Effect of holding at room temperature on initial chromatin configuration and in vitro maturation rate of equine oocytes.

The relationship of holding time in media at room temperature (approximately 22 degrees C) to initial chromatin configuration and rate of in vitro maturation (IVM) of equine oocytes was determined. Only oocytes having a complete, compact cumulus were used in this study. Oocytes were removed from ovaries 3.5-8 h after slaughter and were put into one of four treatment groups: (1) immediate/fix (IF) = immediate fixation following removal from the ovary; (2) delay/fix (DF) = fixation after oocytes were held 1-4 h in medium at room temperature; (3) immediate/mature (IM) = immediate placement into maturation medium at 5% CO2 at 38.2 degrees C; and (4) delay/mature (DM) = placement into maturation medium at 5% CO2 at 38.2 degrees C after oocytes were held 1-4 h in medium at room temperature. Chromatin configurations in fixed oocytes were classified as fluorescent nucleus (FN), condensed chromatin (CC), fibrillar, intermediate, or fibrous germinal vesicle (GV). Other classifications were Metaphase I, II, or degenerating/abnormal. Oocytes held at room temperature before fixation (DF) had a lower proportion of oocytes in the fibrous GV, fibrillar and intermediate configurations than did oocytes fixed immediately (IF; 1/54, 2% versus 15/51, 30%, respectively, P < 0.001). Oocytes held before fixation tended to have a higher percentage of both the CC and FN configurations than did oocytes fixed immediately (CC: 22/40, 55% versus 11/36, 31%, respectively, P = 0.056; FN: 17/40, 43% versus 10/36, 28%, respectively, P = 0.066). Holding of oocytes did not affect the rate of resumption of meiosis or the rate of degeneration in culture; however, of oocytes resuming meiosis, more oocytes in the delayed than in the immediate maturation group had reached MII by 24h culture (14/15, 93% versus 8/15, 53%, respectively, P = 0.018).