Showing papers by "F. Ivy Carroll published in 1997"

N-substituted phenyltropanes as in vivo binding ligands for rapid imaging studies of the dopamine transporter.

Abstract: Variously substituted phenyltropanes are proven as superb binding ligands for the dopamine transporter (DAT). In this study, we examine four N-substituted phenyltropanes which are derivatives of RTI-55 as in vivo binding ligands in mice. In this series, the methyl group on the nitrogen was replaced by a propyl (RTI-310), an allyl (RTI-311), a butyl (RTI-312), or a fluoropropyl (RTI-313) group. The in vitro binding potencies of these compounds at rat striatal DAT varied somewhat but were about 1 nM. While these compounds did not display marked selectivity for the dopamine transporter, they were more selective than RTI-55. Injection of the radiolabeled compound into mice resulted in striatal-to-cerebellar ratios that varied from about 4.5-6.5. The ratios peaked most rapidly for RTI-311 and RTI-313, at about 20 min. Pharmacological inhibition studies indicated that these compounds were binding to DATs in the striatum, as expected. These findings suggest that some compounds of this type may be excellent in vivo binding ligands for rapid imaging studies of the DAT.

Dopamine-Transporter Uptake Blockers

Abstract: The dopamine transporter (DAT), a protein located on presynaptic nerve terminals (1-3),plays a major role in the reuptake of released dopamine. Uptake of DA is sodium-and chloride-ion-as well as temperature-and time-dependent, and is inhibited by a variety of compounds, including cocaine. Even though cocaine binds to several sites in the brain, cocaine abuse has been shown to be related to binding at the DAT site. Thus, analysis of the binding potencies of a series of cocaine analogs demonstrates that, in animal models, the reinforcing properties of these analogs correlate only with binding potencies at the DAT site. The DAT site has been called a cocaine receptor (4,5); it may be the initial site responsible for producing cocaine’s drug reinforcement. The cDNA for the DAT has been cloned from rat (6–8),mouse (9,10),bovine (11),and human (12) brains, and these clones exhibit 92, 93, and 88% homology to human DAT (hDAT), respectively. A transporter with 92% homology and with neuronal DAT properties has also been characterized from African Green Monkey kidney (COS-7) cell lines (13). The hydrophobicity profile of the DAT indicates 12 possible membrane-spanning regions with the amino and carboxy termini located intracellularly. The protein from human and rat brains contains three and four extracellular N-glycosylation sites, respectively (Fig. 1). A detailed review of structural information about the DAT has recently been published (14).

Turnover of Rat Dopamine Transporter Protein in rDAT-LLC-PK1 Cells

Abstract: We have determined the turnover of the dopamine transporter protein in a porcine kidney epithelial cell line, LLC-PK1, that stably expresses a rat dopamine transporter (rDAT-LLC-PK1). A cocaine analog, RTI-76 (3s-(p-chlorophenyl)tropan-2s-carboxylic acid-p-isothiocyanatophenylethyl ester), was used to block dopamine transporter binding irreversibly in intact cells, and the rate of reappearance of transporter binding was measured. RTI-76 inhibited [125I]RTI-55 binding to the dopamine transporter rapidly (within 10 min.) with an IC50 of 23.8 nM. After exposure to RTI-76, dopamine transporter binding reappeared in the cells over time with monoexponential kinetics with a half-life of 23 ± 1.6 h. Cell lines stably expressing DAT and irreversible binding ligands will be useful to study perturbations of transporter activity and their consequent effects on the transporter protein half-life.

Imaging Transporters for Dopamine and Other Neurotransmitters in Brain

Abstract: There has been substantial progress in the imaging of dopamine transporters. The earliest positron emission tomography (PET) experiments used carbon-11 labeled nominfensine (1,44); whereas recent investigations by both PET and single photon emission computed tomography (SPECT) utilize a variety of ligands with a variety of properties and address many biological questions (see below). The earliest ligands utilized, [11C]nomifensine and [11C]cocaine, were somewhat limited because of low signal-to-noise ratios. Nevertheless, [11C]cocaine as a ligand continues to be popular, because it is the same authentic substance that is abused around the world. The best, current 3-substituted phenyltropane ligands, such as RTI-55, have vastly improved signal-to-noise ratios. These successes are in no small part because of the development of the chemistry of cocaine analogs and other analogs of inhibitors for transporters (Fig. 1) (4,8). Despite this progress, several important things remain to be achieved. We need to develop methods to carry out rigorous calculations of Bmax instead of relying on more inexact but easier measures of relative occupancy and binding potential. We also need useful ligands for all transporters in brain.