Effects of Cryopreservation on the Meiotic Spindle, Cortical Granule Distribution and Development of Rabbit Oocytes

Authors

  • E Jiménez-Trigos,

    1. Institute of Science and Animal Technology, Laboratorio de Biotecnología de la Reproducción, Universidad Politécnica de Valencia, Valencia, Spain
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  • C Naturil-Alfonso,

    1. Institute of Science and Animal Technology, Laboratorio de Biotecnología de la Reproducción, Universidad Politécnica de Valencia, Valencia, Spain
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  • JS Vicente,

    1. Institute of Science and Animal Technology, Laboratorio de Biotecnología de la Reproducción, Universidad Politécnica de Valencia, Valencia, Spain
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  • F Marco-Jiménez

    1. Institute of Science and Animal Technology, Laboratorio de Biotecnología de la Reproducción, Universidad Politécnica de Valencia, Valencia, Spain
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Author’s address (for correspondence): Francisco Marco Jiménez, Laboratory of biotechnology of reproduction. Institute of Science and Animal Technology (ICTA) at the Polytechnic University of Valencia, C/Camino de Vera s/n, 46022 Valencia, Spain. E-mail: fmarco@dca.upv.es

Contents

Although much progress has been made in oocyte cryopreservation since 1971, live offspring have only been obtained in a few species and in rabbits. The aim of our study was to evaluate the effect of vitrification and slow freezing on the meiotic spindle, cortical granule (CG) distribution and their developmental competence. Oocytes were vitrified in 16.84% ethylene glycol, 12.86% formamide, 22.3% dimethyl sulphoxide, 7% PVP and 1% of synthetic ice blockers using Cryotop as device or slow freezing in 1.5 m PROH and 0.2 m sucrose in 0.25 ml sterile French mini straws. Meiotic spindle and CG distribution were assessed using a confocal laser-scanning microscope. To determine oocyte competence, in vitro development of oocytes from each cryopreservation procedure was assessed using parthenogenesis activation. Our data showed that oocytes were significantly affected by both cryopreservation procedures. In particular, meiotic spindle organization was dramatically altered after cryopreservation. Oocytes with peripheral CG distribution have a better chance of survival in cryopreservation after slow-freezing procedures compared to vitrification. In addition, slow freezing of oocytes led to higher cleavage and blastocyst rates compared to vitrification. Our data showed that, in rabbits, structural alterations are more evident in vitrified oocytes than in slow-frozen oocytes, probably as a consequence of sensitivity to high levels of cryoprotectants. Slow-freezing method is currently the recommended option for rabbit oocyte cryopreservation.

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