Immunoreactivity for Gonadotropin-Releasing Hormone in Microdialyzed Perfusates of Bovine Mature Follicles In Vitro

The presence of gonadotropin-releasing hormone (GnRH)-like protein has been previously demonstrated in the bovine follicle. This GnRH-like protein was purified and concluded to be histone H2A. However, neither GnRH peptide nor specific GnRH-immunoreactivity (GnRH-IR) has been demonstrated in the bovine follicle so far. Thus, this study focused on the detection of specific GnRH-IR using the second-antibody enzyme immunoassay in isolated bovine mature follicles, and on an examination of the direct effect of luteinizing hormone (LH), endothelin-1 (ET-1), and cytokines on the GnRH-IR using an in vitro microdialysis system (MDS). We further examined a cross-reactivity of the GnRH antibody with bovine histone H2A. GnRH-IR was detected in microdialyzed perfusate from isolated bovine mature follicles at 4.40 ± 0.35 pg/ml (mean ± SEM). Bovine histone H2A showed no GnRH-IR at all. Heat treatment of the extract (100 C for 10 min) did not affect the GnRH-IR. Single infusion of LH, ET-1, or cytokines into the MDS did not affect the GnRH-IR. However, infusion of ET-1 after LH exposure increased the GnRH-IR. These results demonstrate the presence of specific GnRH-IR, that is different from histone H2A, in microdialyzed perfusate of isolated bovine mature follicles in vitro, suggesting that the GnRH-IR may reflect a role of GnRH-like peptide or some peptide structurally similar to GnRH in the local regulation of mature follicles