Dopamine and its receptors are extremely widespread throughout the body, and are especially prevalent in the nervous system. Basic and clinical neuroscience literature has reported that the dopamine systems are important in the psychomotor disturbances associated with schizophrenia, Parkinson's disease, Huntington's disease, Tardive dyskinesia and cocaine addiction. Consequently, determining the cellular locations and potential interactions among dopamine receptors is an important endeavor in targeting appropriate drug therapies for these neurological disturbances. Two dopamine receptor subclasses can be pharmacologically described. The receptors are designated as either D1 or D2 by their cyclic AMP interactions and the type of ligand recognized by the binding site. D1 receptors stimulate adenylyl cyclase and bind substituted benzazepines; D2 receptors inhibit cyclic AMP formation and are inhibited by neuroleptic drugs. These two subtypes can be distinguished using autoradiography of their ligand binding, and studies show a heterogeneous distribution of the subtypes throughout the CNS. Different genes for these subtypes have been isolated recently and are expressed in high abundance in the nervous system. To date, five different dopamine receptors have been cloned in the rat, (i.e, D1A, D1B, D2, D3, and D4). Two of the receptors exhibit RNA splice variants. The hypothesis to be examined is that dopamine receptors are prevalently co-localized within identified neurons of the mesostriatal and mesocorticolimbic systems. We will document this relationship in the two major mesencephalic dopamine systems, since the behavioral manifestation of dopamine signalling in these two pathways is very different. The proposed experiments will examine the intact brain, and focus on the neostriatum (CN), the substantia nigra (SN), the ventral tegmental area (VTA), and the prefrontal cortex (Pfx), since these four regions contribute to the mesostriatal (SN, CN) and mesocorticolimbic (VTA, Pfx) systems. Cellular determination of dopamine receptor protein will be established using subtype selective antisera in order to distinguish the five cloned receptor isoforms and their variants in these two systems. A sensitive in situ transcription method will be employed to visualize nascent cDNA generated from specific dopamine receptor primers. The information gathered should provide a basis for establishing receptor co- incidence and potential synergies which may be involved in mediation of dopaminergic responses in the mesostriatal versus the mesocorticolimbic systems.
