Characterization of ligand binding to Dopamine D3 receptor using fluorescence anisotropy and radioligand binding

Abstract

There are five subtypes of dopamine receptors that play a role in the dopaminergic system. Due to their limited distribution and involvement in cognitive and emotional functions, Dopamine D3 receptors are attractive pharmacological targets for treatment of drug addiction and neuropsychiatric disorders. D3 receptor ligands have been labelled with a fluorescent dye or a radioisotope for direct monitoring of ligand binding to the receptor. However, there are not many fluorescent ligands that are available for studying D3 receptor. A2-TAMRA is a novel fluorescent ligand with high affinity for D3 receptor. Binding of [3H]-methylspiperone to D3R was studied in parallel to validate results from A2-TAMRA binding to D3R. The D3 receptor ligands had similar affinities in inhibiting A2-TAMRA and [3H]-methyl spiperone binding to D3 receptor, since a very good correlation (R2 = 0.94) was obtained between both methods. The affinities for the known antagonists had a good correlation with previously published data. Selectivity of A2-TAMRA towards two different subtypes was also studied and we found that A2-TAMRA prefers D3 receptor over D1 receptor.