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Hum. Reprod. Advance Access published online on June 30, 2004
Human Reproduction, doi:10.1093/humrep/deh313
© 2004 by European Society of Human Reproduction and Embryology
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Received November 18, 2003
Accepted April 22, 2004

Article

Cryopreservation of metaphase II human oocytes effects mitochondrial membrane potential: implications for developmental competence

Amy Jones 1*, Jonathan Van Blerkom 2, Patrick Davis 2, Andrew A. Toledo 1

1 Reproductive Biology Associates, 1150 Lake Hearn Dr., Suite 600, Atlanta, GA 30342, USA
2 Department of Molecular, Cellular and Developmental Biology University of Colorado, Boulder, CO 80309, USA; Colorado Reproductive Endocrinology, Rose Medical Center, Denver, CO 80220, USA

* To whom correspondence should be addressed. E-mail: hesikaij{at}aol.com.


  Abstract

BACKGROUND: Current outcome results with embryos derived from thawed MII human oocyes are significantly lower than with embryos cryopreserved at the pronuclear stage. Here, we investigated whether freezing-thawing was associated with changes in oocyte mitochondrial polarity ({Delta}{Psi}m) that could influence competence by altering ATP levels or the ability of the cytoplasm to regulate intracellular Ca2+. METHODS: Fresh and thawed uninseminated and unfertilized MII oocytes were stained with the {Delta}{Psi}m-specific probe JC-1 to detect clusters of high-polarized mitochondria (J-aggregate positive) and with the Ca2+- specific probe Fluo-4 to measure changes in intracellular levels of this cation. ATP content per oocyte was measured directly and cortical granules were visualized with a cortical granule-specific probe. RESULTS: A significant difference between fresh and thawed MII oocytes existed for pericortical J-aggregate fluorescence and for the ability of the cytoplasm to increase free Ca2+ in response to ionophore exposure. No significant difference in ATP contents was measured and cryopreservation was not associated with an apparent release of cortical granules. CONCLUSION: Irreversible loss of high {Delta}{Psi}m in thawed oocytes may be associated with defects in Ca2+ signalling after insemination and could have downstream consequences for normal embryogenesis.

Keywords: ATP; calcium; embryo competence; mitochondrial polarity; oocyte cryopreservation.
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