Multiple Conformations of Native and Recombinant Human 5-Hydroxytryptamine2A Receptors Are Labeled by Agonists and Discriminated by Antagonists Juan F. Lopez-Gimenez, Maria Villason, Hose Brea, M. Isabel Loza, Jose M. Palacios, Guadalupe Mengod, and M. Teresa Vilaro
Mol Pharmacol 60:690-699, 2001
(
http://molpharm.aspetjournals.org/cgi/reprint/60/4/690.pdf)
AbstractWe 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-HT
2A receptor antagonistsby using new selective antagonists of 5-HT
2A (MDL100,907) and 5-HT
2C (SB242084) receptors together with ketanserin and mesulergine. Autoradiographic competition experiments were performed with these antagonists in human brain regions where (+/-)-[
125I]DOI labels almowst exclusively 5-HT
2A receptors (frontal cortex and strioisomers). Furthermore, the effect of uncoupling receptor/G protein complexes on antagonist competition was studied with guanosine-5'-(?,?-imido)triphosphate [Gpp(NH)p]. Competition experients with (+/-)-[
3H]1-(4-bromo-2,5-dimethoxyphenyl)-2-aminopropane [(+/-)-[-
3H]DOB] were also performed on membranes from Chinese hamster ovary cells (CHOFA4) expressing cloned human 5-HT
2A receptors. In both systems, ketanserin and MDL100,907 displayed bihpasic competition profiles, whereas SB242084 and mesulergine competed monophasically. In absence of antagonist, 100 ?M Gpp(NH)p decreased brain (+/-)-[
125I]DOI specific binding to 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 (K
dh=0.8 nM, K
dl=31.22 nM). Addition of 100 ?M Gpp(NH)p resulted in a single low-affinity (K
d=24.44 nM) site with unchanged B
max. [
3H]5-HT showed no specific binding to 5-HT
2A receptors. These results conform with the extended ternary complex model of receptor action that postulates the existance 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.
It`s getting more and more complicated. I almost long for the 50s when there`s been almost nothing known about the human brain.