Comparative Evaluation of Nuclear Morphology of Equine Oocytes Aspirated in vivo and Stained with Hoechst and Orcein

Abstract

Nuclear maturation of equine oocytes was assessed immediately after in vivo collection. A double-staining technique (Hoechst and orcein) was used on the same oocytes to visualize nuclear morphology, i.e. to evaluate the chromatin configurations of each oocyte after Hoechst in relation to the nuclear morphology after orcein staining. The proportion of oocytes evaluated as germinal vesicle stages was significantly (p < 0.02) lower after Hoechst (14.5%) than after orcein staining (29.0%), while the incidence of the so-called dense chromatin stage was assessed to be higher (p < 0.05) after Hoechst than after orcein staining (14.5 vs. 6.5%). There was no difference between Hoechst and orcein staining in the incidence of diakinesis and germinal vesicle breakdown stages, respectively (44.9 vs. 42.0%), and the same applied for metaphase I (11.6 vs. 8.0%), metaphase II (7.2 vs. 8.0%) and degenerated stages (7.2 vs. 6.5%). It was concluded that the interpretation of the meiotic stages may differ between Hoechst and orcein staining and in a large proportion of equine oocytes the nuclear border may not be visualized on orcein staining.

Establishment of conditions for ovum pick up and IVM of jennies oocytes toward the setting up of efficient IVF and in vitro embryos culture procedures in donkey (Equus asinus)

“…At collection, jennies oocytes contained a germinal vesicle, either with distinct chromatin filaments or with chromatin partly condensed. These meiotic stages were observed in equine oocytes immediately after in vivo collection [44,45]. After 24-hour IVM, some jennies oocytes contained condensed chromatin, and metaphase 1 oocytes were observed after 30 hours.…”

Beta‐mercaptoethanol supplementation of in vitro maturation medium does not influence nuclear and cytoplasmic maturation of equine oocytes

“…The meiotic stage of each oocyte after Hoechst staining was characterized as previously described (Bézard, Bøgh, Duchamp, Hyttel, & Greve, ): GV: diffuse fluorescence of the oocyte nucleus often presenting a cloudy or partly filamentous chromatin pattern (Fig. a); diakinesis (Dia): distinct fine filamentous pattern of often loop‐shaped slightly condensed chromatin, forming an irregularly shaped web (Fig.…”

Ovarian function and morphology in the mare after multiple follicular punctures

“…In the mare, ultrasound-guided follicular puncture has so far mainly been used for research purposes (Bracher et al 1993;Cook et al 1993;Meintjes et al 1995;Alm et al 1997;Brück et al 1997Brück et al , 2000Goudet et al 1997;Carnevale et al 2000;Cochran et al 2000;Weber et al 2001;Bézard et al 2002;Bøgh et al 2003). Embryo development and foaling rates after IVF or intracytoplasmic sperm injection of equine oocytes are still of limited success (Palmer et al 1991;Grøndahl et al 1995;Goudet et al 1998;McKinnon et al 1998).…”