Functional oxytocin receptors in bovine granulosa cells

The aim of this study was to identify the presence of functional oxytocin (OT) receptors on bovine granulosa cells. Freshly prepared bovine granulosa cells from small (3-5 mm in diameter) or preovulatory (mature) follicles were examined for OT receptors by a radioreceptor assay. Scatchard analysis revealed that both binding capacity and affinity in granulosa cells from small follicles were significantly higher than those in granulosa cells from mature follicles (p < 0.01). With use of a reverse transcriptase polymerase chain reaction analysis, expression of OT receptor mRNA was detected in granulosa cells obtained from both small and mature follicles. When the granulosa cells obtained from small follicles were cultured in Dulbecco's Modified Eagle's Medium and Ham's F-12 medium (1:1 [v:v]) with 10% calf serum up to 72 h, as the period of culture was prolonged, the concentration of OT receptor decreased with increases of progesterone and OT release in the medium. However, the binding affinity was not changed during culture for 72 h. When bovine follicular oocytes with cumulus oophorus were cultured for 24 h in tissue culture Medium-199 with 10% fetal calf serum and OT (0-10 nM), the percentages of oocytes reaching maximum cumulus expansion were significantly increased at 0.5, 1, and 10 nM OT, although nuclear maturation in oocytes surrounded by compact cumulus cells was not affected by the addition of OT. Coexposure with OT antagonist blocked the stimulatory effect of OT on cumulus expansion, confirming the specificity of the effect. Furthermore, anti-OT rabbit serum inhibited the percentages of oocytes with expanded cumulus compared to those supplemented with normal rabbit serum (p <0.05). The overall results indicate the presence of functional OT receptors in bovine granulosa cells and support the hypothesis that OT plays a role (or roles) in regulating the function of granulosa cells as an autocrine factor during follicular growth.