title: Sensitivity to chilling of medaka (Oryzias latipes) embryos at various developmental stages.
abstract: As an essential step toward cryopreservation of fish embryos, we examined the chilling sensitivity of medaka (Oryzias latipes) embryos at various developmental stages. Embryos at the 2-4 cell, 8-16 cell, morula, blastula, and early gastrula stages were suspended in Hanks solution. They were chilled to various temperatures (usually 0 degrees C), kept for various periods (usually 20 min), then cultured for up to 14 d to determine survival (assessed by the ability to hatch). Embryos at the 2-4 cell stage were the most sensitive to chilling to 0 degrees C, but sensitivity decreased as development proceeded. The survival rate of 2-4 cell embryos was affected after 2 min of chilling at 0 degrees C; although the rate decreased gradually as the duration of chilling increased, 38% of them still survived after 40 min of chilling. Embryos at the 2-4 cell stage were sensitive to chilling at 0 or -5 degrees C, but much less sensitive at 5 or 10 degrees C. The survival rate of 2-4 cell embryos subjected to repeated rapid cooling and warming was similar to that of those kept chilled. When early gastrula embryos were preserved at 0 or 5 degrees C, the hatching rate did not decrease after 12 and 24h of chilling, respectively, but then decreased gradually as storage was prolonged; however, 3-10% of the embryos hatched even after storage for 10 d. In conclusion, although later-stage medaka embryos would be suitable for cryopreservation (from the perspective of chilling sensitivity), chilling injury may not be serious in earlier stage embryos.
authors.affiliation: Laboratory of Animal Science, College of Agriculture, Kochi University, Nankoku, Kochi 783-8502, Japan.
authors.forename: Delgado M
authors.initials: DM
authors.lastname: Valdez
authors.forename: Akira
authors.initials: A
authors.lastname: Miyamoto
authors.forename: Takao
authors.initials: T
authors.lastname: Hara
authors.forename: Keisuke
authors.initials: K
authors.lastname: Edashige
authors.forename: Magosaburo
authors.initials: M
authors.lastname: Kasai
mesh_terms: D000818:Animals; D003080:Cold Temperature; D046149:Embryo Culture Techniques / Q000662:veterinary; D047108:Embryonic Development; D005775:Gastrula; D009990:Oryzias / Q000196:embryology*; D013997:Time Factors; D014021:Tissue Preservation / Q000662:veterinary*