Dopamine Receptor Binding and Quantitative Autoradiographic Study - Drugs of Abuse: Neurological Reviews and Protocols - page 292

pharmacological similarity of these sites with the D 1 and D 2 receptors, it is
often not possible to distinguish between these sites in a heterogeneous tissue
with currently available radioligands. As such the site labeled by any given
radioligand in a such a tissue (i.e., brain) will, of course, be dependent on the
relative affi nities of that ligand for the related subtypes and the population of
sites present in the tissue employed. Thus, although interactions with specifi c
D 1 - or D 2 -like subtypes may be inferred, it is likely that sites labeled by these
radioligands in brain tissue represent a mixture of the related subtypes. Several
radioligands that appear to exhibit selectivity for the D 3 receptor have been
synthesized including [ 3 H]7-hydroxy-2-(di- n -propylamino)tetralin ([ 3 H]7-OH-
DPAT) and [ 3 H]PD 128907. These compounds label similar populations of
sites in rat brain which are generally consistent with that reported for the D 3
receptor (7-9) .
This chapter outlines protocols for radioligand binding and autoradiographic
assays for different subtypes of dopamine receptors in brain. The scope of the
chapter is limited to the widely used, well established protocols for binding
and autoradiographic assays that employ commercially available, tritiated and
iodinated ligands. Radioligand binding assays, or “grind and bind” studies,
can be used to measure receptor density and affi nity in homogenized tissue
samples. This method is rapid and highly quantitative; however, localization
of binding sites is limited by the accuracy of dissection and any variation in
binding within the brain region of interest will be obscured by homogenization.
Receptor autoradiography, on the other hand, has a high degree of anatomical
resolution. However, even when performed in a quantitative manner, the
method is only semiquantitative compared to radioligand binding. Accordingly,
receptor autoradiography is most suited to determining receptor localization
and relative densities in various brain areas or between different treatment
groups. In addition, receptor autoradiography can require extended periods of
time for the exposure of autoradiograms.
Protocols are outlined for saturation analysis of radioligand binding for
D 1 -like receptors using the antagonist ligand [ 3 H]SCH 23390 and D 2 -like
receptors using the antagonist ligand [ 3 H]spiperone ( see Note 1 ). A protocol
for the putatively selective labeling of D 3 sites using [ 3 H]PD 128907 is also
presented. For autoradiographic studies, protocols are outlined for D 1 -like
receptors using the antagonist ligand [ 3 H]SCH 23390, D 2 -like receptors using
the antagonist ligand [ 125 I]iodosulpiride, and D 3 sites using agonist [ 3 H]PD
128907. Procedures are generally similar for the respective assays; however,
specifi c details, such as buffer composition, radioligand concentration, or
incubation time may vary for each ligand. Ligand-specifi c details for each
assay are presented at the relevant step of the protocol.