Yasuko Hyodo-Taguchi

Phenotypic rescue of the albino mutation in the medakafish (Oryzias latipes) by a mouse tyrosinase transgene

Mutations of the tyrosinase gene are one common cause of a similar phenotype in all vertebrates, known as albinism. In an attempt to contribute to an understanding of the genetic hierarchy governing the development of pigmentation, we have used a mouse tyrosinase minigene under the control of its 5.2 kb upstream promoter region to rescue two different albino mutations in the medakafish, Oryzias latipes. Around hatching stages an almost perfect phenocopy of the wildtype pigmentation was obtained. Subsequent ectopic melanin overproduction indicated a possible incompatibility of the heterologous mouse promoter for stable expression during the entire ontogenesis. Like in some tyrosinase transgenic mouse lines a strong variegation effect was observed. The transgene-mediated pigmentation phenotype was obtained up to the eighth offspring generation. The phenotypic effects of the tyrosinase transgene in different albino mutant strains places the i3-locus upstream and the b-locus downstream of the tyrosinase locus i1 in the genetic hierarchy leading to wildtype pigmentation.

Short- and long-term effects of low-dose prenatal X-irradiation in mouse cerebral cortex, with special reference to neuronal migration

Abstract To elucidate the short- and long-term effects of ionizing radiation on cell migration in the developing cerebral cortex, we labeled proliferating cells on day 14 of gestation of mice with bromodeoxyuridine (BrdU) followed by a single exposure to 0.1–1 Gy of X-rays. The brains of embryos on day 17 and offspring at 2, 3 and 8 weeks after birth were processed for BrdU immunohistochemistry to trace the movements of BrdU-labeled cells. The location of BrdU-labeled neurons in the cerebral cortex was quantitatively analyzed between irradiated animals and non-irradiated controls. We have demonstrated that the initial migration of BrdU-labeled cells from the matrix cell zone towards the cortical plate during embryonic periods was decelerated when exposed to X-rays of 0.25, 0.5 and 1 Gy on embryonic day 14, and that aberrantly placed neurons in the cerebral neocortex were noted in younger animals that were irradiated prenatally, whereas such derangement was less pronounced in mature animals. These observations suggest that some modification process might have occurred during the postnatal period.

Effects of tritiated water on germ cells in medaka embryos

Embryos of medaka, Oryzias latipes, were exposed to tritiated water and 137Cs gamma rays continuously from the one-cell stage until hatching (10 days at 26 degrees C). Germ cells in the gonads of newly hatched fry were counted in histological sections and compared with controls. The accumulated dose for 50% survival of germ cells was 195 rad for tritium beta rays and 350 rad for 137Cs gamma rays. Female progeny were produced using Yamamotos method. The 50% survival doses for female germ cells treated in a manner similar to that described above were 140 rad for beta rays and 305 rad for gamma rays. When embryos of medaka were irradiated with gamma rays below an accumulated dose of 475 rad or treated with tritiated water at a concentration of 0.2 mCi/ml or lower, the dose response of the germ cells showed an exponential relationship. It appeared that there was no threshold or significant dose-rate effect for either beta or gamma rays on germ cell survival, and that tritium beta rays were more effective than 137Cs gamma rays in germ cell killing.

RBE of fast neutrons for inhibition of hatchability in fish embryos irradiated at different developmental stages.

Embryos of the fish, Oryzias latipes at four different developmental stages were irradiated with various doses of 2 MeV fast neutrons from a Van de Graaff or 200 kVp x-rays. The hatchability, morta...

Heritable malformations in the progeny of the male medaka (Oryzias latipes) irradiated with X-rays

Heritable malformations were examined in the progeny of X-irradiated male medaka (Oryzias latipes) by three-generation crosses. Two X-irradiated male fish were pair-mated with non-irradiated females to produce F1 founders, and each F1 fish was pair-mated with a non-irradiated fish to produce F2 progeny. For detection of recessive mutations, pair-matings between F2 siblings were performed for each F1 family. Morphogenesis of the embryos of each generation was observed using a stereomicroscope, throughout the entire period of embryonic development. In the F1 embryos, the frequencies of dominant lethals and malformations were increased by the X-irradiation. Two out of 30 F1 pairs produced a number of malformed and lethal F2 embryos, indicating inheritance of high rates of the dominant lethals in the two F1 families. Moreover, F2 sib-pairs offspring of which exibited high rates of dominant lethals were found in 10 out of 28 F1 families. Recessive lethal mutations, which were associated with a particular phenotype, were found in 2 out of the 28 F1 families. These results indicate that the heritable malformations induced by X-irradiation can be studied in the medaka.

