Effect of follicular components on meiotic arrest and resumption in horse oocytes.

Two experiments were conducted to evaluate the effect of follicular components on the maintenance of meiotic arrest in horse oocytes. In Expt 1, oocytes were incubated for 24 h with follicular fluid, or with granulosa cells suspended either in medium or in follicular fluid at 25 x 10(6) cells ml-1. None of the treatments resulted in significant maintenance of the germinal vesicle stage over that of non-suppressive control. Culture with follicular fluid plus granulosa cells resulted in a significantly higher proportion of oocytes at metaphase I compared with controls. In Expt 2, oocytes were divided into those originally having compact or expanded cumuli. Oocytes were cultured with sheets of mural granulosa or sections of follicle wall, or after injection into intact dissected follicles. After incubation, half of the oocytes from each suppressive treatment were matured for 24 h. All three suppressive treatments were effective in maintaining oocytes at the germinal vesicle stage (no significant difference from control oocytes fixed directly after removal from the follicle). However, no treatment maintained normal viability of oocytes, as significantly fewer oocytes were at metaphase II after all the suppression-maturation treatments compared with the maturation control. The highest rate of post-suppression maturation was found in the mural granulosa treatment. Within this treatment, the proportion of oocytes in metaphase II was significantly higher for oocytes with expanded than for oocytes with compact cumuli (31% versus 11%, respectively; P < 0.05). Suppression by injection into an intact follicle was associated with a lack of progression to metaphase II during subsequent maturation.(ABSTRACT TRUNCATED AT 250 WORDS)

In vitro maturation of horse oocytes: characterization of chromatin configuration using fluorescence microscopy.

The chromatin configuration of resting horse oocytes and the time course of in vitro oocyte maturation was characterized using a fluorescent, DNA-specific label. Oocytes were classified as having either compact (CP) or expanded (EX) cumuli at the time of collection. Centrifugation of oocytes was effective in allowing visualization of the germinal vesicle. Two main chromatin configurations were found in oocytes known to have a germinal vesicle: condensed chromatin (CC), in which the chromatin formed a dense mass surrounding the nucleolus; and fluorescing nucleus (FN), in which the entire nucleus, containing diffuse or spotty chromatin, was visible. The proportion of CC to FN was higher for oocytes with EX cumuli. At time 0, 78% of CP oocytes and 73% of EX oocytes were in the germinal vesicle stage. Significantly more EX than CP oocytes were in metaphase I or II at time 0. In both CP and EX groups, maturation had not begun after 8 h of incubation. Maximal maturation occurred after 24 h for oocytes in the EX group, whereas CP oocytes continued to mature between 24 and 32 h. The percentage of EX oocytes in metaphase I did not change between 24 and 32 h, indicating a possible arrest of some EX oocytes at metaphase I. There was no difference in the percentage of oocytes at metaphase II between the CP and EX groups after 32 h of incubation.