Gonadal development and reproductive cycle of Aplysia kurodai inhabiting the coastal waters of Jeju Island, Korea were investigated based on monthly changes of gonadosomatic index, gametogenesis, and developmental phases of ovotestis. A. kurodai was simultaneous hermaphrodite; the ovotestis generally embedded in the posterior dorsal surface of the brownish digestive gland. The ovotestis is composed of a large number of follicles, and both oocytes and sperm are produced in the same follicles. In the sampling periods, the adult A. kurodai population have characteristic of seasonal pattern present during only 10 months. The reproductive cycle can be grouped into the following successive stages in the ovary: inactive (December to February), active (December to April), mature and spawning (April to September). The gonadal development of A. kurodai coincided with rising temperature, and spawning occurred from April to September, when the temperature was high. The histological observations of the ovotestis suggested that this species have a single spawning season that extend over six months.

Light characteristics are very specific in the aquatic environment. Fish vision and different light spectra perception are related to each species’ natural habit. Light is one of the main environmental conditions and can be easily manipulated in artificial rearing settings. Cholecystokinin (CCK) and mucus-secreting goblet cells are the main regulators of digestion. In this study, we established whether the light spectrum (natural condition, full spectrum: green, 520 nm; red, 590 nm, and blue, 480 nm) influences growth performance and digestive activity related to CCK mRNA expression and mucus-secreting goblet cell activity in order to develop a good management protocol and optimal rearing system for the longtooth grouper. For each light spectrum, fish were reared 12 weeks under a flow-through system and fed commercial pellet diets once daily. At the end of the experiment, the final body weights differed among the fish reared under different light spectra. The highest growth performance value was observed in fish reared under the green light condition. On the other hand, the growth performances of fish in the natural and blue light conditions were drastically decreased in last 3 weeks of the experiment. CCK mRNA expression and mucus-secreting goblet cell activity were significantly higher in the fish under green light condition than in the fish under the natural, red, and blue light conditions. Rearing of the longtooth grouper under the green light condition had positive effects on fish growth performance and digestion. We recommend that the appropriate light spectrum for the artificial culture of the longtooth grouper is the green light condition from the perspective of growth performance and the synergistic effects of CCK and mucus-secreting goblet cells. However, longer light treatment periods are needed in future investigations to clarify the effects of light spectrum on the longtooth grouper. Together with the findings of the present study, such studies would result in better understanding of the digestive physiology and contribute to the development of optimal rearing management for commercial production of the longtooth grouper.

One of challenging goals of developmental biology is understand of genetic inheritability of developmental programming. This challenging task first requires identification of all genetic factors and their functional dependences, revealing holistic view of genetic organization of cellular and organismal development. Because of high complexity of genotype as well as phenotype, genetic dissection of developmental programs could be even untouchable by combinatorial explosion of the number of possible associations. Therefore, modern genetics needs to be more systematic and predictive. Recently we proposed network-guided approach for genetics of complex traits. First, we construct probabilistic functional gene networks for cells or organisms by benchmarking and integrating heterogeneous multi-omics data that are in general publicly available. Then, using guilt-by-association, and other algorithms of network propagation of known biological information, we predict gene functions, phenotypic effect of loss-of-function, and epistatic interaction. The information can contribute to reconstruction of map between genotype and phenotype. The network-guided genetics method has been effectively applied for various organisms; from simple microbe yeast, to multicellular animal C. elegans, and to the human.

237Np is very important material in the fission products of nuclear reactors. Resonance integral(RI) tests of this material is necessary to check between the experiments and the evaluated data. Such feedback to the evaluated data is very important to correct data and improve of codes. The RI for the 237Np(n,γ)238Np reaction were measured by using the 46-MeV electron linear accelerator (linac) at the Research Reactor Institute, Kyoto University (KURRI). The measurement was performed in the energy region from 0.005 eV and 10 keV. RI obtained as 804.7 barns, compared with those of the evaluated data in JENDL-4.0 and Mughabghab.