Cranial nerves and brain fiber systems of the medaka fry as observed by a whole-mount staining method

The cranial nerves and the brain fiber systems of the medaka (Oryzias latipes) fry are revealed by a whole-mount staining method. Newly hatched fry of an albino strain of the medaka were fixed, partially digested with trypsin, treated in 1% Triton X-100, and finally immunohistochemically stained using anti-neurofilament protein (70K+160K+210K) antibodies. Since both head skin and eyes were colorless in the albino fish, the three-dimensional distribution of of nerve fibers in the brain could be readily observed in whole specimens without interference of pigment cells. All cranial nerves and main fiber systems in the adult fish were differentiated in the fry brain. Using this method, the distribution of nerves to the ocular muscles and the periorbital pit organs was shown.

Purification and partial characterization of the muscle LDH-A4 and -B4 isozymes and the respective subunits of the fish, Oryzias latipes

Abstract 1. 1. The muscle LDH-A 4 and -B 4 isozymes of the inbred and outbred fish of a Japanese cyprinodont, Oryzias latipes , were purified and some of the protein chemical as well as enzymological characteristics were examined. 2. 2. The molecular weights of the A and B subunits were about 36.0 kD and 36.5 kD, respectively. There was a marked similarity in amino acid composition between the corresponding subunits of this fish and the salmon, Oncorhynchus tshawytscha . 3. 3. The optimal pHs of both A 4 and B 4 isozymes were almost the same, being about 6.5 for pyruvate reduction and about 10.0 for lactate oxidation. 4. 4. The B 4 enzyme was markedly more heat-stable and of higher affinity to the substrates in the reciprocal reactions than the A 4 enzyme.

Effects of Tritiated Water on Germ Cells in Medaka: II. Diminished Reproductive Capacity Following Embryonic Exposure

To determine the effects of early radiation-induced germ cell loss on lifetime reproduction, the fecundity and fertility of fish exposed to chronic irradiation as embryos were investigated. Embryos of the medaka, Oryzias latipes, were exposed to tritiated water (0.05-2.0 mCi/ml) and137 Cs γ rays (6.1-254 rad/day) continuously from the morula stage until hatching (10 days at 26°C). Newly-hatched fry were removed from the radioactive environment. At 4-8 months after hatching, pairs consisting of irradiated males and nonirradiated females or the reverse were mated. When irradiated females were mated with unirradiated males, both the total number of ovipositions and the number of eggs per fish decreased with increasing dose. However, most eggs laid were fertilized and hatched normally in all groups. Tritium β rays were more effective than137 Cs γ rays in reducing the fecundity of the irradiated females. In pairs where the male had been irradiated, the total number of ovipositions did not decrease, but the num...

Mortality response of goldfish after exposure to fast neutrons

ETOH, H., HYODO-TAGUCHI, Y., AND MARUYAMA, T. Mortality Response of Goldfish after Exposure to Fast Neutrons. Radiat. Res. 59, 513-520 (1974). The mortality response of goldfish to whole-body irradiation with 2 MeV fast neutrons from a Van de Graaff accelerator was studied and compared with the effects of 200 kVp X-rays. Although the dose ranges leading to equivalent injury were different with the two types of radiation, the dose vs survival time relationship, and the distribution pattern of time of death after irradiation with fast neutrons were similar to those observed after X-irradiation. Mean lethal dose was used to determine the relative biological effectiveness of neutrons. The LDs5(30) was 375 rad for neutrons and 1238 rad for X-rays, from which the RBE of neutrons was calculated to be 3.4. The histological changes in hematopoietic tissue (body kidney) and intenstine to 400 or 1000 rad of neutrons were examined. In fish irradiated with 400 or 1000 rad of neutrons, the marked reduction in the number of hematopoietic cells occurred 5-10 postirradiation days. Severe lesions in the intestine were observed only in 1000 rad-irradiated fish at 10 days after irradiation. These results were compared with previous X-irradiation results in fish.

MEDAKA, A USEFUL EXPERIMENTAL SYSTEM FOR CHEMICAL AND ENVIRONMENTAL CARCINOGENESIS

Publisher Summary This chapter describes the application of the medaka in chemical and radiation carcinogenesis. The medaka, Oryzias latipes, is a convenient system to study chemical and environmental carcinogenesis at molecular, cellular, and tissue levels. It is economical and easy to breed. Chemicals can be administered as solution added to the water in aquaria or be injected into the fish body. The fish is sensitive to water-soluble carcinogens including diethylnitrosamine, methylazoxymethanol (MAM) acetate, and N-Methyl-N´-nitro-N-nitrosoguanidine. The presence of cytochrome P-450 system indicates a possibility to use this fish as a test system for other carcinogens that need metabolic activation. Tumor formation can frequently be detected from the body surface as experienced in melanomas and hepatomas. The small size of the fish permits routine serial sectioning for histological examination.