AGONISTS AND ANTAGONISTS DIFFERENTIALLY REGULATE THE HIGH-AFFINITY STATE OF THE D2(L) RECEPTOR IN HUMAN EMBRYONIC KIDNEY 293-CELLS

Studies with radiolabeled antagonists have revealed that both agonists and antagonists induce up-regulation of D2 dopamine receptors in cell s transfected to express D2(L) or D2(S) receptors. The regulation indu ced by agonists, but not antagonists, was synergistic with cAMP analog ues, and differences in the time courses of the effects of agonists an d antagonists have been observed. These findings have been extended by using a radiolabeled agonist to investigate agonist- and antagonist-i nduced regulation of the high affinity state of the D2(L) dopamine rec eptor in transfected HEK 293 cells. Exposure to agonists decreased the proportion of receptors in the high affinity, agonist-preferring stat e. Exposure to antagonists, however, led to an increase in the density of receptors with a high affinity for agonists. The effects of both a gonists and antagonists on the agonist-preferring receptors occurred w ithout a lag and were time and dose dependent. Inhibition of forskolin -stimulated cAMP accumulation by agonists was not affected by exposure of the cells to the antagonist (-)-sulpiride. Desensitization was see n after exposing cells to the agonist quinpirole for 1.5 hr, suggestin g that the rapid loss of high affinity binding sites represents an unc oupling of the receptor from the G protein that mediates inhibition of adenylyl cyclase. Pretreatment of cells with the protein synthesis in hibitor cycloheximide did not block the quinpirole-induced loss of rec eptors with a high affinity for agonists. The effect of (-)-sulpiride on high affinity binding sites was blocked by cycloheximide, but only after incubation of cells for sufficient time to induce an increase in the total number of receptors. After incubation of cells with (-)-sul piride for a short time, the increase in the number of receptors with a high affinity for agonists was unaffected by cycloheximide. These re sults suggest that the increase in agonist binding after brief exposur e to an antagonist is due to interactions of the receptor with one or more G proteins that are not coupled to inhibition of adenylyl cyclase , whereas the increase in agonist binding at later time points is asso ciated with the antagonist-induced up-regulation.