In order to simulate a free surface flow in a trench channel, a three-dimensional incompressible unsteady Reynolds-averaged Navier-Stokes (RANS) equations are closed with the model. The artificial compressibility (AC) method is used. Because the pressure

This experiment was carried out to investigate the appropriate chilling requirements for breaking dormancy by treating the dormant plant of Hanabusaya asiatica with low temperature (4℃) for different time periods. The rates of sprouting and flowering were higher with longer treatment periods at low temperature. In addition, the growth and flowering of the plant were better when it was potted after treatment at a low temperature for 90 days. The abscisic acid levels and polyphenoloxidase activity of the dormant plant increased during the low temperature treatment, reached a climax 90 days and decreased thereafter. The catalase activity was the lowest after the low temperature treatment for 90 days and increased subsequently. The peroxidase activity increased and showed a sharp rise after the low temperature treatment for more than 90 days. Considering the physiological activities of the enzymes, the changes in the abscisic acid levels, and the characteristics of growth and flowering after sprouting of the plant, the appropriate cold periods required for breaking dormancy could be 90 days.

The amphetamine derivative 3,4-methylenedioxymethamphetamine (MDMA) is a potent monoaminergic neurotoxin with the potential to cause serotonergic neurotoxicity, but has become a popular recreational drug. Little has been known about the cellular effects induced by MDMA. This report shows that MDMA inhibits neuronal cell growth and differentiation. MDMA suppressed neuronal cell growth. The results of quantitative real-time PCR analysis showed that Egr-1 expression is elevated in mouse embryo and neuroblastoma cells after MDMA treatment. Transiently transfected Egr-1 interfered with the neuronal differentiation of neuroblastoma cells such as SH-SY5Y and PC12 cells. These findings provide evidence that the abuse of MDMA during pregnancy may impair neuronal development via an induction of Egr-1 over-expression.

From “The Love Song of J. Alfred Prufrock” to “Old Possum’s Book of Practical Cats,” he often used such animal imagery as cats, possum, and other animals in his poems. Eliot was haunted by animal images in that he expressed his inner self through animal imagery. In particular, he frequently used reserved animals to show his true self. In his poems, Eliot articulated significance of such animals as possum, cat, rat, and rabbit because these animals hide their inner nature in their minds. In addition, animals react intuitional response as shown in possum and rats, animals are the perfect imagery to manipulate human mind. Therefore, Eliot used animal imagery to manifest human archetypal minds. In “The Love Song of J. Alfred Prufrock,” Eliot connotes the imagery of a cat. In “Gerontion,” he called Christ the tiger because he was frighted by Christ as the absolute judge. In The Waste Land, the death that rats create is not a death which brings resurrection and new life like Jesus Christ, but brings about no life at all in the waste land. Especially, Eliot called himself “possum” in his letters to Pound. Even in his unpublished poem, “Cow,” the cow shows his panic emotion from his first marriage. Therefore, Eliot’s use of animal imagery makes significant roles to understand one of his major themes.

Lipid metabolites involved in cellular regulation as signaling mediators. Prostaglandins (PGs), metabolites of lipid are involved to pregnancy at the time of implantation but the functional roles of PGs on embryo development are still controversy and largely unknown. In previous report, the levels of and at embryos of morula stage and blastocyst stage were explored (Cheon et al., 1998). In this study, the previous suggestion was confirmed and the possible downstream mediator of prostaglandin and prostaglandin on the expansion and hatching of mouse embryo was examined. As expected, developmental rate of the blastocyst to expanded stage was a concentration-response curve that showed the highest expansion rate at 10 , but at 100 , the rate was decreased. In contrast to the , stimulated expansion without toxicity at highest concentration. Cotreatment of PGs with indomethacin overcame the inhibitory effects of indomethacin in expansion. Exogenous PGs also improved the development of expanded embryos to the hatching stage. Besides, PGs receptors' transcripts detected at blastocyst. was caused of calcium fluctuation in the blastocyst but did not. The changes of intracellular calcium concentration were different between indomethacin pretreated embryos and non-treated embryos. Based on these results it is suggested that PGs work as paracrine and/or autocrine factors through calcium and the others which were not identified in this study.

