Reproduction Abstracts

Reproductive ageing in females is associated with reduced oocyte maturation potential and developmental competence. Oocyte chromatin configuration and metabolism, have recently been established as correlates of oocyte developmental competence in a number of species. Here, we have evaluated the effect of reproductive ageing on oocyte chromatin morphology and amino acid turnover in germinal vesicle (GV)-staged, ovine oocytes.

Cumulus–oocyte complexes were harvested from follicles >2 mm diameter from abattoir-derived adult (n=133) and prepubertal (n=135) sheep ovaries. Denuded GV oocytes were cultured individually in 1 μl drops of serum-free IVM medium containing 50 μM cilostamide for 6 h (t6) at 38 °C and 5% CO_2. Chromatin morphology was assessed by fluorescent microscopy at time 0 (t0) and t6 after staining in 10 μg/ml DAPI. Amino acid metabolism was assayed in spent culture media using HPLC.

Chromatin morphology was significantly different in GV oocytes from prepubertal vs adult sheep (P<0.05). At t0, no difference in chromatin morphology was observed; most oocytes contained net-like, uncondensed chromatin (58.8% adult vs 70.4% prepubertal; P>0.05). At t6, significantly more adult oocytes (60.4%, n=53) than prepubertal oocytes (26.5% n=55, P<0.05) had progressed to the clumped chromatin configuration associated with condensation before GVBD. Amino acid profiling revealed adult oocytes consumed more isoleucine (P=0.001) and leucine (P=0.026), and produced more glutamic acid (P<0.001) and alanine (P<0.001), but less tyrosine (P<0.001) than prepubertal oocytes. Aspartic acid metabolism switched from low production to consumption as age increased.

In conclusion, reproductive ageing affected both amino acid metabolism and the dynamics of chromatin configuration in GV-staged sheep oocytes in vitro.