Office of Research Integrity
Use of Zebrafish Fry
The MUSC IACUC must evaluate all experiments on zebrafish fry that are allowed to develop greater than 120 hrs following fertilization. Any experiments performed on zebrafish fry that are older than 120 hours must be described in an approved IACUC protocol, and the number of animals used in the experiments must be tracked and remain consistent with the number approved in the protocol. Furthermore, any experiments on juveniles at less than 120 hrs from fertilization must also be described in an IACUC-approved protocol provided that those animals are expected to survive to greater than 120 hrs in age. Zebrafish embryos that are manipulated before the 120-hour limit and are euthanized before that time are not regulated by the IACUC.
The molecular development of the zebrafish embryo proceeds similarly to that of chick and mouse, but the environment in which this happens is completely different. The zebrafish embryo does not develop internally in the mother and there is no calcified shell to protect the embryo because this would hinder fertilization. Zebrafish females spawn unfertilized eggs, which are quickly fertilized. These eggs sink to the bottom of the river/tank. Even though a one-cell stage embryo can survive (albeit poorly) outside of the chorion membrane if it is carefully removed, viability without a chorion membrane is not synonymous to chick embryo survivability outside of its eggshell. Based on these differences, the IACUC considered a time point that is more related to when chick and mouse embryos make their transition to viability. In determining that it is appropriate to regulate experiments on zebrafish fry only after 120 hrs, the IACUC considered the following developmental traits of the species:
- The jaw is not developed until between day four and five.
- The swim bladders do not fully inflate until between day four and five.
- The yolk is not fully utilized and absorbed until day five to seven. Even though the embryos can eat by day five, they can subsist without food until day seven or eight.
- Vertebrae begin to calcify on day seven, and they are all not fully calcified until between days 16 to 21 (Du et. al., 2001, Dev Bio 238:239).
- The recommendation and guidelines provided to all IACUCs related to zebrafish fry (Borski & Hodson, 2003, ILAR J. 44:286).