Effects of methanol and developmental arrest on chilling injury in zebrafish (Danio rerio) embryos.
Stage-dependent chilling sensitivity has been reported for many species of fish embryos. Most of these studies reveal that developmental stages beyond 50% epiboly are less sensitive to chilling, but the chilling sensitivity accelerates rapidly at subzero temperatures. In this study, the effects of methanol and developmental arrest on chilling injury were studied using zebrafish (Danio rerio) embryos at 64-cell, 50% epiboly, 6-somite, prim-6 and long-bud stages. Embryos were exposed to methanol or anoxic conditions before they were cooled to 0 or -5 degrees C with slow (1 degrees C/min), medium (30 degrees C/min) or fast ( approximately 300 degrees C/min) cooling rates and were held at these temperatures for different time periods. Embryo survival was evaluated in terms of the percentage of treated embryos with normal developmental appearance after 3-day culture. Experiments on the effect of methanol on chilling sensitivity of the embryos showed that the addition of methanol to embryo medium increased embryo survival significantly at all developmental stages and under all cooling conditions. Higher concentration of methanol treatment generally improved embryo survival when embryos were cooled at a fast cooling rate of 300 degrees C/min. Experiments on the effect of developmental arrest on chilling sensitivity of embryos showed that embryos at 50% epiboly and prim-6 stages underwent developmental arrest almost immediately after 15 min oxygen deprivation. After 4h in anoxia, the survival rates of the embryos were not significantly different from their respective aerobic controls. Anoxia and developmental arrest had no effect on the chilling sensitivity of zebrafish embryos.
Zhang
Tiantian
T
Luton Institute of Research in the Applied Natural Sciences, University of Luton, 2 Adelaide Street, Bedfordshire LU1 5DU, UK. tiantian.zhang@luton.ac.uk
Liu
Xiang-Hong
XH
Rawson
David M
DM
D000818:Animals; D003080:Cold Temperature; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary; D004305:Dose-Response Relationship, Drug; D004625:Embryo, Nonmammalian / Q000187:drug effects*; D047108:Embryonic Development; D005260:Female; D000432:Methanol / Q000494:pharmacology*; D009584:Nitrogen / Q000737:chemistry; D010100:Oxygen / Q000737:chemistry; D011247:Pregnancy; D013997:Time Factors; D015027:Zebrafish / Q000196:embryology* / Q000254:growth & development
D009584:Nitrogen; D010100:Oxygen; D000432:Methanol