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Estimation of the apparent affinity of the striatal dopamine receptors for the radioligand [3H]spiperone([3H]spiroperidol)

The characteristics of the receptor population labeled by the ligand [3H]spiperone were determined by several experimental procedures. Varying the assay volumes, and hence both the receptor and ligand concentrations, did not alter the specificity for the dopamine (DA) receptor. The density of binding sites estimated from saturation analyses varied little (6%) over a greater than 50‐fold tissue concentration range. In contrast, variation in tissue concentration did alter the apparent affinity of [3H]spiperone for the DA receptors more than 16‐fold, as determined from saturation analyses. This was most marked at large receptor or tissue concentrations. The standard correction for depletion of the free ligand by that bound to the receptors reduced the range to threefold or 200%. Two separate measurements of the DA receptor affinity, using the rates of association and dissociation and the shift in Ki for DA, gave affinity measurements in the same range as that obtained using low concentrations of receptors in saturation analyses. Therefore, three separate estimates for the affinity of [3H]spiperone binding to the striatal DA receptors agreed that it is probably in the range of 10–15 pM, and single saturation experiments reach this level when the tissue concentration is very low, ie, less than 15 μg protein per 1 ml assay volume.