LOCALIZATION OF DOPAMINE CARRIERS BY BTCP, A DOPAMINE UPTAKE INHIBITOR, ON NIGRAL CELLS CULTURED INVITRO

BTCP, N-[1-(2-benzo(b)thiopenyl)cyclohexyl]piperidine, a derivative of phencyclidine, acted as a potent dopamine (DA) uptake blocking agent on primary cultures of dopaminergic neurons obtained from substantia nigra (IC50 = 70 nM). This value was closely related to IC50 determined for reference DA uptake inhibitors such as nomifensine (70 nM) or benztropine (50 nM), showing the specificity of BTCP towards the DA carrier. Thus, we used BTCP as a tool to visualize the DA uptake complexes on cultures, a model which preserves the integrity of the neurons. The [H-3]BTCP binding sites directly visualized by radioautographical (RAG) labelling seemed to follow the fibres (axons or dendrites) of neurons in culture whereas the cell bodies were not labelled. The [H-3]DA uptake visualized by RAG labelling, was inhibited either partially by BTCP at a concentration near its IC50 or totally by a high concentration of BTCP, all over the dopaminergic neurons (neurites and somas) immunostained with an anti-DA antiserum. Thus, the distribution of DA carriers can be investigated by a suitable tool, BTCP, a powerful and selective DA uptake blocker. These carriers have been visualized by radioautography with tritiated BTCP along the neurites, and the uptake can be totally blocked by a high concentration of BTCP all over DA neurons in vitro.