S 14506: novel receptor coupling at 5-HT1A receptors

Abstract S 14506 is chemically related to the inverse agonist at 5-HT 1A receptors, spiperone, but S 14506 behaves as one of the most potent agonists known at these receptors, both in vitro and in vivo. In hippocampal membranes, the specific binding of [  3 H]-S 14506 ( K d =0.79±0.2 nM; B max =400±32 fmol/mg protein) to 5-HT 1A receptors resembled that of an antagonist in that it was increased by GppNHp, whereas GppNHp reduced the binding of the classic agonist [  3 H]-8-OH-DPAT ( K d =1.5±0.5 nM; B max =303±20 fmol/mg protein). Manganese, magnesium and calcium reduced the binding of [  3 H]-S 14506 to 5-HT 1A receptors whereas the binding of [  3 H]-8-OH-DPAT was increased. Further, sodium markedly reduced the binding of [  3 H]-8-OH-DPAT, without affecting the binding of [  3 H]-S 14506. [  3 H]-S 14506 also bound with high affinity to h 5-HT 1A receptors stably expressed in membranes of CHO cells ( K d =0.13±0.05 nM; B max =2.99±0.60 pmol/mg protein): the B max was double that of [  3 H]-8-OH-DPAT. GppNHp strongly decreased [  3 H]-8-OH-DPAT binding but scarcely changed [  3 H]-S 14506 binding; calcium, magnesium and manganese had little effect on [  3 H]-S 14506 binding in CHO cells. Antagonists (WAY 100635, WAY 100135) and inverse agonists (spiperone and metitepine) displaced [  3 H]-S 14506 binding with high affinity and Hill slopes close to unity, whereas agonists (5-HT and 5-CT) displayed low affinity with low Hill slopes: partial agonists (buspirone, ipsapirone) showed intermediate properties. In fusion proteins of h 5-HT 1A receptors with G iα1 the compound potently increased high-affinity GTPase, with a steeper Hill slope than for 5-HT, which may indicate positive cooperativity. The maximum response for S 14506 in these assays was equivalent to 5-HT, indicating it to be a full agonist. In molecular modelling studies, using a three-site model of the 5-HT 1A receptor, S 14506 spanned between the 5-HT recognition site and the “arginine switch” (DRY microdomain) postulated to activate the interaction of the receptor with the G protein. Thus it is possible to synthesise ligands at G-protein-coupled receptors which are highly potent agonists, but which are structurally related to inverse agonists and show some features of antagonist/inverse agonist binding.