It is suggested that FGF/Ras/MEK/Erk signaling plays crucial roles in specification and cell division of the mesodermal precursor cells in ascidian embryos. To investigate how the number of cell division in tissue precursor cells is determined, we have characterized Wee1 homolog, Hr-Wee1 of the ascidian Halocynthia roretzi. We found that the Hr-Wee1 mRNA is expressed both maternally and zygotically. Maternal transcript is localized to the cytoplasm in the animal cells, while zygotic expression is seen in cells of the endoderm lineage from 32-cell to 110-cell stages. Zygotic in situ signal is detected in the A-line neural plate cells of neurulae, and in epidermal cells of the head region of tailbud embryos. Embryos treated with MEK signaling inhibitor showed a similar pattern to normal embryos in expression of Hr-Wee1. Therefore, it is likely that MEK signaling does not affect the maternal and zygotic expression of Hr-Wee1.

This study was carried out to investigate the effects of wormwood (Artemisia sp.) addition on the growth performance, nutrients utilization and abdominal fat deposition of broiler chickens. Two hundred twenty five and two-day old Arboracre strain male commercial broiler chicks were distributed to 5 treatments with wormwood supplementation levels; C(0%), T₁ (1%), T₂ (3%), T₃ (5%), and T₄ (10%) and with 3 replications each with 5 birds for five weeks. Body weight gain during the experiment was improved in T₁ (1723.0g) compared with that of T₃ (1557.7g) and T₄ (1450.7g) (P＜0.05). Feed intake was significantly (P＜0.05) increased as the levels of wormwood addition increased (C: 2653.8g, T₁: 2852.0, T₂: 2900.3, T₃: 2900.7g, T₄: 2954.7g). Feed conversion rate (feed/gain) was significantly (P＜0.05) increased as the levels of wormwood addition increased (C: 1.55, T₁: 1.66, T₂: 1.70, T₃: 1.86, T₄: 2.04). The days reaching to 2.0㎏ of body weight were expected to be 43.2 days in control, whereas those of group T₁ were reduced to 42.6 days by 0.6 day from control. Nutrient utilization and abdominal fat deposition in the experiment were significantly decreased (P＜0.05) and small intestine contents in the broilers was significantly increased (P＜0.05) as the levels of wormwood addition increased. Therefore, although there was no significant improvement for the performance of broiler chickens with the dietary supplementation of wormwood meal, less than 1% addition of wormwood to broiler diets might have beneficial for human health by reducing the abdominal fat deposition of the broiler chickens.

The canine major histocompatibility complex (MHC) is referred to dog leukocyte antigens (DLA), which is known to be the most polymorphic genetic system in canine species. Many cloned dogs have been produced since Snuppy, first cloned dog, there was no research about genetic identity of MHC among cloned animals. Recently in Lee’s group, two non-transgenic cloned beagles (BG1, 2) were produced by somatic cell nuclear transfer (SCNT) using fetal fibroblast (BF). Also, four transgenic cloned beagles (Ruppy 1-3, 5) were generated using transgenic BF transfected with Red fluorescent protein (RFP) gene. We hypothesize that non-transgenic (BG1, 2) and transgenic (Ruppy 1-3, 5) cloned beagles derived from identical donor cells have the same immunological genetic characteristic except for RFP gene insertion in the genome. Thus, the aim of this study is to confirm the immunological identity of DLA class II in cloned beagles produced using same nuclear donor cell. Genomic DNA was extracted from blood of BG1, BG2, Ruppy 1, 2, 3 and 5. Genomic DNA of normal two control beagle, no correlation with BF was also investigated for rulling out the possibility that beagles were inbred. Forward and reverse primers used for DLA-DQA1 and DQB1 respectively were DQAF: 5’-TAAGGTTCTTTTCTCCCTCT-3’ and DQAR: 5’-GGACAGATTCAGTGAAGAGA-3’ DQBR:5’-CTCACTGGCCCGGCTGTCTC-3’ and DQBR: 5’-CACCTCGC CGCTGCAACGTG-3’. Polymerase Chain Reaction (PCR) products were purified, sequenced directly using the Big Dye Terminator kit. Sequencing analysis was performed on an automated 3730xl DNA analyzer. In experiment 1, sequence of DLA-DQ alpha 1 (DQA1) and DLA-DQ beta 1 (DQB1) exon 2, hypervariabel region, was compared in BG1 and BG2. Experiment 2 also compared the sequence of DQA1 and DQB1 among Ruppy 1, 2, 3 and 5. Experimental 3 compared sequence of DQA1 and DQB1 among all cloned dogs (BG1, BG2 and Ruppy 1-3, 5). As a result, BG1 and BG2 have same allele for DQA1 and DQB1 as we expected. They share DQA1*00101 and DQB1*02901 in experiment 1. In experiment 2, Ruppy 1, 2, 3 and 5 also have identical DQA1*00101 and DQB1*02901 allele. No discrimination between transgenic dogs and cloned dogs was seen in DQA1 and DQB1 Allele in experiment 3. DQA1, DQB1 allele was identified as *00101 and *02901 in all dogs. We provided the allele identity of DQA1and DQB1 in cloned beagles, which can be used as preliminary data for immunological related studies. In conclusion, transgenic cloned dogs despite of red fluorescent protein genes being inserted in their nuclear DNA were immunologically compatible with non-transgenic cloned dogs. We demonstrated that cloned beagles produced using identical nuclear donor were immunologically compatible.

