Influence of oocyte collection technique on initial chromatin configuration, meiotic competence, and male pronucleus formation after intracytoplasmic sperm injection (ICSI) of equine oocytes*

Abstract

There is a great variability in the success of horse oocyte maturation and fertilization among laboratories. This study was conducted to determine if the meiotic and developmental competence of horse oocytes could be dependent on the method of oocyte collection, i.e., aspiration of follicular fluid with a vacuum apparatus, or opening follicles and scraping the granulosa layer. Horse oocytes were recovered from abattoir ovaries by aspiration or scraping and classified as having compact (Cp), expanded (Ex), or partial (P) cumuli. In Experiment 1 (Part A in May and Part B in October), oocytes were fixed immediately after collection to assess whether the collection method influenced the initial chromatin configuration of oocytes. In Experiment 2, in vitro maturation rates of oocytes recovered by aspiration or scraping were compared. In Experiment 3, oocytes were matured in vitro and submitted to intracytoplasmic sperm injection (ICSI). Initial chromatin configuration differed according to collection method in that there was a significantly higher prevalence of diffuse chromatin within the germinal vesicle in oocytes recovered by scraping than in oocytes recovered by aspiration (29/87, 33% and 28/166, 17%, respectively; P < 0.01). Maturation of oocytes to metaphase II did not significantly differ between scraped and aspirated oocytes (56/101, 55.4 % vs. 65/106, 61.4%, respectively). The overall pronucleus formation rate after ICSI of oocytes recovered by scraping was not significantly different than that of oocytes recovered by aspiration (50/99, 52.6% vs. 50/85, 68.5 %, respectively); however, the rate of abnormal fertilization was significantly higher for oocytes collected by aspiration (14/73, 19% vs. 6/94, 6%, respectively; P <0.05). These results demonstrate that the collection method affects the population of recovered oocytes and may contribute to differences in results observed among laboratories working with horse oocytes.

Effect of Oocyte Recovery Techniques on &lt;i&gt;in Vitro&lt;/i&gt; Production of Swine Embryos

“…Although they reported that less MII stage oocytes were observed after in vitro maturation in the ovary slicing group compared to the other three oocyte recovery techniques, no significant difference was observed between puncture and aspiration groups. These similar oocyte maturation rates were also observed in our study and agree with data obtained in equine [6].…”

“…Data presented as least squares means ± SEM. structures were recovered only on visible follicles at the ovary surface by the aspiration method when compared to the scraping method (follicles were opened with a scalpel blade and the granulosa cell layer scraped with a curette) [6] [7].…”

“…The comparison between these two techniques has been performed on bovine [4] [5], equine [6] [7], goats [8], sheep [9] and camels [10], but not in swine.…”

Factors affecting developmental competence of equine oocytes after intracytoplasmic sperm injection

“…Many laboratories have selected oocytes with compact cumuli (Cp) for culture and discarded oocytes with expanded cumuli (Ex) (Fulka & Okolski 1981, Willis et al 1991, Choi et al 1993, Goudet et al 1998, Li et al 2000, Tremoleda et al 2001, as is done in most other species. However, nuclear maturation rates of Ex equine oocytes have been shown to be similar to (Zhang et al 1989, Hinrichs et al 1993, Dell'Aquila et al 2001, or higher than (Hinrichs et al 1995, Alm & Hinrichs 1996, Hinrichs & Williams 1997, Hinrichs & Schmidt 2000, Choi et al 2002b, those for Cp oocytes. Information on the developmental competence of these two types of oocytes is limited.…”

Somatic cell nuclear transfer in horses: effect of oocyte morphology, embryo reconstruction method and donor cell type

“…On average it is possible to recover 3 -4 immature oocytes per ovary from the ovaries of slaughtered animals (Galli & Lazzari 2001; oocyte retrieval from live donors by OPU is slightly more variable and can be from 3 to 6 oocytes per session of OPU , Lorenzo et al 2002. The maturation rate of horse oocytes is also quite variable, averaging between 25 and 70% in published studies (Hinrichs & Williams 1997, Carneiro et al 2001, Dell'Aquila et al 2001, Bogh et al 2002, Choi et al 2002, Lorenzo et al 2002. Interestingly, in the horse the recovered cumulus-oocyte complexes are a mixed population with either a compact or an expanded cumulus.…”