Pindolol-insensitive 3H-5-hydroxytryptamine binding in the rat hypothalamus; identity with 5-hydroxytryptamine7 receptors

摘要

class="enumerated" style="list-style-type:decimal">Pindolol-insensitive [³H]-5-hydroxytryptamine ([³H]-5-HT) binding to rat hypothalamic membranes was pharmacologically and functionally characterized to resolve whether this procedure selectively labels 5-HT7 receptors.Consistent with a previous report, 3 μM and not 100 nM pindolol was required to occupy fully 5-HT1A and 5-HT1B receptors. Remaining [³H]-5-HT binding was saturable (KD, 1.59±0.21 nM; Bmax, 53.8±3.1 fmol.mg protein⁻¹).Displacement of [³H]-5-HT with metergoline and 5-CT revealed shallow Hill slopes (<0.5) but seven other compounds had slopes >0.8 and pKi values and the rank order of affinity were significantly correlated (r=0.81 and 0.93, respectively) with published [³H]-5-HT binding to rat recombinant 5-HT7 receptors.In the presence of pindolol, 5-HT-enhanced accumulation of [³²P]-cyclic AMP was unaffected by the 5-HT4 antagonist RS39604 (0.1 μM) or the 5-ht6 antagonist Ro 04-6790 (1 μM) but significantly attenuated by mesulergine (250 nM), ritanserin (450 nM) or methiothepin (200 nM) which have high affinity for the 5-HT7 receptor.Intracerebroventricular pretreatment with the serotonergic neurotoxin 5,7-dihydroxytryptamine, 5,7-DHT, elevated the [³H]-5-HT Bmax 2 fold, indicating that the hypothalamic 5-HT7 receptor is post-synaptic to 5-HT nerve terminals and regulated by synaptic 5-HT levels.These results suggest that, in the presence of 3 μM pindolol, [³H]-5-HT selectively labels hypothalamic binding sites consistent with functional 5-HT7 receptors.