• The Sea:JOURNAL OF THE KOREAN SOCIETY OF OCEANOGRAPHY
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• v.13 no.2
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• pp.140-146
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• 2008

Marine ecotoxicological standard method using fish larvae was established with the standard test species of Oryzias latipes(Japanese Medaka) and Paralichthys olivaceus(flounder) and with the 7 day $LC_{50}$ as endpoint. Test method referred to the USEPA(1994) with the replacement of test species found in the Korean water. Standard test species were selected in terms of the species supply and ecological importance in Korean waters. Japanese medaka can be reared with small tanks in the lab and has wide tolerance on salinity, and flounder eggs can be easily obtained from commercial fish hatcheries. General conditions for larval fish toxicity test are as follows. The possible salinity ranges for toxicity test were $0{\sim}35\;psu$ for medaka and >20 psu for flounder. Test type was designated as static non-renewal test if the dissolved oxygen in the test chamber does not fall below 4.0 mg/L. Ages of test species were selected as 7 days after hatched for medaka(about 5 mm TL) and 25 days for flounder(about 10 mm TL) because of the low natural mortality after these periods. Test can be accepted when the survival rates are over 80% in control. Also, species sensitivity on standard reference materials(copper, cadmium or zinc) must be provided with the toxicity test results.

• Journal of Animal Reproduction and Biotechnology
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• v.39 no.1
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• pp.19-30
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• 2024

Background: Although an understanding of the proliferation and differentiation of fish female germline stem cells (GSCs) is very important, an appropriate threedimensional (3D) research model to study them is not well established. As a part of the development of stable 3D culture system for fish female GSCs, we conducted this study to establish a 3D aggregate culture system of ovarian cells in marine medaka, Oryzias dancena. Methods: Ovarian cells were separated by Percoll density gradient centrifugation and two different cell populations were cultured in suspension to form ovarian cell aggregates to find suitable cell populations for its formation. Ovarian cell aggregates formed from different cell populations were evaluated by histology and gene expression analyses. To evaluate the media supplements, ovarian cell aggregate culture was performed under different media conditions, and the morphology, viability, size, gene expression, histology, and E2 secretion of ovarian cell aggregates were analyzed. Results: Ovarian cell aggregates were able to be formed well under specific culture conditions that used ultra-low attachment 96 well plate, complete mESM2, and the cell populations from top to 50% layers after separation of ovarian cells. Moreover, they were able to maintain minimal ovarian function such as germ cell maintenance and E2 synthesis for a short period. Conclusions: We established basic conditions for the culture of O. dancena ovarian cell aggregates. Additional efforts will be required to further optimize the culture conditions so that the ovarian cell aggregates can retain the improved ovarian functions for a longer period of time.

• Korean Journal of Ichthyology
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• v.28 no.4
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• pp.215-222
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• 2016

Triploidy was induced in the marine medaka, Oryzias dancena by cold shock treatment ($0^{\circ}C$) of fertilized eggs for 30, 45, or 60 min, applied two minutes after fertilization. The triploid genotype was induced by each of the thermal shock regimes tested. The best result was obtained when the eggs were treated for 45 min, which induced triploidy in all the resulting fish. Triploidy was confirmed using chromosomal and flow cytometer analyses, and erythrocyte measurements. The surface areas and volumes of the erythrocytes of triploid fish were significantly larger than those of diploid fish, and their chromosome number (3N=72) was 1.5 times greater that for the diploids (2N=48). Based on a flow cytometer analysis, the triploid fish had approximately 1.5 times the cellular DNA content (2.40 pg/cell) of the diploid specimens (1.61 pg/cell). Data from this study provide the basis for the development of unique models for studying reproductive confinement in transgenic fish.

• Molecular & Cellular Toxicology
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• v.4 no.4
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• pp.293-299
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• 2008

To screen the differentially expressed genes in cadmuim-exposed marine medaka fish (Oryzias javanicus), a candidate marine test fish for ecological toxicity, the differential display polymerase chain reaction (DD-PCR) was carried out, since the genome-wide gene expression data are not available in this fish species yet. A total of 35 clones were isolated from cadmium-exposed fish and their nucleotide sequences were analyzed. The differentially expressed gene candidates were categorized to response to stimulus (3); ion binding (3); DNA binding (1); protein binding (6); carbohydrate binding (1); metabolic process (4); biological regulation (3); cellular process (2); protein synthesis (2); catalytic activity (2); sense of sight (1); immune (1); neurohormone (1); signaling activity (1); electron carrier activity (1) and others (3). For real-time quantitative RT-PCR, we selected catalase, glucose-6-phosphate dehydrogenase, heat shock protein 70, and metallothionein and confirmed that cadmium exposure enhanced induction of these four genes.

• Molecular & Cellular Toxicology
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• v.3 no.sup4
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• pp.34.1-34.1
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• 2007

• Korean Journal of Environmental Biology
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• v.26 no.3
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• pp.220-225
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• 2008

In this study, alterations in whole proteome expression patterns in the head of Japanese medaka (Oryzias latipes) was investigated following acute or chronic exposure to benzo(a)pyrene (BaP) (25 ${\mu}g$ L$^{-1}$) for 48 hrs and 15 days, respectively. The results showed that 9 and 6 protein spots were statistically different, relative to controls, in response to acute and chronic BaP exposure, respectively. In the acute exposure group, 5 spots were up regulated and 4 spots were downregulated, while in the chronic exposure group, 4 spots were upregulated and 2 spots were downregulated. Three of these spots were common to both the acute and chronic BaP exposure groups and were identified using LC-MS/MS followed by database searching. These 3 spots were found to be associated with structural proteins belonging to the actin and keratin families. These data suggest that acute and chronic exposures to BaP may affect tissue morphology in the head of Japanese medaka.

