Dopamine receptor of the porcine anterior pituitary gland. Solubilization and characterization.
In the anterior pituitary gland, dopamine controls the release of prolactin from the mammotrophs. The dopamine receptors in the porcine gland have been shown to exist in two different affinity states of equal proportion, one bearing high affinity for agonists and labeled by 3H-agonist-ligands and the other displaying low affinity for agonists. Both forms of the receptor can be labeled by 3H-antagonist-ligands. Dopamine receptors from porcine anterior pituitary membranes can be solubilized with retention of their ability to interact with specific dopaminergic ligands. Treatment of membrane preparations with 1% digitonin resulted in the solubilization of 20-25% of the specific binding sites labeled by [3H] spiroperidol with a specific activity of about 100 fmoles/mg. The receptor was a glycoprotein as assessed by the interaction of these binding sites with agarose-immobilized lectin. [3H]Spiroperidol binding in solubilized preparations was saturable, of high affinity (KD = 570 pM), and to a single class of stereoselective binding sites. Agonist competition for [3H]spiroperidol binding indicated that, whereas the solubilized receptor retained its dopaminergic specificity, the high-affinity interactions of the receptor with agonists present in membranes and sensitive to guanine nucleotides were lost in solubilized preparations. Thus, the KD values calculated from the agonist competition curves for [3H]spiroperidol corresponded to the agonist affinities for the low-affinity state of the receptor documented in membranes. However, high-affinity agonist binding and its sensitivity to guanine nucleotides were preserved when the membrane-bound receptor was prelabeled with the agonist [3H]N-n-propylnorapomorphine prior to solubilization. These results suggest that a component that confers agonist high-affinity binding and guanine nucleotide responsiveness to the receptor is lost during solubilization unless a stable complex is formed with the agonist prelabeled receptor prior to solubilization.
D000818:Animals; D006150:Guanine Nucleotides / Q000494:pharmacology; D037102:Lectins; D010903:Pituitary Gland, Anterior / Q000032:analysis*; D011954:Receptors, Dopamine / Q000187:drug effects / Q000302:isolation & purification* / Q000378:metabolism; D012995:Solubility; D013134:Spiperone / Q000378:metabolism; D013552:Swine; D014316:Tritium; D014909:Wheat Germ Agglutinins
D006150:Guanine Nucleotides; D037102:Lectins; D011954:Receptors, Dopamine; D014909:Wheat Germ Agglutinins; D014316:Tritium; D013134:Spiperone