• Journal of Life Science
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• v.26 no.2
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• pp.164-173
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• 2016

Isotocin (IT), a nonapeptide homolog of oxytocin in mammals, has been suggested to be involved in physiological processes including social behaviors, stress responses, and osmoregulation in teleost fish. To study its structure and function, the gene encoding the IT precursor was cloned from the genomic DNA and brain cDNA of the cinnamon clownfish, Amphiprion melanopus. The IT precursor gene consists of three exons separated by two introns, and encodes an open reading frame of 156 amino acid (aa) residues, comprising a putative signal peptide of 19 aa, a mature IT protein of 9 aa, a proteolytic processing site of 3 aa, and 125 aa of neurophysin. Tissue-specific analysis of the IT precursor transcript indicated its expression in the brain and gonads of A. melanopus. To examine its osmoregulatory effects, the salinity of the seawater (34 ppt) used for rearing A. melanopus was lowered to 15 ppt. Histological analysis of the gills indicated the apparent disappearance of an apical crypt on the surface of the gill lamella of A. melanopus, as pavement cells covered the surface upon acclimation to the lower salinity. The level of Na⁺/K^+-ATPase activity in the gills was increased during the initial stage of acclimation, followed by a decrease to its normal level, suggesting its involvement in osmoregulation and homeostasis. The only slight increase in the level of IT precursor transcript in the A. melanopus brain upon low-salinity acclimation suggested that IT played a minor role, if any, in the process of osmoregulation.

• Development and Reproduction
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• v.15 no.2
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• pp.121-131
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• 2011

The aim of this study was to investigate the egg development and the color change of Amphiprion melanopus, by the live food and thyroid hormone. After fertilizing, it took 168 h until hatching. The size of hatching larvae were $3.5{\pm}0.3$ mm, and some individuals had small yolk sac. As reaching the particular age, they underwent the color change (metamorphosis) - Days after hatching (DAH) 10: black, DAH 15~20: the 3 white stripes appear, DAH 30: the dorsal & the caudal white stripe begin to disappear. DAH 90: body color become red and orange, adult: cinnamon body color and a white stripe on head. - The group fed enriched Artemia with Tigriopus japonicus grew the faster than with only Artemia. The fin coloration of the one fed the only T. japonicus has a vivid red and orange color. $T_3$-treated group (4 ppm) grew the faster than the others (0, 2, 6 ppm). The size of the 6 ppm group was the smallest. The results suggest T. japonicus can be used as a supplement with rotifer and Artemia for the juvenile development and coloration. $T_3$ is considered to be involved in the metamorphosis and color formation of Amphipirion sp.

• Journal of Aquaculture
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• v.20 no.4
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• pp.239-247
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• 2007

We studied spawning, hatching and larval growth of Amphiprion melanopus under laboratory conditions. Each time a fish spawned about 440 to 650 eggs. The eggs were hatched 9 days later. Nest colour of the eggs changed from clear yellow orange to greyish violet with processing of embryogenesis. With regard to spawning according to the substrates, the fish prefered red clay pot and ceramic pot to volcanic scoria and live rock. Hatching rate (%) of the eggs on the former substrates was significantly higher than that on the latter. The number of eggs varied with the food stuff supplied to the fish. The fish fed Tetra bits (commercial diet), meat of short necked clams and mussels, and astaxanthin showed significantly higher fecundity with 638 eggs than that fed only Tetra bits (477 eggs) or Tetra bits with meat of shellfish without astaxanthin ($549{\sim}553\;eggs$). Higher temperature ranging from 24 to $30^{\circ}C$ induced shorter incubation period and smaller total length, but higher hatching rate. The optimum salinity and L:D cycle with $60\;{\mu}mol$ photon $m^{-2}s^{-1}$ for hatching were 33 psu and 16:8, respectively. The newly hatched larvae about 4.6 mm in total length grew to 7.5 mm in one week, 18.8 mm in a month and 37.9 mm in 70 days after hatching.

• Animal cells and systems
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• v.16 no.5
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• pp.391-399
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• 2012

We cloned cDNA-encoding arginine vasotocin (AVT) from the brain of the cinnamon clownfish Amphiprion melanopus, and that was predicted to encode a protein of 153 amino acids. We examined changes in the expression of AVT mRNA in the brain and arginine vasotocin receptor (AVTR) mRNA and osmotic stress transcription factor 1 (OSTF1) mRNA in the gills of the cinnamon clownfish using quantitative real-time PCR in an osmotically changing environment (seawater (35 psu) ${\rightarrow}$ brackish water (BW, 17.5 psu) and BW with prolactin [PRL]). The expression of AVT, AVTR, and OSTF1 mRNA in the brain and gills increased after transfer to BW, and the expression was repressed by PRL treatment. AVT-immunoreactive cells were almost consistently observed in the telencephalon. The plasma $Na^+$ and $Cl^-$ levels decreased in BW, but the level of this parameter increased in BW with PRL treatme during salinity change. These results suggest that AVT, AVTR, and OSTF1 play important roles in hormonal regulation in osmoregulation organs, and that PRL improves the hyperosmoregulatory ability of cinnamon clownfish in BW environment.

