Recently a novel member of the transforming growth factor beta (TGFbeta) superfamily termed growth/differentiation factor-9 (GDF-9) was shown to be expressed in ovaries of mice and humans, and to be essential for normal follicular development beyond the primary (type 2) follicle stage in mice. In the present study, the gene for ovine GDF-9 was isolated and characterized, and expression of GDF-9 mRNA in ovaries of domestic ruminants was examined. The predicted amino acid sequence of ovine GDF-9 is 77% and 66% homologous to human and mouse GDF-9, respectively. Specific hybridization using homologous 35S-antisense probes was restricted to oocytes. In contrast to similar studies in mice in which GDF-9 was first detected beginning at the primary (type 2) follicle stage, in ovine and bovine ovaries GDF-9 mRNA was expressed beginning at the primordial (type 1) follicle stage. The observed timing and pattern of GDF-9 expression in oocytes of domestic ruminants is consistent with a role for GDF-9 in the initiation and maintenance of folliculogenesis in these species, and supports the general concept that early stages of follicular growth and development are regulated by intraovarian factors.

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We have evaluated the in vivo and in vitro regulation and temporal expression of messenger RNA (mRNA) for prostaglandin (PG) G/H synthase-2 (PGHS-2) and two specific PG receptors, PGF2alpha receptor (FP receptor) and PGE receptor EP3 subtype (EP3 receptor), in bovine preovulatory follicular cells and luteal cells. An in vivo study showed that PGHS-2 mRNA was not detected in granulosa cells and was highly but transiently induced by the LH surge before ovulation. FP and EP3 receptor mRNAs were present at extremely low concentrations in granulosa or thecal cells and did not increase before ovulation. Messenger RNA for FP receptor increased more than 500- and 2500-fold at 24 and 48 h after ovulation, respectively, and these high amounts were maintained at midluteal phase. On the other hand, mRNA for EP3 receptor remained low with FP receptor mRNA 1000-fold greater than EP3 receptor mRNA in the corpus luteum. In vitro culture of bovine granulosa cells using hCG, forskolin, and phorbol didecanoate demonstrated that induction of FP receptor mRNA was mediated through protein kinase (PK) A. In contrast, EP3 receptor mRNA was stimulated through PKC. PGHS-2 was acutely ( < 12 h) increased by PKA, and to a lesser extent by PKC. Temporal expression of FP receptor mRNA is not consistent with the involvement of FP receptor in ovulation and suggests that PKA stimulates PGHS-2 and FP receptor mRNA by distinct mechanisms.

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