Maturation, fertilization, and the structure and function of the endoplasmic reticulum in cryopreserved mouse oocytes.
Oocyte cryopreservation is a promising technology that could benefit women undergoing assisted reproduction. Most studies examining the effects of cryopreservation on fertilization and developmental competence have been done using metaphase II-stage oocytes, while fewer studies have focused on freezing oocytes at the germinal vesicle (GV) stage, followed by in vitro maturation. Herein, we examined the effects of vitrifying GV-stage mouse oocytes on cytoplasmic structure and on the ability to undergo cytoplasmic changes necessary for proper fertilization and early embryonic development. We examined the endoplasmic reticulum (ER) as one indicator of cytoplasmic structure, as well as the ability of oocytes to develop Ca(2+) release mechanisms following vitrification and in vitro maturation. Vitrified GV-stage oocytes matured in culture to metaphase II at a rate comparable to that of controls. These oocytes had the capacity to release Ca(2+) following injection of inositol 1,4,5-trisphosphate, demonstrating that Ca(2+) release mechanisms developed during meiotic maturation. The ER remained intact during the vitrification procedure as assessed using the lipophilic fluorescent dye DiI. However, the reorganization of the ER that occurs during in vivo maturation was impaired in oocytes that were vitrified before oocyte maturation. These results show that vitrification of GV-stage oocytes does not affect nuclear maturation or the continuity of the ER, but normal cytoplasmic maturation as assessed by the reorganization of the ER is disrupted. Deficiencies in factors that are responsible for proper ER reorganization during oocyte maturation could contribute to the low developmental potential previously reported in vitrified in vitro-matured oocytes.
Lowther
Katie M
KM
Department of Cell Biology, University of Connecticut Health Center, Farmington, Connecticut 06030, USA.
Weitzman
Vanessa N
VN
Maier
Donald
D
Mehlmann
Lisa M
LM
D000818:Animals; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D015925:Cryopreservation; D004721:Endoplasmic Reticulum / Q000502:physiology*; D005260:Female; D005306:Fertilization / Q000502:physiology*; D005307:Fertilization in Vitro; D005615:Freezing / Q000009:adverse effects; D008540:Meiosis / Q000502:physiology; D051379:Mice; D009865:Oocytes / Q000648:ultrastructure; D009866:Oogenesis / Q000502:physiology*