Identification and Role of Serotonin 5‐HT_1A and 5‐HT_1B Receptors in Primary Cultures of Rat Embryonic Rostral Raphe Nucleus Neurons

Abstract

Abstract: Autoregulatory mechanisms affecting serotonin [5‐hydroxytryptamine (5‐HT)] release and synthesis during the early period of development were investigated in dissociated cell cultures raised from embryonic rostral rat rhombencephalon. The presence of 5‐HT 1A and 5‐HT 1B receptors in serotoninergic neurons was assessed using binding assays. The involvement of 5‐HT 1A and 5‐HT 1B receptors in the control of the synthesis and release of [ 3 H]5‐HT was studied using biochemical approaches with several serotoninergic receptor ligands. A mean decrease of 30% in [ 3 H]5‐HT synthesis and release was observed in the presence of 5‐HT (10 −8 M ), the 5‐HT 1A agonist 8‐hydroxy‐2‐(di‐ n ‐propylamino)tetralin (8‐OH‐DPAT), the 5HT 1B/1A agonist 5‐methoxy‐3‐(1,2,5,6‐tetrahydro‐4‐pyridinyl)‐1 H ‐indole (RU 24969), the 5‐HT 1B agonist 3‐(1,2,5,6‐tetrahydropyrid‐4‐yl)pyrrolo[3,2‐ b ]pyrid‐5‐one (CP‐93, 129), and the 5‐HT 1D/1B agonist sumatriptan. Inhibition of 5‐HT synthesis and release induced by 8‐OH‐DPAT was blocked by chiral N ‐tert‐butyl‐3‐[1‐[1‐(2‐methoxy)phenyl]piperazinyl]‐1‐phenylpropionamide dihydrochloride quaternary‐hydrate (WAY 100135) (10 −7 M ) or methyl 4‐[4‐[4‐(1,1,3‐trioxo‐2 H ‐1,2‐benzoisothiazol‐2‐yl)butyl]‐1‐piperazinyl]‐1 H ‐indole‐2‐carboxylate (SDZ 216–525) (10 −7 M ), and that of CP‐93, 129 was blocked by methiothepin (10 −7 M ). Paradoxically, extracellular levels of [ 3 H]5‐HT increased in the presence of 8‐OH‐DPAT and RU 24969 at 10 −6 M . 5‐HT uptake experiments showed that these two agonists interacted with the 5‐HT transporter. 5‐HT 1 binding sites (620 fmol/mg of protein) and 5‐HT 1A (482 fmol/mg of protein) and 5‐HT 1B (127 fmol/mg of protein) receptors were detected in 12‐day in vitro cell cultures. Experiments carried out with tetrodotoxin suggested that 5‐HT 1A receptors are located on nerve cell bodies, whereas 5‐HT 1B receptors are located on the nerve terminals. We concluded that autoregulatory mechanisms involving 5‐HT 1A and 5‐HT 1B autoreceptors are functionally mature in cells from rostral raphe nuclei during the early period of development.