RT Journal Article SR Electronic T1 Multiple Conformations of Native and Recombinant Human 5-Hydroxytryptamine2A Receptors Are Labeled by Agonists and Discriminated by Antagonists JF Molecular Pharmacology JO Mol Pharmacol FD American Society for Pharmacology and Experimental Therapeutics SP 690 OP 699 VO 60 IS 4 A1 Juan F. López-Giménez A1 Marı́a Villazón A1 José Brea A1 M. Isabel Loza A1 José M. Palacios A1 Guadalupe Mengod A1 M. Teresa Vilaró YR 2001 UL http://molpharm.aspetjournals.org/content/60/4/690.abstract AB We have expanded previous studies with the 5-hydroxytryptamine (5-HT)2 receptor agonist (±)-1-(2,5-dimethoxy-4-[125I]iodophenyl)-2-aminopropane [(±)-[125I]DOI] in human brain that had shown biphasic competition curves for several 5-HT2A receptor antagonists by using new selective antagonists of 5-HT2A (MDL100,907) and 5-HT2C (SB242084) receptors together with ketanserin and mesulergine. Autoradiographic competition experiments were performed with these antagonists in human brain regions where (±)-[125I]DOI labels almost exclusively 5-HT2A receptors (frontal cortex and striosomes). Furthermore, the effect of uncoupling receptor/G protein complexes on antagonist competition was studied with guanosine-5′-(β,γ-imido)triphosphate [Gpp(NH)p]. Competition experiments with (±)-[3H]1-(4-bromo-2,5-dimethoxyphenil)-2-aminopropane [(±)-[3H]DOB] were also performed in membranes from Chinese hamster ovary cells (CHOFA4) expressing cloned human 5-HT2A receptors. In both systems, ketanserin and MDL100,907 displayed biphasic competition profiles, whereas SB242084 and mesulergine competed monophasically. In absence of antagonist, 100 μM Gpp(NH)p decreased brain (±)-[125I]DOI specific binding by 40 to 50% and (±)-[3H]DOB specific binding to CHOFA4 cells by 30%. The remaining agonist-labeled uncoupled sites were still displaced biphasically by ketanserin and MDL100,907, with unaltered affinities. Saturation experiments were performed in CHOFA4 cells. (±)-[3H]DOB labeled two sites (Kdh= 0.8 nM,Kdl = 31.22 nM). Addition of 100 μM Gpp(NH)p resulted in a single low-affinity (Kd = 24.44 nM) site with unchanged Bmax. [3H]5-HT showed no specific binding to 5-HT2A receptors. These results conform with the extended ternary complex model of receptor action that postulates the existence of partly activated receptor conformation(s) (R∗) in equilibrium with the ground (R) and the activated G protein-coupled (R∗G) conformations. Thus, both in human brain and CHOFA4 cells, the agonists possibly label all three conformations and ketanserin and MDL100,907 recognize with different affinities at least two of these conformations. The American Society for Pharmacology and Experimental Therapeutics