Julie K. Staley

The dopamine transporter (DAT) was localized in normal human brain tissue by light microscopic immunocytochemistry by using highly specific monoclonal antibodies. Regional distribution of DAT was found in areas with established dopaminergic circuitry, e.g., mesostriatal, mesolimbic, and mesocortical pathways. Mesencephalic DAT-immunoreactivity was enriched in the dendrites and cell bodies of neurons in the substantia nigra pars compacta and ventral tegmental area. Staining in the striatum and nucleus accumbens was dense and heterogeneous. Mesocortical DAT immunoreactivity in motor, premotor, anterior cingulate, prefrontal, entorhinal/perirhinal, insular, and visual cortices was detected in scattered varicose and a few nonvaricose fibers. Varicose fibers were relatively enriched in the basolateral and central subnuclei of amygdala, with sparser fibers in lateral and basomedial subnuclei. Double-labeling studies combining DAT and tyrosine hydroxylase (TH) immunostaining in the ventral mesencephalon showed two subpopulations of dopaminergic neurons differentiated by the presence or absence of DAT-immunoreactivity in the A9 and A10 cell groups. In other dopaminergic cell groups (All, A13-A15), TH-positive hypothalamic neurons showed no detectable DAT-immunoreactivity. However, fine DAT-immunoreactive axons were scattered throughout the hypothalamus, particularly concentrated along the medial border, with more coarse axons present along the lateral border. These findings demonstrate that most mesotelencephalic dopamine neurons of human brain express high levels of DAT throughout their entire somatodendritic and axonal domains, whereas a smaller subpopulation of mesencephalic dopamine cells and all hypothalamic dopamine cell groups examined express little or no DAT. These data indicate that different subpopulations of dopaminergic neurons use different mechanisms to regulate their extracellular dopamine levels.

The mesolimbic dopaminergic system plays a primary role in mediating the euphoric and rewarding effects of most abused drugs. Chronic cocaine use is associated with an increase in dopamine neurotransmission resulting from the blockade of dopamine uptake and is mediated by the activation of dopamine receptors. Recent studies have suggested that the D3 receptor subtype plays a pivotal role in the reinforcing effects of cocaine. The D3 receptor-preferring agonist 7-hydroxy-N,N-di-n-propyl-2-aminotetralin (7-OH-DPAT) is a reinforcer in rhesus monkeys trained to self-administer cocaine, but not in cocainenaive monkeys. In vitro autoradiographic localization of [3H]-(+)-7-OH-DPAT binding in the human brain demonstrated that D3 receptors were prevalent and highly localized over the ventromedial sectors of the striatum. Pharmacological characterization of [3H]-(+)-7-OH-DPAT binding to the human nucleus accumbens demonstrated a rank order of potency similar to that observed for binding to the cloned D3 receptor expressed in transfected cell lines. Region-of-interest analysis of [3H]-(+)-7-OH-DPAT binding to the D3 receptor demonstrated a one- to threefold elevation in the number of binding sites over particular sectors of the striatum and substantia nigra in cocaine overdose victims as compared with age-matched and drug-free control subjects. The elevated number of [3H]-(+)-7-OH-DPAT binding sites demonstrates that adaptive changes in the D3 receptor in the reward circuitry of the brain are associated with chronic cocaine abuse. These results suggest that the D3 receptor may be a useful target for drug development of anticocaine medications.

OBJECTIVE: Evidence of a relationship between genotype and binding availability was assessed for the dopamine and serotonin transporter genes. METHOD: The authors assessed dopamine transporter genotype at the SLC6A3 3' variable number of tandem repeats (VNTR) polymorphism and serotonin transporter genotype at the SLC6A4 promotor VNTR polymorphism in 30 healthy subjects who also underwent single photon emission computed tomography with [(123)I]beta-CIT. RESULTS: Subjects homozygous for the 10-repeat allele at the SLC6A3 locus demonstrated significantly lower dopamine transporter binding than carriers of the nine-repeat allele. There was no effect of SLC6A4 genotype upon serotonin transporter binding. CONCLUSIONS: These findings suggest that genetic variation at the SLC6A3 3' VNTR polymorphism may modify dopamine transporter function.

UNLABELLED: A polymorphism involving a variable number of tandem repeats (VNTR) has been described in the 3' untranslated region of the gene (SLC6A3) coding for the dopamine transporter (DAT). This polymorphism has 2 common alleles, designated as 10-repeat (*10R) and 9-repeat (*9R), that have been linked with several human clinical phenotypes. Previous investigations of the effects of the SLC6A3 polymorphism on DAT availability in smaller samples of humans have yielded divergent results. METHODS: We assessed genotype at the SLC6A3 promoter VNTR polymorphism in 96 healthy European Americans (age range, 18-88 y) who also underwent SPECT with (123)I-2beta-carbomethoxy-3beta-(4-iodophenyl)tropane (beta-CIT) for measurement of striatal DAT protein availability. A ratio of specific to nondisplaceable brain uptake (i.e., V(3)'' = [striatal -occipital]/occipital), a measure proportional to the binding potential, was derived. For this analysis, 9-9 homozygotes and 9-10 heterozygotes were grouped as SLC6A3 *9R carriers and contrasted with SLC6A3 *10R homozygotes. RESULTS: The SLC6A3 *9R carriers had significantly higher striatal DAT availability (V(3)'') than did the SLC6A3 *10R homozygotes, controlling for age (F(1,93) = 6.25, P = 0.014, analysis of covariance). The *9R carriers (n = 41, 49.8 +/- 19.5 y) had a mean increase in striatal DAT availability of 8.9% relative to the *10R homozygotes (n = 53, 49.9 +/- 19.2 y). Striatal subregion analysis revealed that the effect of DAT genotype was significant for both the caudate and the putamen. CONCLUSION: These results support the interpretation of higher DAT levels in association with the *9R allele in European Americans and may relate to previously observed associations between DAT genotype and neuropsychiatric diseases.