Characterization of a cloned rat serotonin 5-HT1A receptor expressed in the HeLa cell line.
We have previously isolated the rat serotonin (5-HT)1A receptor gene (G21Y2) and now report the expression and characterization of this receptor. The BamHI/Xbal fragment of this gene was cloned into Rc/RSV and stably transfected into HeLa cells by the calcium phosphate method. For determination of specific 5-HT1A receptor binding, [3H]8OH-DPAT was used as the radioligand and incubated with HeLa cell membranes. The cells expressed specific and saturable binding of [3H]8OH-DPAT with a Kd value of 0.3 nM and a Bmax value of 2 pmol/mg protein. GTP (50 microM) added to the incubation mixture increased the Kd value to 3 nM indicating that the expressed receptor is coupled to a G protein. The specific binding was inhibited by selective 5-HT1A partial agonists, such as buspirone, ipsapirone, gepirone, tandospirone, zalospirone and SUN8399 with Ki values of 1-30nM, whereas other neurotropic drugs except for spiperone (Ki = 46 nM) and nemonapride (Ki = 2.3 nM) were effective only at concentrations of more than 100 microM. The potencies of these compounds to inhibit [3H]8OH-DPAT from its specific binding sites were similar to their affinities determined in rat hippocampus binding studies. These data suggest that the expressed receptor is a 5-HT1A-type similar to 5-HT1A receptors in the rat hippocampus.
Fujiwara
Y
Y
Department of Neuropsychiatry, Okayama University Medical School, Japan.
Tomita
H
H
Hikiji
M
M
Kashihara
K
K
Otsuki
S
S
Ohnuki
T
T
Hamagishi
Y
Y
Oki
T
T
Sora
I
I
Roeske
W R
WR
D017371:8-Hydroxy-2-(di-n-propylamino)tetralin / Q000378:metabolism*; D000818:Animals; D014150:Antipsychotic Agents / Q000494:pharmacology; D001667:Binding, Competitive; D003001:Cloning, Molecular; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D051381:Rats; D011985:Receptors, Serotonin / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D017366:Serotonin Receptor Agonists / Q000378:metabolism / Q000494:pharmacology; D013134:Spiperone / Q000494:pharmacology; D014162:Transfection
D014150:Antipsychotic Agents; D011985:Receptors, Serotonin; D017366:Serotonin Receptor Agonists; D013134:Spiperone; D017371:8-Hydroxy-2-(di-n-propylamino)tetralin; D006160:Guanosine Triphosphate