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.
Dickenson
J M
JM
Institute of Cell Signalling, School of Biomedical Sciences, Queen's Medical Centre, Nottingham, UK.
Hill
S J
SJ
D000818:Animals; D016466:CHO Cells; D005576:Colforsin / Q000494:pharmacology; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism*; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D007295:Inositol Phosphates / Q000378:metabolism*; D009119:Muscle Contraction; D009131:Muscle, Smooth, Vascular / Q000502:physiology; D044302:Receptor, Serotonin, 5-HT1B; D011985:Receptors, Serotonin / Q000187:drug effects* / Q000378:metabolism; D015290:Second Messenger Systems; D017366:Serotonin Receptor Agonists / Q000494:pharmacology*; D018170:Sumatriptan / Q000494:pharmacology*
D018076:DNA, Complementary; C470302:HTR1B protein, human; D007295:Inositol Phosphates; D044302:Receptor, Serotonin, 5-HT1B; D011985:Receptors, Serotonin; D017366:Serotonin Receptor Agonists; D005576:Colforsin; D018170:Sumatriptan; D000242:Cyclic AMP; D019204:GTP-Binding Proteins