Biol Reprod Lalor Postdoctoral Fellowships -- Application Deadline January 15, 2009
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BOR - Papers in Press, published online ahead of print September 3, 2008.
Biol Reprod 2008, 10.1095/biolreprod.108.069880
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BIOLOGY OF REPRODUCTION 80, 175–183 (2009)
DOI: 10.1095/biolreprod.108.069880
© 2009 by the Society for the Study of Reproduction, Inc.

Artificial Fertilization by Intracytoplasmic Sperm Injection in a Teleost Fish, the Medaka (Oryzias latipes)1

Satoshi Otani , Toshiharu Iwai , Shingo Nakahata  3, Chiharu Sakai , and Masakane Yamashita  2

Laboratory of Reproductive and Developmental Biology, Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810, Japan

ABSTRACT

Intracytoplasmic sperm injection (ICSI) is a technique that has been successfully used for assisting reproduction in mammals. However, this method is still not reliable in nonmammalian species, including teleosts. We succeeded in producing medaka individuals by ICSI with a rate of 13.4% (28 hatched embryos out of 209 eggs fertilized by ICSI), the best value reported so far in teleosts, including zebrafish and Nile tilapia. Although the technique was based on that developed for mammalian eggs, some critical modifications were made to adjust it to the medaka egg, which has a thick and hard envelope (the chorion) and a single sperm entry site (the micropyle). Medaka ICSI was performed by injecting a demembranated spermatozoon into an egg cytoplasm through the micropyle 10–15 sec after egg activation induced by a piezo-actuated vibration, the site and timing of sperm penetration being consistent with those in normal fertilization in medaka. To increase the efficiency of ICSI in medaka, we found that the fertilization by ICSI should precisely mimic the fertilization by insemination with intact sperm, both spatially and temporally. The success rate of ICSI was highly variable in batches of eggs (ranging from 0% to 56%), suggesting that the conditions of eggs are important factors in stabilizing the production of individuals by ICSI. The success in medaka ICSI provides a basis for future research to understand the basic mechanisms in gamete biology of teleosts as well as for development of new technology that can yield valuable applications in fisheries science.

artificial fertilization, assisted reproductive technology, developmental biology, fertilization, gamete biology, ICSI, in vitro fertilization, medaka, sperm injection, teleost fish

FOOTNOTES

1Supported by the Program for Promotion of Basic Research Activities for Innovative Biosciences from the Bio-Oriented Technology Research Advancement Institution to M.Y.

Correspondence: 2FAX: 81 11 706 4456; e-mail: myama{at}sci.hokudai.ac.jp

3 Current address: Division of Tumor and Cellular Biochemistry, Department of Medical Sciences, Faculty of Medicine, University of Miyazaki, Miyazaki 889-1692, Japan.






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Copyright © 2009 by the Society for the Study of Reproduction.