Intracytoplasmic Sperm Injection (ICSI) in Zebrafish Eggs

Poleo, G. A. and T. R. Tiersch, Aquaculture Research Station, Louisiana Agricultural Experimental Station, Agricultural Center, Louisiana State University, Baton Rouge, LA 70820

Direct injection of sperm into the cytoplasm of oocytes was first reported in sea urchin and amphibians 30 years ago. More recently, this technique has been found to be useful in humans to overcome some of the problems related to male infertility. In domestic animals intracytoplasmic sperm injection (ICSI) as become an area of intensive research due to its potential for stock improvement. This technique also has been recognized as a valuable method for the preservation of endangered species with difficulties to reproduce in captivity. However, aside from the original studies of sperm injection in echinoderms, other work has not been reported in aquatic organisms. Using zebrafish Danio rerio as a model we were able to inject single sperm into eggs and obtain phenotypically normal larvae. As observed in other organisms, we also found that injection of sperm can cause lethal damage. However, unlike mammals, in which inviable embryos only undergo early development, zebrafish developed to the post-gastrula stage despite altered phenotypes. From 74 eggs injected with single sperm cells, five embryos developed to the gastrula stage, two developed abnormally past gastrulation and one developed normally and hatched. Measurement of the DNA content of the normal fish, by flow cytometry, showed it to be diploid. However, with this experiment we were not able to distinguish the contribution of male and female gametes. To overcome this we have begun use of a zebrafish line possessing a recessive phenotype for pigment (gold) and fin shape (long fins) as an egg donor, together with sperm from zebrafish carrying dominant phenotype for pigment (wild type).  These preliminary results show that ICSI is possible in fish and opens the door to new areas of research, ranging from basic reproductive biology to the production of hybrids with commercial value that otherwise would not be possible. Another potential application is the use of sperm preserved by methods other than freezing, which would reduce storage costs.

Copyright © 1999
Texas Chapter of the American Fisheries Society
Last modified: 01/07/09 06:45:17 AM