Densities and distribution of D1-dopamine and D2-dopamine receptors were investigated in vitro using [3H]SCH 23390 and [3H]raclopride in receptor binding assays and autoradiography on human post mortem whole hemisphere slices to serve as anatomical correlates to PET studies using [11C]SCH 23390 and [11C]raclopride. In addition, the levels of dopamine and its metabolites were determined by HPLC in various brain regions. Both dopamine receptor subtypes, as well as dopamine, HVA and DOPAC, were primarily found in the basal ganglia. Very high densities of D1-dopamine receptors were found particularly in the medial caudate nucleus, whereas D2-dopamine receptors were evenly distributed throughout the caudate. The densities of D1- and D2-dopamine receptors were similar in the caudate nucleus and the putamen, whereas there were 4 to 7 times higher densities of the D1- than of the D2-dopamine receptors in several limbic and neocortical regions. The receptor distribution in the autoradiographic study was consistent with that demonstrated in the living human brain using [11C]SCH 23390 and [11C]raclopride.

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Neuropeptide Y (NPY) is a regulatory molecule in both the central and peripheral nervous system (1), and at least two types of NPY receptors have been described, Y1 and Y2 receptors (Y1-R and Y2-R) (2). Both in vitro (3) and in vivo (4-6) studies have demonstrated high-affinity 1251-labeled NPY binding sites on dorsal root ganglion (DRG) neurons, and these sites may be of both the Y1-R and the Y2-R types (3-6). Recently, the molecular cloning ofthe cDNA sequence of the Y1-R has been reported (7-9), and it could be shown that NPY Y1-R mRNA is strongly expressed in DRG neurons, mainly of the small type (10,11). In the present study we have raised an antibody to the C-terminal portion of the Y1-R and used immunohistochemistry to analyze the cellular distribution of the receptor protein at the spinal level and also, in a preliminary way, in the brain. METHODSProduction of Antiserum. A C-terminal peptide (amino acids 355-382) of the NPY Y1-R (7-9) was synthesized, coupled to thyroglobulin by using glutaraldehyde, and injected into six rabbits. Sera obtained after repeated boosters were used for immunostaining.Immunocytochemisby. Adult male Sprague-Dawley rats (200-to 300-g body weight; HALAB, Stockholm) were anesthetized, and the left lumbar 4, 5, and 6 (L4, L5, and L6) dorsal roots and the sciatic nerve were compressed unilaterally with a forceps for 3 sec. After 24 hr the operated rats and normal rats were anesthetized and fixed by perfusion with picric acid-containing formalin. Brain, spinal cord, DRGs, and sciatic nerves were processed for immunostaining with antiserum Y1-2D using both immunofluorescence (dilution 1:4000) or avidin-biotinylated horseradish peroxidase complex (ABC) (1:15,000) staining. Antisera against the 8-opioid receptor (12) or -opioid receptor (8-or -opioid-R; unpublished data) were also used (1:400 or 1:4000, respectively). Double staining was carried out with a mixture of the rabbit Y1-R antiserum (1:4000) and a monoclonal mouse antibody against calcitonin gene-related peptide (CGRP) (1:400; Celltech, Slough, Berkshire, U.K.) and a mixture of fluorescein isothiocyanate-conjugated donkey anti-rabbit and lissamine rhodamine B sulfonyl chloride (LRSC)-conjugated donkey anti-mouse antibodies (1:40; both fromThe Jackson Laboratory). The sections were examined under a Nikon Optiphot II microscope or a Bio-Rad MRC-600 laser scanning confocal imaging system. The specificity of the antisera was tested by absorption with synthetic peptides (1 pM for 24 hr at 40C.

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