The dopamine transporter/cocaine receptor (DAT) is the site at which cocaine exerts rewarding/reinforcing effects. Cloning of rat and human transporter CDNAS provided primary sequence information and CDNA clones that facilitated production of antibodies that allow detection of the transporter protein immunoreactivity and MRNA expression in tissue extracts and in tissue sections, refining our information about the cellular distribution of transporter expression. Conjugated peptide preparations and biosynthesized fusion protein fragments elicited antibody responses in immunized rabbits; one of these sera recognized the transporter in immunohistochemical analyses. In situ hybridization studies continued to suggest high levels of transporter MRNA expression in neurons of the substantia nigra pars compacta and somewhat lower levels of expression in neurons of the ventral tegmental area in both rat and human. Retinal and arcuate nucleus neurons expressed even lower levels of MRNA in rats. Immunohistochemical studies revealed dense transporter immunoreactivity in fine, beaded neural processes found densely in the striatum and nucleus accumbens, but significantly less cell body staining than that found in adjacent control sections stained for tyrosine hydroxylase. Study of both coexpressed MRNAS revealed less tyrosine hydroxylase MRNA expression that transporter MRNA expression, despite good coregistration of expression. Studies of development revealed initial MRNA expression in ventral midbrain as early as embryonic day 14, with steadily increasing expression until adult levels were reached after birth. This work confirms the exquisitely cell- specific expression of this gene in zones that include the mesolimbic/ mesocortical neurons important for reward and reinforcement, suggests a prominent and rapid distribution of transporter protein away from the perikaryon soon after synthesis, and indicates the possibility of significant developmental effect of cocaine on transporter expressed in utero. Transporter MRNA in cocaine-treated and withdrawn animals displayed moderate downregulation in cells of the midline ventral tegmental area. This observation correlates with data on reduced nucleus accumbens transporter binding densities, and suggests a specific mechanism that could contribute to cocaine """"""""craving"""""""" and other clinical withdrawal phenomena.
