Hypoxanthine (HX) inhibition of in vitro meiotic resumption in goat oocytes.

ABSTRACT

To improve in vitro maturation and to understand the mechanism for meiotic resumption of oocytes, meiotic progression, and its control by hypoxanthine (HX) were studied in goat oocytes. Ovaries were obtained from a local abattoir, and cumulus-oocyte complexes (COCs) and follicular fluid were collected from follicles of different surface diameters (SDs). The meiotic competence and progression of oocytes were observed, and the concentration of HX in the follicular fluid and culture media was measured by high-performance liquid chromatography (HPLC). Full meiotic competence of goat oocytes was acquired in follicles of >/=1.5 mm in SD with 90% of the oocytes developing to metaphase II (MII) stage after 24 hr in culture. The HX concentration in follicular fluid decreased with follicle development, from the highest level of 1.16 mM in /=5 mm follicles. HX inhibited meiotic resumption of goat oocytes in a concentration-related manner but this inhibitory effect declined gradually. When we renewed the medium at 4 hr of HX-199 (TCM-199 supplemented with 4 mM HX) culture, the percentage of oocytes with intact germinal vesicle (GV) did not increase but decreased significantly instead. HPLC measurement of HX in the HX-199 culture drops indicated that the HX concentration declined from 0 hr to 4 hr of culture and after medium renewal at 4 hr of culture. By adding dibutyryl cAMP (db-cAMP) at medium renewal, we found that db-cAMP held up the decline of GV percentages. Together, these results were consistent with the possibility that the decline of HX inhibitory effect was not due to HX depletion but rather due to the negative feedback of the metabolites on its further uptake by oocytes. Goat oocytes were capable of normal nuclear maturation and activation after temporal arrest by HX, but prolonged exposure to HX induced spontaneous activation.