Ethical Euthanasia and Short-Term Anesthesia of the Chick Embryo

Fertilized chicken eggs are employed as an alternative to mammalian models. The chorioallantoic membrane (CAM) of the chick embryo is widely used for examination of angiogenesis, xenotransplants and for virus production. Unfortunately, it is mostly not taken into account that the chick embryo's ability to experience pain starts to develop at day 7 of incubation. In our view, this model is only in accordance with the 3R principles if an appropriate anesthesia of the chick embryo in potentially painful procedures is provided. Although many experimental approaches are performed on the non-innervated CAM, the euthanasia of the embryo strongly requires a more humane technique than the commonly used methods of freezing at -20 degrees C, decapitation or in ovo fixation with paraformaldehyde without prior anesthesia. However, protocols describing feasible and ethical methods for anesthesia and euthanasia of avian embryos are currently not available. Therefore, we established an easy and reliable method for the euthanasia and short-term anesthesia of the chick embryo.