Functional Dopamine-1 Receptors and DARPP-32 Are Expressed in Human Ovary and Granulosa Luteal Cellsin Vitro1

The catecholamines norepinephrine and dopamine (DA) are present in the human ovary; in particular, in follicular fluid. Norepinephrine activates ovarian α- and β-adrenergic receptors and modulates ovarian steroidogenesis, but the significance of ovarian DA is unclear. We examined whether a DA receptor of the D1-subtype (D1-R) is present in human ovary and in cultured human granulosa luteal cells (GC). Using RT-PCR, we cloned complementary DNAs from adult human ovarian and GC messenger RNAs, which are identical to the human D1-R sequence. In ovarian sections, D1-R protein was identified (by immunohistochemistry) in granulosa cells of large antral follicles, cells of the corpus luteum, as well as in cultured GC. An immunoreactive band of approximately Mr 50,000 was found in cultured luteinized GC using the same antiserum in Western blots. The D1-R in these cells was functional, because DA, alone or in the presence of the β-receptor antagonist propranolol, caused cellular contraction. The selective D1-R agonist SKF-38393 induced a similar change in cytomorphology and increased the levels of media cAMP. SKF-38393 failed, however, to significantly affect basal and hCG-stimulated progesterone release in vitro, indicating that the activation of the D1-R was not directly linked to synthesis of progesterone, the major steroid of human GC. Estradiol synthesis likewise was not affected. Using RT-PCR and immunohistochemistry, we found that GC express DA- and cAMP-regulated phosphoprotein of Mr 32,000 (DARPP-32), a protein typically associated with neurons bearing the D1-R. In cultured GC, DA and SKF-38393 induced increased threonine-phosphorylation of DARPP-32, even in the presence of propranolol but not in the presence of D1-R antagonist SCH-23390. Taken together, the presence of DA and a functional DA receptor and DARPP-32 indicate that a novel, physiological regulatory pathway involving DA exists in the human ovary.