The transcription factors, DMRT1 and FOXL2, play a role in fish sex differentiation of the bipotential precursor into the male and female pathway, respectively. In order to provide the molecular background for understanding hormonal regulation in sexual determination and differentiation in the red-spotted grouper, Epinephelus akaara, one of commercially important epinephelines, and is often used to study protogynous sex change. First, we amplified the partial sequence of two genes (DMRT1 and FOXL2) from the gonad of red-spotted grouper. Also, we surveyed the tissue-specific and sex-specific expression pattern of each genes by RT-PCR. The effects of 17α-methyltestosterone (MT) in the sexually immature gonad of red-spotted grouper were investigated by Real-time quantitative RT-PCR. Fish were treated with MT-dipping method from around 70 days after hatching (DAH) for two month. DMRT1 and FOXL2 cDNA flagments consist of 489 and 836 base pairs (bp) and encodes a protein of 162 and 278 amino acids, respectively. RT-PCR revealed that DMRT1 mRNA was expressed higher level in the testis. Foxl2 was expressed extensively in the neural and peripheral tissues with its highest level in the ovary, indicating a potential role for Foxl2 in the brain-pituitary-gonad axis. Real-time quantitative RT-PCR analyses showed that DMRT1 mRNA expression was upregulated in the MT-treated fish. These results suggest that the sex inversion of red-spotted grouper by MT might be due to the suppression of FOXL2 gene expression, and resulting in the induction of the 11-KT secretion.

Controllable transgenic expression systems in transgenic animal model are valuable to the development of therapeutic approaches in human medical fields. The aim of this study was to 1) produce a transgenic cloned dog using inducible tetracycline vector system, and 2) investigate whether the transgenic cloned dog could be induced the transgene expression using doxycycline (Doxy). Canine fetal fibroblasts were infected with retroviral vectors designed to express the enhanced green fluorescent protein (eGFP) gene under the control of tetracycline-inducible promoter. For somatic cell nuclear transfer (SCNT), nucleus of an in vivo matured oocyte was removed and an eGFP expressed cell cultured with 1 ㎍/㎖ of Doxy was injected. After electrical fusion and chemical activation, the reconstructed embryos were transferred to a recipient and pregnancy diagnosis was performed by ultrasonography. Experiment I evaluated the mean fluorescence intensity (MFI) of infected cells while the cells were cultured in the presence of 1 ㎍/㎖ of Doxy for 5 days, and then in the absence of Doxy for 7 days using fluorescence-activated cell sorter. Experiment II was designed to produce an eGFP controllable transgenic cloned dog via SCNT. For verification of transgenic dog, experiment III was performed Southern Blot analysis and observation in vivo regulation of eGFP expression in the cloned dog treated with 100 ㎎/㎏ of Doxy every 2 days for 2 weeks under ultraviolet light. In experiment IV, western blot was used to detect eGFP increase and decrease in skin tissues of transgenic dog under the presence or absence of Doxy. In the results of experiment I, the MFI for infected cells was rapidly increased to approximately 42.3 times after 3 day-treatment compared to pre-treatment and quickly decreased 3 days after ceasing the treatment. In experiment II, a total of 203 embryos were transferred to nine recipients and three pregnant delivered three pups (Tet-on eGFP 0, Tet-on eGFP 1, and Tet-on eGFP 2) by C-sec and Tet-on eGFP 2 among them is still alive. All cloned pups were genetically identical to the donor cell. Tet-on eGFP 2 showed an apparent in vivo eGFP expression on her body after Doxy administration in experiment III. The result of Sothern blotting showed that the transgene insertion was detected from the three cloned dogs and all organs of Tet-on eGFP 1. Experiment IV indicated that a robust eGFP expression in skin tissue of Tet-on eGFP 2 rapidly increased after Doxy treatment and gradually decreased to basal level on 9 weeks after ceasing the treatment. In conclusion, we report here for the first time an inducible transgenic system in canine species and it can stably induce the transgene expression at intended time. This study has demonstrated the capacity to generate transgenic model dog which could regulate the transgene and it would contribute to human medical research fields.

