Water and DMSO membrane permeability characteristics of in-vivo- and in- vitro-derived and cultured murine oocytes and embryos

Although embryo cryopreservation is routine for many mammalian species, itis important to know how the fundamental cryobiology of these cells changeswith development. Progressive cleavage divisions result in a reduction inthe blastomere surface area available for water and cryoprotectant masstransport. Therefore, the membrane permeability of murine oocytes, zygotes,2-cell, 4-cell, and 8-cell embryos to water (Lp), and dimethylsulphoxide(PDMSO), and the reflection coefficient, sigma (sigma) were determined.Oocytes or zygotes were recovered, cumulus cells removed, then cultureduntil use. Oocytes and embryos were immobilized and perfused with treatmentsolutions at 24 degrees C. Osmotically induced cell volume changes overtime were videotaped followed by image analysis. The Lp values in thepresence of dimethylsulphoxide (DMSO) were 0.77, 0.81, 0.94, 0.86, and 1.10microm/min/atm, and the PDMSO values were 1.85, 2.04, 2.41, 1.95, and1.25x10(-3) cm/min for oocytes, zygotes, 2, 4, and 8-cell embryosrespectively. The Lp values in the presence of DMSO were significantly (P< 0.05) higher than those in the absence of DMSO. Treating the wholeembryo as a single osmotic entity leads to significantly (P < 0.05)elevated PDMSO estimates relative to those based upon measurements ofindividual blastomeres. These data indicate that both Lp and PDMSOestimates are lower when predicted on an individual blastomere basis. Thedata also show that neither Lp nor PDMSO differ among oocytes, zygotes,2-cell and 4-cell embryos. However, the significantly higher Lp and lowerPDMSO of the 8-cell stage support the hypothesis that fundamentalcryobiological differences may require developmental stage- specific embryocryopreservation protocols.