Abstract
We report a novel method of rapid and quantitative evaluation of the degree of radiation-induced apoptosis in the developing brain, using medaka (Oryzias latipes). We also show the radiation effects on the hisogenesis of medaka brain. Embryos at stage 28 were irradiated with 1-10 Gy of X-rays. Twenty to twenty-four hours after the irradiation, living embryos were stained with a vital dye, acridine orange (AO), for 1-2 h, and whole-mount brains were examined under an epifluorescence microscope. The numbers of AO-stained apoptotic bodies and single nuclei increased in a dose-dependent manner, and AO-stained large apoptotic bodies (rosette-shaped clusters of nuclear fragments) were found in the optic tectum. We used the number of rosette-shaped clusters/tectum as an index of the degree of radiation-induced brain cell death. The results showed that the number of rosette-shaped clusters/tectum in the irradiated embryos exposed to higher than 2 Gy was highly significantly different from that in the nonirradiated control embryos, whereas no difference was detected at 1 Gy. Thus, the threshold dose for brain cell death in medaka embryos is about 1 Gy and this dose is probably comparable to those for rodents and humans. The dead cells disappeared thereafter, and the irradiated embryos continued to develop apparently normally. The grown irradiated (10 Gy) embryos, however, had smaller brains and eyes than the nonirradiated control embryos. At hatching, the irradiated (10 Gy) embryos exhibited histological abnormalities in the brain and retina, although most of them hatched normally and survived. The results show that medaka embryos are useful for studies of the developmental neurotoxic effects of radiation.