The objective of this study was to investigate the effects of oxygen tension during in vitro maturation of porcine oocytes on the nuclear maturation and differences in gene expression. Cumulus-oocyte complexes (COCs) were collected from ovaries obtained at a local slaughterhouse, matured for 44 hours in TCM199 supplemented with porcine follicular fluid (pFF) under 5% or 20% oxygen concentration. In results, oxygen tension had no significant effects on nuclear maturation. Relative poly(A) mRNA abundance of MnSOD, CCNB1, LDHA, G6PD, BCL, GPX1, IGFR2, GLUT1, BAX, GREM, PTGS2 was analysed in cumulus cells. GLUT1, G6PD, LDHA were up-regulated in the cumulus cells matured in low oxygen, suggesting a higher glucose uptake and an increase in anaerobic glycolysis, whereas CCNB, MnSOD were up-regulated in the cumulus cells matured in high oxygen, which suggest a higher activity of mitosis-promoting factor and antioxidant response. In conclusion, cumulus cells increase in glucose metabolism via anaerobic glycolysis under low oxygen concentration and show significant change in antioxidant against oxidant damage or apoptotic response under high oxygen concentration. For such an effect of cumulus cells, oocytes could be matured normally regardless of various oxygen concentration.

This study is to survey the ultrastructure of gamete cells and micropyle of pre-fertilized and post-fertilized eggs after HCG hormone treatment by transmission and scanning electron microscopy (TEM and SEM) in E. akaara, E. bruneus and E. septemfasciatus. These fishes are economical importance species for Jeju coastal area resources. In spite of its an importance resources, details studies on the ultrastructural aspects of gamete cells for its reproductive biology have not been undertaken. Morphological features of ovulation process have been studied during its normal occurrence in the reproductive cycle of these fish by light microscopy. Moreover, it has been studied for many years to induce spawning by environmental factors (day length, water temperature etc) or injection of HCG for ovulation in these species. Studies on the micropyle was mainly focused on the eggs of insects, fresh water and a few sea water fishes. Micropylar structure of fish displays morphological characteristics of interspecies-specific by inhabitant environment and spawning feature. On the other hand, it is an importance cue for a taxonomical indicator and identification fish eggs. SEM studies were performed on growing and mature oocytes obtained by stripping and cannulation from 3 grouper species sampled between July and August in spawning season. The outer layer of chorion of preovulatory growing stage oocytes could be divided into four layers; zona pellucida, follicular cell layers consisted of granulosa and thecal cells layer and the most outer ovigerous lamella. Ovulation process of mature stage oocytes initiated by rupture of ovigerous lamella and ovulated by contraction of follicular cell layers. Besides, the micropylar shape of ripe stage oocytes in E. akaara, E. bruneus and E. septemfasciatus presented volcano or crateriform-like cylindrical form. Internal structure of micropylar vestibule displayed cylindrical clockwise 8 or 10 spiral arrangement structure in these species. The micropyle diameter and apparatus at the animal pole differ significantly among the 3 species. The difference in their diameters suggests species-specific in the correlation between spermatozoal head size and micropylar diameter for polyspermy prevention and hybridization during fertilization. Besides, after artificial fertilization, the vestibule morphologically transformed into dom-shape and pillar-shape for fertilization cone formation. Pores of various sizes in the 3 grouper species were somewhat regularly distributed in concentric circles only around the micropyle. In particular, large pores had numerous gill filament-shaped projections connected to oolemma. These structures are suggested to be related to gas exchange, osmoregulation, and micronutrient influx or efflux between eggs and water during fertilization and egg development. In addition, spermatozoa ultrastructure was examined in 3 grouper species. TEM investigation revealed that, in all species, spermatozoa display a round head, a nucleus containing highly condensed, filamentous chromatin clusters, no acrosome, a short midpieces consisting of numerous mitochondria and the proximal and distal centrioles and a flagellum exhibiting the typical axoneme structure (9+2). Especial, both E. akaara and E. bruneus display regular laternal fins in flagella, but in E. septemfasciatus, no fins in flagella with hook shape tails.

