Biol Reprod
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BOR - Papers in Press, published online ahead of print October 23, 2002.
Biol Reprod 2002, 10.1095/biolreprod.101.002394
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BIOLOGY OF REPRODUCTION 68, 87–94 (2003)
DOI: 10.1095/biolreprod.101.002394
© 2003 by the Society for the Study of Reproduction, Inc.

Embryo

Artificial Expression of Aquaporin-3 Improves the Survival of Mouse Oocytes after Cryopreservation1

Keisuke Edashige2,a, Yohei Yamajia, F.W. Kleinhansb, and Magosaburo Kasaia

a Laboratory of Animal Science, College of Agriculture, Kochi University, Nankoku, Kochi 783-8502, Japan b Department of Physics, Indianapolis University and Purdue University Indianapolis, Indianapolis, Indiana 46202

Successful cryopreservation of mammalian cells requires rapid transport of water and cryoprotective solutes across the plasma membrane. Aquaporin-3 is known as a water/solute channel that can transport water and neutral solutes such as glycerol. In this study we examined whether artificial expression of aquaporin-3 in mouse oocytes can improve water and glycerol permeability and oocyte survival after cryopreservation. Immature mouse oocytes were injected with aquaporin-3 cRNA and were cultured for 12 h. Then the hydraulic conductivity (LP) and glycerol permeability (PGLY) of matured oocytes were determined from the relative volume changes in 10% glycerol in PB1 medium at 25°C. Mean ± SD values of LP and PGLY of cRNA-injected oocytes (3.09 ± 1.22 µm min-1 atm-1 and 3.69 ± 1.47 x 10-3 cm/min, respectively; numbers of oocytes = 25) were significantly higher than those of noninjected oocytes (0.83 ± 0.02 µm min-1 atm-1 and 0.07 ± 0.02 x 10-3 cm/min, respectively; n = 13) and water-injected oocytes (0.87 ± 0.10 µm min-1 atm-1 and 0.08 ± 0.02 x 10-3 cm/min, respectively; n = 20). After cryopreservation in a glycerol-based solution, 74% of cRNA-injected oocytes (n = 27) survived as assessed by their morphological appearance, whereas none of the water-injected oocytes survived (n = 10). When cRNA-injected oocytes that survived cryopreservation were inseminated in vitro, the penetration rate was 40% (n = 48) and the cleavage rate was 31% (n = 70), showing that oocytes retain their ability to be fertilized. This is the first report to show that artificial expression of a water/solute channel in a cell improves its survival after cryopreservation. This approach may enable cryopreservation of cells that have been difficult to cryopreserve.

1 This study was supported by grants-in-aid for scientific research from the Japanese Society for the Promotion of Science (RFTF Program); the Ministry of Education, Sports, Culture, and Science of Japan; and the Inamori Foundation.

2 Correspondence. FAX: 81 88 864 5219; keisuke{at}cc.kochi-u.ac.jp






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Copyright © 2003 by the Society for the Study of Reproduction.