Dual coupling and down regulation of human FSH receptor in CHO cells

Abstract

The FSH receptor is a member of the family of G protein-coupled receptors that activate adenylyl cyclase. The binding of agonist to cell surface receptors leads to a reduction in the intensity of the response to continuous stimulation, a process that is usually referred to as desensitization. Although the exact mechanism is not fully understood, the molecular cloning of the FSH receptor has made it possible to study desensitization in transfected cell lines. In this experiment FSH-induced desensitization was studied using Chinese hamster ovary cells expressing a functional human FSH receptor (CHO-FSHR cells). Stimulation of the CHO-FSHR cells with 10 ng/ml human FSH resulted in a decreased sensitivity to a second FSH stimulation. This decrease in FSH-induced cAMP production was observed within 2 h, and exposure of cells to FSH for 20 h led to a 70–80 % inhibition of cAMP formation. Moreover, the desensitization effect observed in CHO cells was mimicked by forskolin and, therefore, was mediated by cAMP. Incubation of cells with 125I-FSH showed an efficient internalization of the ligand in the CHO-FSHR cells. The CHO-FSHR cells rapidly internalized approximately 30% of the receptor-associated 125I-FSH by 2 h and 50% by 4 h. The responsiveness of individual CHO-FSHR cells to FSH was studied and administration of human FSH (30 ng/ml) induced a rapid rise in cytosolic calcium, reaching a peak at 6 sec. The data that human FSH can increase intracellular calcium in cells transfected with the FSH receptor cDNA reveal the possibility for the human FSH receptor to couple to both adenylyl cyclase and phospholipase C cascades.