Human 5-HT1B receptor stimulated inositol phospholipid hydrolysis in CHO cells: synergy with Gq-coupled receptors.

We have previously reported that the transfected Gi/Go protein-coupled human adenosine A1 receptor (expressed at 200 fmol/mg of protein) and the endogenous 5-HT1B receptor (not detectable using radioligand binding) suppress forskolin-stimulated cyclic AMP accumulation and stimulate increases in [Ca2+]i in Chinese hamster ovary cells (CHO). In addition, co-activation of the adenosine A1 receptor (but not the 5-HT1B receptor) potentiates the hydrolysis of inositol phospholipids elicited by receptors coupled to Gq-proteins (Dickenson and Hill, 1996. Eur. J. Pharmacol. 320, 141-151). In order to establish whether this difference in ability to modulate Gq-coupled receptor responses is a consequence of low 5-HT1B receptor density, we have stably transfected CHO-KI cells with the human 5-HT1Dbeta cDNA (the human homologue of the rodent 5-HT1B receptor). We initially isolated a clonal cell line (designated CHO5-HT1B cells) displaying moderate specific [3H]5-HT binding (pKd of 8.17+/-0.07 and a Bmax of 140 fmol/mg protein). In CHO5-HT1B cells, the selective human 5-HT1B/1D receptor agonist sumatriptan produced a concentration-dependent inhibition of forskolin-stimulated cyclic AMP accumulation (pEC50=7.92+/-0.04). Sumatriptan also elicited a moderate and pertussis toxin-sensitive increase in [3H]inositol phosphate formation in CHO-5HT1B cells (pEC50=6.51+/-0.05). Finally, sumatriptan synergistically enhanced P2U purinoceptor stimulated [3H]inositol phosphate accumulation through a pertussis toxin-sensitive mechanism. These findings clearly show the significance of 5-HT1B receptor expression level in determining whether 5-HT1B receptor activation can modulate the accumulation of [3H]inositol phosphates elicited by a Gq-protein coupled receptor. The observation that 5-HT1B receptor activation can potentiate Gq-coupled receptor stimulated second messenger responses may have an important physiological role in the regulation of vascular smooth muscle contraction.