Mammalian reproduction is regulated by a feedback circuit of the key reproductive hormones such as GnRH, gonadotropin and sex steroids on the hypothalamic-pituitary-gonadal axis. In particular, the onset of female puberty is triggered by gain of a pulsatile pattern and increment of GnRH secretion from hypothalamus. Previous studies including our own clearly demonstrated that genistein (GS), a phytoestrogenic isoflavone, altered the timing of puberty onset in female rats. However, the brain-specific actions of GS in female rats has not been explored yet. The present study was performed to examine the changes in the activities of GnRH neurons and their neural circuits by GS in female rats. Concerning the drug delivery route, intracerebroventricular (ICV) injection technique was employed to eliminate the unwanted actions on the extrabrain tissues which can be occurred if the testing drug is systemically administered. Adult female rats (PND 100, 210-230 g BW) were anaesthetized, treated with single dose of GS (/animal), and sacrificed at 3 hrs post-injection. To determine the transcriptional changes of reproductive hormone-related genes in hypothalamus, total RNAs were extracted and applied to the semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). ICV infusion of GS significantly raised the transcriptional activities of enhanced at puberty1 (EAP-1, p<0.05), glutamic acid decarboxylase (GAD67, p<0.01) which are known to modulate GnRH secretion in the hypothalamus. However, GS infusion could not change the mRNA level of nitric oxide synthase 2 (NOS-2). GS administration significantly increased the mRNA levels of KiSS-1 (p<0.001), GPR54 (p<0.001), and GnRH (p<0.01) in the hypothalami, but decreased the mRNA levels of LH- (p<0.01) and FSH- (p<0.05) in the pituitaries. Taken together, the present study indicated that the acute exposure to GS could directly activate the hypothalamic GnRH modulating system, suggesting the GS's disrupting effects such as the early onset of puberty in immature female rats might be derived from premature activation of key reproduction related genes in hypothalamus-pituitary neuroendocrine circuit.

This study attempted to verify the possibility of using germ cell aspiration (GCA) method as a non-fatal technique in studying the life-history of equilateral venus, Gomphina veneriformis (Veneridae) and granular ark, Tegillarca granosa (Arcidae). Using twenty-six gauge 1/2" (12.7mm) needle, GCA was carried out in equilateral venus through external ligament. In granular ark, GCA was performed by preventing closure of the shells by inserting a tongue depressor between the shells while still open. The success rate of sex identification using the GCA method was 95.6% for the equilateral venus (n=650/680) and 94.3% for the granular ark (n=707/750). Mortality of equilateral venus, which spent 33 days under wild conditions, was 13.8% (n=90/650) while the mortality of granular ark, which spent 390 days under wild conditions, was 2.4% (n=17/707). Although we believe that GCA does not appear to cause death in equilateral venus or granular ark, the success rate in employing of this methodology may differ depending on the level of proficiency of the researcher and reproductive stage of the bivalve. This study concludes that GCA is a convenient non-fatal methodology, which can be employed to identify sex and investigate gonadal maturity in Gomphina veneriformis and Tegillarca granosa.

We investigated the androgenic effects of 11-ketotestosterone (11-KT) on gonadal sex reversal and spermatogenesis in honeycomb grouper Epinephelus merra by method of gonadal biopsy. 11-KT was injected intramuscularly at a concentration of 1 and body weight. The proportion of cross sectional area of the gonad occupied by each germ cell type was measured and compared pre- and post-injection group. During the sex change phase, the distribution ratio of oocytes was decreased in all fish of 11-KT treatment group while the distribution ratio of spermatocytes was increased than pre-injection group. In male phase, all fish of 11-KT treatment group shown the increased distribution ratio of spermatocytes, but the distribution ratio of spermatozoa was decreased than pre-injection group. The present results suggest that 11-KT can stimulate degeneration of oocytes, proliferation of spermatocytes and spermiation in honeycomb grouper.