• Korean Journal of Ichthyology
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• v.21 no.3
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• pp.141-148
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• 2009

Sex differentiation and gonad development were investigated in a marine medaka species, Oryzias dancena (Beloniformes; Teleostei). The average time to hatch was 11 days post-fertilization (dpf) at $25^{\circ}C$. Primordial germ cell (PGC) was first observed at 5 dpf and migrated to presumptive gonadal area between the gut and pronephric duct at 9 dpf. Male and female gonads were morphologically differentiated at 12 days post-hatching (dph). Early oocytes at perinucleolus stage as well as the formation of spermatid and efferent duct were observed at 28 dph. At 6 weeks of age, the ovary exhibited yolk granulation in many oocytes, while testis possessed a considerable number of spermatogonia and spermatids. The first ovulation was observed in 9-week-old females, and at the same age, males contained fully-matured spermatozoa. Data obtained in this study indicate that the gonad differentiation of O. dancena is the typical type of differentiated gonochorism.

• Korean Journal of Ichthyology
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• v.21 no.4
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• pp.227-238
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• 2009

The egg development and morphological changes of larvae, juveniles and adults of Oryzias dancena were observed. Fertilized eggs were incubated at $25{\pm}1^{\circ}C$; the process of embryonic development was observed by light microscopy and based on diagnostic features of the developing embryos. The average time to hatch was 11 days after fertilization. The hatched larvae averaged $4.40{\pm}0.24mm$ in total length (TL). The yolk sacs of the larvae were almost absorbed at 3 days after hatching and $4.55{\pm}0.23mm$ TL. At 21 days after hatching, the larvae were $7.23{\pm}0.73mm$ TL and had reached the juvenile stage. First ovulation was about 9 weeks after hatching and at $22.58{\pm}2.73mm$ TL.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.43 no.5
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• pp.462-473
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• 2010

Inductions of hybrids and reciprocal hybrids between Oryzias dancena and O. javanicus (ODJ and OJD) were conducted and backcross hybrids between female O. dancena and male ODJ were also produced for biological and cytogenetic analysis. Embryonic development of ODJ and OJD were compared with those of their parents. Developmental time was fastest in O. dancena and ODJ, followed by O. javanicus and OJD. Oryzias dancena hatched 11 days (d) after fertilization, ODJ at 13 d, O. javanicus at 14 d and OJD at 15 d. The abnormality of external morphology rate in ODJ was 10.6%; however, OJD showed a high degree of abnormality, over 90%. The proportion of males was 90.0% and 31.3% for ODJ and OJD, respectively. Cytogenetic analysis was conducted to obtain basic information for genetic identification of O. dancena, O. javanicus and their hybrids. The karyotypes of all experimental groups showed 2n=48 chromosomes and the fundamental number (FN) was 48. The first pair carried secondary constrictions near the centromeric regions. Erythrocyte area and volume were $9.8\;{\pm}\;0.5\;{\mu}m^2$ and $18.2\;{\pm}\;1.0\;{\mu}m^3$, respectively, for O. dancena, $8.3\;{\pm}\;0.5\;{\mu}m^2$ and $15.8\;{\pm}\;1.5\;{\mu}m^3$ in O. javanicus, and $18.3\;{\pm}\;0.5\;{\mu}m^2$ and $15.7\;{\pm}\;1.3\;{\mu}m^3$ in ODJ. Erythrocyte area and volume in ODJ were similar to those of O. javanicus. In backcross hybrids between female O. dancena and male ODJ, all embryos failed to develop and died in the late gastrula stage.

• Molecules and Cells
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• v.23 no.3
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• pp.287-303
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• 2007

Expressed sequence tags (ESTs) and differentially expressed cDNAs from the self-fertilizing fish, Kryptolebias marmoratus were mined to develop alternative biomarkers for endocrine-disrupting chemicals (EDCs). 1,577 K. marmoratus cDNA clones were randomly sequenced from the 5'-end. These clones corresponded to 1,518 and 1,519 genes in medaka dbEST and zebrafish dbEST, respectively. Of the matched genes, 197 and 115 genes obtained Unigene IDs in medaka dbEST and zebrafish dbEST, respectively. Many of the annotated genes are potential biomarkers for environmental stresses. In a differential display reverse transcriptase-polymerase chain reaction (DD RT-PCR) study, 56 differential expressed genes were obtained from fish liver exposed to bisphenol A. Of these, 16 genes were identified after BLAST search to GenBank, and the annotated genes were mainly involved in catalytic activity and binding. The expression patterns of these 16 genes were validated by real-time RT-PCR of liver tissue from fish exposed to bisphenol A. Our findings suggest that expression of these 16 genes is modulated by endocrine disrupting chemicals, and therefore that they are potential biomarkers for environmental stress including EDCs exposure.