5-HYDROXYTRYPTAMINE 5-HT1D RECEPTORS MEDIATING INHIBITION OF CYCLIC-AMP ACCUMULATION IN MADIN-DARBY CANINE KIDNEY (MDCK) CELLS

5-Hydroxytryptamine 5-HT1B/5-HT1D receptors are members of the same receptor subfamily, but display a different pharmacology (Hartig et al. (1992) Trends Pharmacol Sci 13:152-159). Whereas several cell lines have been reported to contain 5-HT1B receptors, none has been described, however, that endogenously expresses well-characterized 5-HT1D receptors. The present study deals with the identification of 5-HT1D receptors inhibiting cyclic AMP accumulation in Madin-Darby canine kidney (MDCK) cells. 5-HT (1 nM- 10 mu M) induced a concentration-dependent inhibition of the cyclic AMP accumulation stimulated by prostaglandin E(1) (1 mu M) in MDCK cells. The maximal effect of 5-HT averaged 50% inhibition and was abolished after a pre-treatment of the cells with pertussis toxin. Other agonists mimicked the effects of 5-HT, with the following rank order of potency (pEC(50) +/- SEM, n greater than or equal to 3): 5-carboxamidotryptamine (8.36 +/- 0.48) > PAPP (p-aminophenylethyl-m-trifluoromethylphenyl piperazine, 7.89 +/- 0.23) > 5-HT (7.35 +/- 0.05) > sumatriptan (6.65 +/- 0.27). PAPP behaved as a partial agonist. 8-OH-DPAT (8-hydroxy-2-(di-n-propylamino)tetralin) was less potent, its maximal effect being not reached at 0.1 mM. Methiothepin, GR127935: (-) propranolol, rauwolscine and ketanserin were all devoid of intrinsic activity (up to 10 mu M or 0.1 mM). Methiothepin (10 nM, 0.1 mu M and 1 mu M) antagonized 5-HT effect (pA(2) 8.57 +/- 0.44, Schild slope 1.17 +/- 0.21, n = 3). GR127935 (1 nM, 10 nM and 0.1 mu M) shifted the curve of 5-HT to the right, but the antagonism was not fully surmountable (apparent pK(B) value, 9.80 +/- 0.16, n = 9). From the shifts obtained with rauwolscine (1 mu M) and (-) propranolol (10 mu M), respective pK(B) values were estimated 6.68 +/- 0.30 and approximate to 5.4 (n = 3 each). PAPP, when tested as an antagonist at 1 mu M, also shifted the curve of 5-HT to the right, with a pK(B) of 8.27 +/- 0.16 (n = 3). Finally, ketanserin (10 mu M) also antagonized the effects of 5-HT, the pK(B) being 6.54 +/- 0.16 (n = 9). The rank orders of agonist and antagonist potencies strongly suggest 5-HT receptors mediating inhibition of cyclic AMP accumulation in MDCK cells to be 5-HT1D receptors. This is the first report of a cell line expressing endogenous, well-characterized, 5-HT1D receptors. With regard to the 5-HT1D receptor subtype involved, the relatively high potency of ketanserin would suggest it to be a 5-HT1D alpha, subtype or a mixture of 5-HT1D chi/S-HT1D beta subtypes. However, caution must be exercised here, owing to the poor knowledge of canine 5-HT1D receptor subtypes.