This is a request for an Independent Scientist Award. The long term objective of this research is the understanding of the biological significance of the neurotransmitter dopamine (DA) in several structures of the central nervous system with respect to its role in the action of drugs of abuse. Two areas of investigation are planned. One area is the examination of the structural features of substrates and transporter affecting kinetics of the human dopamine and norepinephrine transporters, hDAT and hNET, respectively. A great deal of work has gone into the study of inhibitors of hDAT and hNET, but features of substrates have been less studied. In particular, hypotheses concerning the interaction of the amine group and the catechol groups with specific amino acids in the transporter will be tested. A recently developed method, rotating disk electrode voltammetry, will be used to observe the time course of uptake and induced efflux. The results of these studies will be put in the context of kinetic studies on a range of substrates. A second area, related to the above studies, is the localization of the binding site of substrates for hDAT and hNET. Previous work by others has identified regions of inhibitor binding using photoaffinity labels. Part of the proposed work is a collaboration to use recent progress in mass spectrometry to identify the specific amino acids covalently linked to the photoaffinity agents. This work will then be extended by using a newly synthesized photoaffinity substrate to localize substrate binding.

Agency
National Institute of Health (NIH)
Institute
National Institute on Drug Abuse (NIDA)
Type
Research Scientist Development Award - Research (K02)
Project #
5K02DA000179-10
Application #
6378242
Study Section
Human Development Research Subcommittee (NIDA)
Program Officer
Hillery, Paul
Project Start
1992-04-01
Project End
2002-08-31
Budget Start
2001-09-01
Budget End
2002-08-31
Support Year
10
Fiscal Year
2001
Total Cost
$119,361
Indirect Cost
Name
Emory University
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
066469933
City
Atlanta
State
GA
Country
United States
Zip Code
30322
Vaughan, Roxanne A; Parnas, M Laura; Gaffaney, Jon D et al. (2005) Affinity labeling the dopamine transporter ligand binding site. J Neurosci Methods 143:33-40
Reed, Brian; Chen, Nianhang; Justice Jr, Joseph B (2003) Dual-electrode voltammetry of catecholamine transport: simultaneous monitoring of uptake and efflux. J Neurosci Methods 126:127-35
Schad, Christina A; Justice Jr, Joseph B; Holtzman, Stephen G (2002) Endogenous opioids in dopaminergic cell body regions modulate amphetamine-induced increases in extracellular dopamine levels in the terminal regions. J Pharmacol Exp Ther 300:932-8
Chen, N; Justice, J B (2000) Differential effect of structural modification of human dopamine transporter on the inward and outward transport of dopamine. Brain Res Mol Brain Res 75:208-15
Gong, W; Neill, D B; Lynn, M et al. (1999) Dopamine D1/D2 agonists injected into nucleus accumbens and ventral pallidum differentially affect locomotor activity depending on site. Neuroscience 93:1349-58
Chen, N; Trowbridge, C G; Justice Jr, J B (1999) Cationic modulation of human dopamine transporter: dopamine uptake and inhibition of uptake. J Pharmacol Exp Ther 290:940-9
Danek Burgess, K S; Justice Jr, J B (1999) Effects of serine mutations in transmembrane domain 7 of the human norepinephrine transporter on substrate binding and transport. J Neurochem 73:656-64
Danek, K; Justice Jr, J B (1998) Voltammetric studies on kinetics of uptake and efflux at catecholamine transporters. Methods Enzymol 296:649-60
Chen, N; Trowbridge, C G; Justice Jr, J B (1998) Voltammetric studies on mechanisms of dopamine efflux in the presence of substrates and cocaine from cells expressing human norepinephrine transporter. J Neurochem 71:653-65
Chen, N; Justice Jr, J B (1998) Cocaine acts as an apparent competitive inhibitor at the outward-facing conformation of the human norepinephrine transporter: kinetic analysis of inward and outward transport. J Neurosci 18:10257-68

Showing the most recent 10 out of 23 publications