Effects of ethanol and dimethylsulphoxide on nuclear and cytoplasmic maturation of bovine cumulus-oocyte complexes

Abstract

The influence of small doses of ethanol or dimethylsulphoxide (DMSO) on in vitro maturation (IVM) of bovine cumulus-oocyte complexes (COC) was examined, either after spontanous maturation or after inhibition of meiosis with 6-dimethylaminopurine (6-DMAP) or 3-isobutyl-1-methylxanthine (IBMX). Subsequent to IVM for 23 hr in semidefined serum-free Earle's TCM199 medium, nuclear maturation was assessed cytogenetically, while the combined cytoplasmic and nuclear maturation was measured indirectly by the oocytes' ability to undergo fertilization and further embryonic development. Embryo development was followed until the blastocyst stages at day 9 after insemination. Neither spontanous nuclear maturation nor cleavage was compromised by IVM in ≤1% (v/v) ethanol or ≤1% (v/v) DMSO, nor was the frequency of polyspermy altered. However, IVM in 0.3% or 1% (v/v) ethanol or in 0.4 or 1% (v/v) DMSO negatively affected blastocyst formation, compared to 0% in the control groups (22% and 23% vs. 34%, P < 0.0001, and 29% and 22% vs. 34%, P < 0.01, respectively), whereas the speed of blastocyst formation, assessed as the D7/D9 blastocyst proportion, was not compromised. In oocytes meiotically inhibited with 2 mM 6-DMAP, the presence of ethanol (0.5%, 1%, and 2% [v/v]) induced germinal vesicle breakdown in a dose-dependent manner (32%, 45%, and 68%, vs. 22%, P < 0.0001), however, the oocytes exhibited no further meiotic progression. In oocytes inhibited with 1 and 2 mM IBMX, the presence of ethanol (0.5%, 1%, and 2% [v/v]) significantly (P < 0.05) enhanced the inhibitory effect in a dose-dependent manner by reducing the proportion of the mature (AI-MII) stages (77%, 68%, and 56% vs. 79%, and 33%, 29%, and 18% vs. 39%, respectively). It is concluded that even small doses of ethanol or DMSO can cause profound negative effects on bovine in vitro maturation and subsequent embryo development. Mol. Reprod. Dev. 55:438–445, 2000. © 2000 Wiley-Liss, Inc.