrations as low as 20 ng/ml of PTX. PTX pretreatment of the 5-HT(2C) bearing cells also unmasked a small stimulatory effect on adenylyl cyclase. When expressed at low density the 5-HT(2C) receptor potentiated forskolin-stimulated cAMP production by 2 fold while still maintaining its ability to enhance PI hydrolysis. A more modest potentiation of cAMP production was noted with low density expression of the 5-HT(2B) receptor. Thus the ability of the 5-HT(2C) receptor to interact with several effecters through at least two different G proteins is, in part, receptor subtype specific but also influenced by receptor density.