Characteristics of drug interactions with recombinant biogenic amine transporters expressed in the same cell type

We characterized the effects of drugs on the uptake of [H-3]neurotransmitte r by and the binding of [I-125](3 beta-(4-iodophenyl)tropane-2 beta-carboxy lic acid methyl ester ([I-125]RTI-55) to the recombinant human dopamine (hD AT), serotonin (hSERT), or norepinephrine (hNET) transporters stably expres sed in human embryonic kidney 293 cells. RTI-55 had similar affinity for th e hDAT and hSERT and lower affinity for hNET (K-d = 1.83, 0.98, and 12.1 nM , respectively). Kinetic analysis of [I-125]RTI-55 binding indicated that t he dissociation rate (k(-1)) was significantly lower for hSERT and the asso ciation rate (k(+1)) was significantly lower for hNET compared with the hDA T. The potency of drugs at blocking [H-3]neurotransmitter uptake was highly correlated with potency at blocking radioligand binding for hDAT and hSERT . Substrates were more potent at the inhibition of [H-3]neurotransmitter up take than radioligand binding. The potency of drugs was highly correlated b etween displacement of [H-3]nisoxetine (K-d = 6.0 nM) and [I-125]RTI-55 fro m the hNET, suggesting that these radioligands recognize similar sites on t he transporter protein. The correlation observed between inhibitory potency for uptake and binding of either ligand at the hNET was lower than correla tions between uptake and binding for hDAT and hSERT. The present results in dicate that the cocaine analog [I-125]RT1-55 has unique binding properties at each of the transporters and that the use of recombinant transporters ex pressed by a single cell type can provide a powerful screening tool for dru gs interacting with biogenic amine transporters, such as possible cocaine a ntagonists.