Abstract
It is important to examine the neurocytotoxic effects of radiation on the developing central nervous system (CNS) because of its high sensitiveness to radiation. Medaka is an ideal model for studying the effects of developing CNS of vertebrates, because the transparency of their eggs and embryos makes it possible to detect morphological abnormalities in the CNS easily using a conventional stereomicroscope. Wild-type and p53(-/-) medaka embryos at developmental stage 28, when neural cells proliferate rapidly especially in the optic tectum (OT), were irradiated with 10 Gy gamma-rays which does not affect to hathability. Time-lapse changes of radiation-induced apoptosis were examined in whole-mount specimens using an acridine orange (AO) assay. By AO assays, apoptotic cells in irradiated wild-type OT appeared as AO-positive small single nuclei at 3h after the irradiation. At 8-10 h after the irradiation, we found two types of AO-stained cells, namely,a single single nuclei and rosette-shaped clusters. At 15 h after the irradiation, the AO-positive clusters became bigger and more obvious, and the number of them gradually decreased at 35 h after the irradiation. On the other hand, in p53(-/-) irradiated OT, AO-positive single nuclei started to appear 3-5 h later than that of wild-type embryos, and the size of AO-positive rosette-shaped clusters was apparently smaller than that of wild-type.
In order to prevent the diffusion of damaging degradation products into the surrounding tissue, apoptotic neuronal cells are quickly phagocytosed by the microglia in the brain. Here, we investigated how microglia act to phagocytose rosette-shaped clusters of apoptotic cells in wild-type and p53(-/-) irradiated brain at 24 h after the irradiation. The gene expressions of apolipoprotein-E (Apo-E) protein, which is specifically expressed in micriglia, determined by whole-mount in situ hybridization revealed a high-level of transcripts in the retinal ganglion cell layer and in the OT, where rosette-shaped clusters of apoptosis were observed on histological sections in both type of irradiated OT. Almost no differences were observed in expression patterns of ApoE in both type of irradiated embryos.
