This laboratory has been actively pursuing the design and development of drugs targeted to monoamine transporters for over a decade. The novel drugs have been indispensable as imaging agents for brain dopamine systems in Parkinson's disease and in cocaine abuse research, as candidate medications for cocaine addiction and Parkinson's disease, and as probes to develop a model of transporter function. We have discovered that a novel class of tropanes that have oxygen or carbon substitutions for nitrogen (nonamines) bind to monoamine transporters with high affinity and are as effective as their amine counterparts at inhibiting monoamine transport in vitro. These data demonstrate that neither ionic nor hydrogen bonding between an amine nitrogen on the ligand and a transporter amine counterion is needed for high affinity binding and blockade of transport. This discovery offers a compelling reason to investigate how drugs bind to and block monoamine transporters. Pilot experiments are designed to determine the critical domains on monoamine transporter molecules that mediate ligand binding and transport using site-directed mutagenesis and chimeric transporters. Point mutations will be made using the GeneEditor Site Directed Mutagenesis System Expression of mutant transporter protein will be confirmed using immunocytochemistry and/or western blotting. We will use a range drugs to determine the effects of mutations on uptake of [3H]