In vitro screening of psychoactive drugs by [35S]GTPγS binding in rat brain membranes

Abstract

We constructed a reproducible, simple, and small-scale determination method of the psychoactive drugs that acted directly on the monoamine receptor by measuring the activation of [35S]guanosine-5′-O-(3-thio)- triphosphate binding to guanine nucleotide-binding proteins (G proteins). This method can simultaneously measure the effects of three monoamines, namely dopamine (DA), serotonin (5-HT), and norepinephrine (NE), in rat brain membranes using a 96-well microplate. Activation of D1 and D2 receptors in striatal membranes by DA as well as 5-HT and NEα2 receptors in cortical membranes could be measured. Of 12 tested phenethylamines, 2,5-dimethoxy-4-chlorophenethylamine (2C-C), 2,5-dimethoxy-4- ethylphenethylamine (2C-E), and 2,5-dimethoxy-4-iodophenethylamine (2C-I) stimulated G protein binding. The other phenethylamines did not affect G protein binding. All 7 tryptamines tested stimulated G protein binding with the following rank order of potency; 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT)>5-methoxy-N,N-diallyltryptamine (5-MeO-DALT)>5-methoxy- α-methyltryptamine (5-MeO-AMT)≥5-methoxy-N,N-methylisopropyltryptamine (5-MeO-MIPT)>5-methoxy-N,N-diisopropyltryptamine (5-MeO-DIPT)>N,N- dipropyltryptamine (DPT)&gelα-methyltryptamine (AMT). This assay system was able to designate psychoactive drugs as prohibited substances in accordance with criteria set forth by the Tokyo Metropolitan government. © 2007 Pharmaceutical Society of Japan.