Methyl 3β‐(4‐[¹²⁵I]Iodophenyl)Tropane‐2β‐Carboxylate In Vitro Binding to Dopamine and Serotonin Transporters Under “Physiological” Conditions

Abstract: Methyl 3β‐(4‐[¹²⁵I]iodophenyl)tropane‐2β‐carboxylate ([¹²³I]β‐CIT) is a single photon emission computed tomographic radiotracer for in vivo labeling of dopamine (DA) and serotonin (5‐HT) transporters. Single photon emission computed tomographic experiments in nonhuman primates showed that [¹²³I]β‐CIT in vivo binding to DA transporters had a much slower washout than binding to 5‐HT transporters. This observation was not predicted from previously published in vitro studies. These studies, performed at 22°C in nonphysiological buffer, reported similar affinity of [¹²⁵I]β‐CIT for DA and 5‐HT transporters. We now report [¹²⁵I]β‐CIT binding parameters to fresh rat membranes at 22°C and 37°C, in a buffer mimicking the composition of cerebrospinal fluid. At both temperatures, binding to DA transporters was best fit by a twosite model, whereas binding to 5‐HT transporters was compatible with one population of sites. At 22°C, [¹²⁵I]β‐CIT showed similar affinity to high‐affinity DA (0.39 nM) and 5‐HT transporter sites (0.47 nM). Increasing the incubation temperature from 22°C to 37°C reduced binding to DA transporters by 60%, whereas binding to 5‐HT transporters was only marginally affected. In vitro kinetic experiments failed to detect significant differences in on or off rates that could explain the observed in vivo kinetics. These experiments thus failed to explain [¹²³ I]β‐CIT in vivo uptake kinetics, suggesting the existence of specific factors affecting the in vivo situation.