title: A trial to cryopreserve immature medaka (Oryzias latipes) oocytes after enhancing their permeability by exogenous expression of aquaporin 3.
abstract: Fish oocytes have not been cryopreserved successfully, probably because it is difficult to prevent intracellular ice from forming. Previously, we have shown in medaka that immature oocytes are more suitable for cryopreservation than mature oocytes or embryos, in terms of permeability. We have also shown in immature medaka oocytes that the exogenous expression of aquaporin 3 (AQP3), a water/cryoprotectant channel, promotes the movement of water and cryoprotectants through the plasma membrane. In the present study, we attempted to cryopreserve immature medaka oocytes expressing AQP3. We first examined effects of hypertonic stress and the chemical toxicity of cryoprotectants on the survival of the AQP3-expressing oocytes. Exposure to hypertonic solutions containing sucrose decreased the survival of oocytes, but the expression of AQP3 did not affect sensitivity to hypertonic stress. Also, AQP3 expression did not markedly increase sensitivity to the toxicity of cryoprotectants. Of the four cryoprotectants tested, propylene glycol was the least toxic. Using a propylene glycol-based solution, therefore, we tried to cryopreserve immature oocytes by vitrification. During cooling with liquid nitrogen, all intact oocytes became opaque, but many AQP3-expressing oocytes remained transparent. This indicates that the expression of AQP3 is effective in preventing intracellular ice from forming during cooling. During warming, however, all the AQP3-expressing oocytes became opaque, indicating that intracellular ice formed. Therefore, the dehydration and permeation by propylene glycol were still insufficient. Further studies are necessary to realize the cryopreservation of fish oocytes.
authors: Laboratory of Animal Science, College of Agriculture, Kochi University, Kochi 783-8502, Japan.
authors: Delgado M
authors: DM
authors: Valdez
authors: Ryoma
authors: R
authors: Tsuchiya
authors: Shinsuke
authors: S
authors: Seki
authors: Naoya
authors: N
authors: Saida
authors: Saori
authors: S
authors: Niimi
authors: Chihiro
authors: C
authors: Koshimoto
authors: Kazutsugu
authors: K
authors: Matsukawa
authors: Magosaburo
authors: M
authors: Kasai
authors: Keisuke
authors: K
authors: Edashige
mesh_terms: D000818:Animals; D051400:Aquaporin 3 / Q000235:genetics* / Q000378:metabolism; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology; D019855:Ethylene Glycol / Q000494:pharmacology; D006982:Hypertonic Solutions / Q000494:pharmacology; D009865:Oocytes / Q000378:metabolism*; D009990:Oryzias; D010539:Permeability; D019946:Propylene Glycol / Q000494:pharmacology
chemical_list: D003451:Cryoprotective Agents; D006982:Hypertonic Solutions; D051400:Aquaporin 3; D019946:Propylene Glycol; D019855:Ethylene Glycol