PH-DEPENDENCE OF LIGAND-BINDING TO D-2 DOPAMINE-RECEPTORS

The binding of a range of ligands to D-2 dopamine receptors in bovine caudate nucleus and recombinant CHO cells expressing the receptor has been determined at different pH values between 4.5 and 8.5. The maximum number of D-2 dopamine receptor binding sites in each tissue was not affected by the change in pH, but the affinity of ligands for binding to the receptors was decreased as the pH was decreased. For classical dopamine antagonists, e.g. spiperone and haloperidol, the data on pH dependence of the dissociation constant for receptor binding indicated that the protonation of a single ionizing group on the receptor (pK(a) similar to 6) influenced the binding process. For antagonists of the substituted benzamide class, the data indicated that the protonation of two ionizing groups (pK(a) between 6 and 7) influenced the ligand binding process. These ionizing residues may correspond to Asp 114 for the classical antagonists and Asp 114 and Asp 80 for the substituted benzamide antagonists. Further evidence for the participation of carboxyl residues in the ligand binding process was obtained from the inhibition by N,N'dicyclohexylcarbodiimide of the binding of [H-3]spiperone and [H-3]YM 09151-2 to D-2 receptors in the recombinant CHO cells.