Zebrafish as a “Biosensor”? Effects of Ionizing Radiation and Amifostine on Embryonic Viability and Development

Abstract

The zebrafish (Danio rerio) has emerged as a popular vertebrate model system for cancer and treatment-related research. Benefits include ease of care, rapid development, optical clarity of embryos, which allows visualization of major organ systems, and opportunities for genetic manipulation. However, specific parameters of radiation sensitivity have not been systematically documented. We investigated the effects of radiation and a radiomodifier on zebrafish viability and embryonic development. Embryos were exposed to ;-radiation (5, 10, or 20 Gy) at sequential times postfertilization and serially assessed for viability and morphologic abnormalities. As expected, lethality and morphologic perturbations were more pronounced earlier in embryogenesis and with higher radiation doses and were partially reversed by amifostine. The effects of radiation and concurrent treatment with amifostine on the developmental organization of the eye and brain were striking. Radiation resulted in hypocellularity and disorganization of the cellular layers of the retina, effects partially reversed by amifostine, as well as lens opacification. Radiation strikingly reduced the volume of brain, but the volume loss was substantially blocked by amifostine. Increased terminal deoxyribonucleotidyl transferase-mediated dUTP nick end labeling signal was noted in both the irradiated eye and brain, but reduced by amifostine. Finally, irradiating embryos resulted in caspase activation detectable in 96-well microplates, which was proportional to the number of embryos and radiation dose; the degree of activation was markedly reduced by amifostine. These results together suggest the power and versatility of the zebrafish in assessing the effects of radiation and radiomodifiers on organ and tissue development. (Cancer Res 2006; 66(16): 8172-81)

Temozolomide-Mediated Radiosensitization of Human Glioma Cells in a Zebrafish Embryonic System

“…Finally, rapid embryonic development facilitates investigations of effects on development. Recent reports have described the effects of radiation on viability and development in early zebrafish embryos, confirming anatomic effects similar to that described for humans (15,16) and showed the usefulness of the model in delineating the toxicity of novel agents (17). The growth and interactions with the host tissues of human melanoma cells transplanted into zebrafish has been described (18)(19)(20).…”

“…We have previously established parameters for testing the effects of IR on zebrafish embryos (15). This allowed us to determine that the ideal stage to test the relative effects of IR on transplanted tumor cells would be at 1 dpf.…”

“…Zebrafish were raised as previously described (15) and all procedures, including embryo collection and handling, were done in accordance with accepted standard operating procedures (21) approved by the Institutional Animal Care and Use Committee at the University of Pennsylvania School of Medicine. Staging of the zebrafish embryos and larvae were performed as described by Kimmel et al (22).…”

“…Acquisition of images was performed as previously described (15). Following transplantation of the U251-RFP cells, embryos were serially examined under a Â100 PlanNeofluor objective mounted on a Nikon TE-200 microscope equipped with epifluorescence optics.…”

“…Treatment (irradiation and exposure to temozolomide) of embryos was performed according to protocols approved by the Department of Environmental Health and Radiation Safety at the University of Pennsylvania. Embryos were either irradiated at 10 Gy or mock-irradiated as previously described (15). Embryos at 1 dpf were exposed to single fractions of 640 kVp g-irradiation at room temperature using a J.L.…”