• Development and Reproduction
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• v.15 no.2
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• pp.133-141
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• 2011

The marine ornamental industry has become a multi-billion dollar industry these days. As developing, however, this industry has been criticized for the indiscriminate captures and the destruction of the surrounding environment. To circumvent these problems, it is suggested to breed the organisms artificially. While clownfishes Amphiprion sp. and Premnas sp. are the most famous ornamental organisms in the trade, few studies are yet available on the culture and commercial production of these fishes. These studies were performed to investigate the spawning periodicity, behavior and the habits during egg incubation, and to provide the information on the mass hatching system. The spawning periodicity and frequency were different in 4 pairs under the constant condition, temperature, salinity and photoperiod. On the contrary, the male's behaviors for egg incubation are almost same in the all. The egg-fanning activity of the male increased as the developing eggs reaching to the hatching day. Based on the above results, we designed a new artificial hatching system, the rotating type (RT), and compared it with the aeration type (AT) and spray type (ST) that were previously described. RT showed higher hatching rate of 87.3% than AT (74.4%) and ST (60.5%). Also, there were no significant differences in the hatching rate regardless of the number (2, 3, 5) of hatching plates. We suggest RT may accommodate various number of hatching plates and constitute a better hatching system for clownfishes.

• Animal cells and systems
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• v.15 no.3
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• pp.219-225
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• 2011

We studied oxidative stress in cinnamon clownfish exposed to hypoosmotic (35 psu ${\rightarrow}$ 17.5 psu and 17.5 psu with prolactin (PRL)) and low temperature ($28^{\circ}C{\rightarrow}24^{\circ}C$ and $20^{\circ}C$) conditions by measuring the expression and activity of Cu/Zn-superoxide dismutase (Cu/Zn-SOD), catalase (CAT), and glutathione peroxidase (GPX). The expression and activity of the antioxidant enzymes were significantly higher after the fish were exposed to $24^{\circ}C$, $20^{\circ}C$, and 17.5 psu, and expression was repressed by PRL treatment. Furthermore, we measured $H_2O_2$ and lipid peroxidation levels and found that they were significantly higher after exposure to the hypoosmotic and low-temperature environments. Additionally, we investigated changes in plasma AST and ALT levels after exposure to low temperature and hypoosmotic stress. These levels increased upon exposure of the clownfish to $24^{\circ}C$, $20^{\circ}C$, and 17.5 psu, but the levels of these parameters decreased in the 17.5 psu with PRL treatment during a salinity change. The results indicate that hypoosmotic and low-temperature conditions induce oxidative stress in cinnamon clownfish and that the parameters tested in this study may be indices of oxidative stress in the cinnamon clownfish.

• Fisheries and Aquatic Sciences
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• v.27 no.6
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• pp.346-355
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• 2024

Groupers are economically important species in the fishery and aquaculture industries in Asian countries. Various species of grouper, including hybrids, have been brought to market even without precise species identification. In this study, we analyzed the structure and expression profile of the gene encoding prolactin (PRL) in the red-spotted grouper Epinephelus akaara based on genomic DNA and cDNA templates. The results showed that the PRL gene consists of five exons encoding an open reading frame of 212 amino acids, including a putative signal peptide of 24 amino acids and a mature structural protein of 188 amino acids. It showed amino acid identities of 99% with Epinephelus coioides, 83% with Amphiprion melanopus, 82% with Acanthopagrus schlegelii, 75% with Oreochromis niloticus, 70% with Coregonus autumnalis, and 67% with Oncorhynchus mykiss, indicating its closer similarity to E. coioides and other groupers but marked distinction from non-teleost PRLs. PRL mRNA expression was detected mostly in the brain, including the pituitary gland, with little expression in other tissues. While the 5-exon structure of the PRL gene of red-spotted grouper and the exon sizes were conserved, the sizes of the introns, particularly the first intron, were markedly different among the grouper species. To examine whether these differences can be used to distinguish groupers of similar phenotypes, exon-primed intron-crossing analysis was carried out for various commercially important grouper species. The results showed clear differences in size of the amplified fragment encompassing the first intron of the PRL gene, indicating that this method could be used to develop species-specific markers capable of discriminating between grouper species and their hybrids at the molecular level.