Solubilization and Characterization of Striatal Dopamine Receptors

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Abstract
Dopamine [DA] receptor binding proteins were solubilized with the detergent 3-(3-cholamidopropyl) dimethylammonio-2-hydroxy-1-propanesulfonate (CHAPSO) from bovine and rat striatal membranes. The binding of the DA antagonist [3H]spiroperidol ([3H]Spi) to the solubilized DA receptors was determined by the polyethyleneglycol method. The CHAPSO-solubilized DA receptor binding proteins remain in the supernatant fraction following centrifugation at 100,000 .times. g for 2 h. The CHAPSO-solubilized DA receptor proteins as well as the prelabeled [3H]Spi-receptor protein complex, bind specifically to wheat germ agglutinin(WGA)-agarose columns, which is consistent with an identification as glycoproteins. HPLC [high performance liquid chromatography] analysis of the CHAPSO-solubilized, prelabeled [3H]Spi-receptor protein complex (CHAPSO preparation) reveals association with a high-MW form, indicating the formation of aggregates and/or micelles. Treatment of the WGA-agarose-bound [3H]Spi-receptor protein complex with digitonin (CHAPSO-digitonin preparation) results in dissociation of the high MW-form into lower-MW forms. The HPLC profile of the prelabeled [3H]Spi-receptor complex in the CHAPSO-digitonin preparation reveals 2 radioactive peaks. The major peak had a retention time of 16 min, corresponding to an apparent MW of 175,000, whereas the minor peak had a retention time of 21 min, corresponding to an apparent MW of 49,000. The CHAPSO-solubilized DA receptor binding proteins are sensitive to modulation by GTP, indicating that the association with the GTP binding component is preserved in the soluble state. The potencies of DA antagonists and agonists for inhibiting the binding of [3H]Spi to CHAPSO-solubilized DA receptor proteins are similar to those for membrane-bound proteins. Chronic treatment with haloperidol increases the Bmax, and does not change the KD for [3H]Spi in the CHAPSO-solubilized and in the membrane-bound preparations. The CHAPSO-solubilized DA receptor proteins retain the binding characteristics of the supersensitive membrane-bound DA receptors.