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Dopamine D1 and D2 Receptors and Their Signal System Present in Coated Vesicles Prepared from Bovine Striatal Tissue

Authors


Address correspondence and reprint requests to Dr. N. Ozaki at Department of Pharmacology, Kanazawa Medical University, Daigaku 1-1, Uchinada-machi, Kahoku-gun, Ishikawa 920-02, Japan.

Abstract

Abstract: Coated vesicles (CVs) isolated from bovine striatal tissue were examined to determine whether they are associated with dopamine signal systems consisting of dopamine D1 and D2 receptors, G proteins, and adenylate cyclase. Dopamine receptors in CVs were characterized by a dopamine D1 receptor antagonist, [3H]SCH 23390, and a dopamine D2 receptor antagonist, [3H]-spiroperidol. The bindings of both ligands were specifically saturable and reversible with a dissociation constant (KD) of 0.65 and 0.5 nM, respectively. Dopaminergic antagonists and agonists inhibited the specific bindings of [3H]SCH 23390 and [3H]spiroperidol in a stereoselective and concentration-dependent manner with an appropriate rank order potency for dopamine D1 or D2 receptors. The regulations of the agonist binding by guanyl-5-ylimidodiphosphate were observed. ADP ribosylation of the CVs with [32P]NAD demonstrated predominant labeling of bands of Mr 47,000–52,000, 42,000–45,000, and 40,000-39,000, which corresponded to the known molecular weights of the α subunits of Gs and Gi proteins. The presence of α and β subunits of G proteins in the CVs was also confirmed by immunoblotting assay. Adenylate cyclase activity, which was stimulated by SKF 38393 and inhibited by dopamine D2 receptor agonists, was present in the CVs. These findings suggest that the dopamine D1 and D2 receptors in the CVs couple with adenylate cyclase via Gs/Gi protein.

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