Altering fish embryos with aquaporin-3: an essential step toward successful cryopreservation.
Fish populations are globally threatened by overharvesting and habitat degradation. The ability to bank fish embryos by cryopreservation could be crucial for preserving species diversity, for aquaculture (allowing circannual fish farming), and for managing fish models used in human biomedical research. However, no nonmammalian embryo has ever been successfully cryopreserved. For fish, low membrane permeability prevents cryoprotectants from entering the yolk to prevent cryodamage. Here, we present evidence of a membrane mechanism hindering cryopreservation of fish and propose a novel solution to this obstacle. Zebrafish (Danio rerio) embryos have rectifying membranes that allow water to leave but not to reenter readily. This feature may be an evolutionary trait that allows freshwater embryos to grow in hypoosmotic environments without osmoregulatory organs. However, this trait may also prevent successful fish embryo cryopreservation because both water and cryoprotectants must move into and out of cells. As a solution, we injected zebrafish embryos with mRNA for the aquaporin-3 water channel protein and demonstrated increased membrane permeability to water and to a cryoprotectant. Modeling indicates that sufficient cryoprotectant enters aquaporin-3-expressing zebrafish embryos to allow cryopreservation.
Department of Reproductive Science, Smithsonian Institution, National Zoological Park and Conservation and Research Center, Washington, District of Columbia 20008, USA. hagedornm@nzp.si.edu
M
M
Hagedorn
S L
SL
Lance
D M
DM
Fonseca
F W
FW
Kleinhans
D
D
Artimov
R
R
Fleischer
A T M S
AT
Hoque
M B
MB
Hamilton
B S
BS
Pukazhenthi
D000818:Animals; D051400:Aquaporin 3; D020346:Aquaporins / Q000235:genetics* / Q000494:pharmacology; D002463:Cell Membrane Permeability; D015925:Cryopreservation / Q000662:veterinary*; D003451:Cryoprotective Agents / Q000378:metabolism; D004625:Embryo, Nonmammalian / Q000378:metabolism*; D015870:Gene Expression; D005818:Genetic Engineering; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics; D009994:Osmolar Concentration; D019946:Propylene Glycol / Q000378:metabolism; D012333:RNA, Messenger / Q000008:administration & dosage; D011993:Recombinant Fusion Proteins; D014162:Transfection; D014882:Water-Electrolyte Balance; D015027:Zebrafish / Q000196:embryology*
D020346:Aquaporins; D003451:Cryoprotective Agents; D008164:Luminescent Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D051400:Aquaporin 3; D019946:Propylene Glycol