Collection and evaluation of the pre-ovulatory equine oocyte

Abstract

A series of experiments was conducted to develop a procedure for consistent, repeatable collection of oocytes from the pre-ovulatory follicle of the mare. Experiment 1 resulted in the collection of 26 oocytes from 30 equine ovaries excised at ovariectomy, necropsy or slaughter. In experiment 2, in situ follicular aspiration with a sterile needle and syringe was performed on 19 mares. From 37 aspirations, four oocytes were recovered for a 10% recovery rate. In experiment 3, ovaries were visualized during each of two aspiration techniques which involved a surgical procedure. Use of the needle and syringe as per experiment 2 resulted in successful oocyte recovery in one of seven (14%) attempts. A continuous irrigation vacuum system, developed for use during laparotomy, resulted in successful collections of oocytes from six of 10 attempts (60%). In experiment 4, oocytes were recovered from seven of 18 attempts (38%) at in situ follicular aspiration using a double lumen needle attached to the continuous irrigation vacuum system. In experiments 2, 3 and 4, certain mares were randomly subjected to stimulation of follicular maturation by exogenous hormones. Oocyte recovery was significantly increased in treated mares as compared with non-treated mares. Twenty-two of the oocytes collected during the previous experiments were examined by light and transmission electron microscopy. Oocytes from both treated and non-treated mares were evaluated. Oocytes were at various stages of meiotic development or degeneration. Eighteen of the non-degenerate oocytes had intact nuclear membranes and four had undergone germinal vesicle breakdown. Organization and appearance of cellular organelles within the oocytes were similar to those of other mammalian oocytes, with the exception of the absence of granular endoplasmic reticulum, typical Golgi complex configuration and absence of a reticulate nucleolus. Four oocytes in more advanced stages of meiosis exhibited varying degrees of nuclear membrane disintegration, chromosome alignment and polar body formation. Exogenous hormonal stimulation before collection did not appear to affect ultrastructural morphology of the follicular oocytes...