• Development and Reproduction
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• v.27 no.3
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• pp.127-135
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• 2023

Effects of changes in photoperiod on the reproductive events in fish are suggested to be mediated mainly via the action of melatonin (MEL). Changing levels of plasma MEL throughout the day and year are suggested to influence the hypothalamus-pituitary-gonadal axis in fish. Therefore, in this study, we aimed to investigate the effects of MEL on oocyte maturation and germinal vesicle breakdown (GVBD) in the marine fish, Chaenogobius annularis, in vitro. Oocytes at three different stages (pre-, mid-, and late-vitellogenesis) were incubated with (a) only MEL (5, 10, 50, 100, 500, and 1,000 pg/mL) and (b) 50 pg/mL of 17α,20β-dihydroxy-4-pregnen-3-one (17α20βP), maturation-inducing hormone (MIH) of this species, and MEL (4-h incubation before addition of MIH). Any single MEL treatment did not significantly induce GVBD. However, treatment with 50 pg/mL MEL or MIH significantly induced GVBD. These results suggest that preincubation with MEL accelerates the effect of MIH on longchin goby oocyte maturation.

• Asian-Australasian Journal of Animal Sciences
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• v.14 no.5
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• pp.693-696
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• 2001

The effect of replacement of in vitro maturation medium completely with the buffalo follicular fluid (buFF) on in vitro oocyte maturation of buffalo oocytes was studied. 5 to 8 buffalo cumulus oocyte complexes were cultured in a single drop with each of the eight media studied i.e., M199+steer serum (10% v/v), M199+steer serum (10% v/v)+PMSG, M199+buFF (10% v/v), M199+buFF (10% v/v)+PMSG, M199+buFF (50% v/v), M199+buFF (50% v/v)+ PMSG, buFF (100%) and buFF+PMSG at $39^{\circ}C$ and 5% $CO_2$ in air for 24 h. Supplementation of M199 with Steer serum alone resulted in IVM rate of 35% only. When the above medium was supplemented with PMSG, the maturation rate rallied to 82%. Significant increase in the maturation rates were observed when M199 was supplemented with increasing levels of buFF. A further increase in the maturation rate was also obtained when PMSG was incorporated into the medium of M199 supplemented with buFF. The rate of maturation was to the tune of 91% when oocytes were matured in buFF alone which was increased non significantly on the addition of PMSG. Highest maturation rate (97%) obtained with M199+buFF (50%v/v)+PMSG did not differ significantly from that obtained by either M199+buFF (10%v/v)+PMSG or buFF+PMSG. It is suggested that buFF alone without any supplementation can form the effective in vitro maturation medium for buffalo oocytes.

• Korean Journal of Animal Reproduction
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• v.19 no.4
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• pp.251-258
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• 1996

These studies were carried out to investigate the effect of gonadotropins (GTH), fetal calf serum (FCS), porcine follicular fluid (pFF) and FCS and pFF fractions obtained by the gel filtration on in vitro maturation of porcine follicular fluid. When the oocytes were cultured in TCM-199, the maturation rate was higher in pFF than in FCS in both with or without GTH and in pFF the maturation rate was higher in with GTH than in without GTH. In case of without GTH, pFF increased maturation rates in TCM-199, but not in Whitten's medium (WM). When the oocytes were cultured in WM supplemented with FCS fractions, the maturation rate(51.6%) of oocytes was significantly (P<0.05) higher in fraction B (about 30∼70 kDa) than in control, FCS and other fractions. When oocytes were cultured in WM supplemented with pFF fractions, fractions B (about 30∼70 kDa) and D (about 1∼10 kDa) were significantly (P<0.05) higher than in control, pFF and other fractions. In conclusiion, the addition of gonadotropins into the maturation media was effective for oocyte maturation. The addition of pFF was more effective than addition of FCS for maturation of porcine oocytes in vitro. And fraction B from FCS and fractions B and D from pFF was effective for oocyte maturation.

• Korean Journal of Animal Reproduction
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• v.14 no.1
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• pp.84-91
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• 1990

Experiments were disigned to define and optimize efficiency of a system whereby pig follicular oocytes could be matured and fertil ized in vitro. The pig oocytes removed from 1- 2 mm and 3-7 mm follicles were cultured in vitro in the mKRB(-BSA) solution containing estrous sow serum (ESS), FCS or dialyzed pig follicular fluid for 24 to 48 hr at 37$^{\circ}C$. The oocytes matured in vitro were evaluated after epididymal spermatozoa-oocyte incubation for 24 hr for pronucleus formation. 50-60% of the oocytes reached metaphase II during 36 to 48 hr of culture. There was no differernce in oocyte matura¬tion between two groups of follicular size but meiosis was slightly faster in the 3-7 mm follicular oocytes. The oocytes matured in mKRB (-BSA) plus 5% ESS, 15% FCS or dialyzed follicular fraction showed slightly higher maturation rates than the control mKRB. in vitro fertilization, pronucleus formation, tended to be increased when mKRBi-BSA) plus 5% ESS or 15% FCS was used for oocyte maturation and in vivo -capacitated spermatozoa were inseminated, respectively. It is concluded that ESS, FCS and dialyzed pig follicular fluid may be effective factors for in vitro maturation and fertilization of pig follicular oocytes.

• The Korean Journal of Zoology
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• v.33 no.1
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• pp.53-62
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• 1990

Fully grown amphibian oocytes undergo their maturation (germinal vesicle breakdown, GVBD) during in vitro follicle culture when they are stimulated with frog pituitary homogenate (FPH) or progesterone. Present experiments were designed to determine whether proteolytic enzymes are involved in the regulation of the matunation process. Treatment of a $\alpha$ -chymoiyypsin inhibitor, N a -tosyl-L-phenylalanine-chloromethyl-ketone(TP) to the oocytes exhibited a biphasic phenomenon, the induction of the maturation without added hormone at relatively low doses (0.001-1 $\mu$M) and inhibition of the hormone induced oocyte maturation at a high dose (100 $\mu$M). Treatment of a trypsin inhibitor, N a -tosyl-L-lysine-chloromethyl ketone(TLCK) to the oocytes did not induce the maturation, but rather suppressed the hormone induced oocyte maturation in a high dose(100 $\mu$ M). Treatment of exogenous iyypsin to the oocyte induced their maturation without added hormone in a dose dependent manner (0.001-1 $\mu$ M). The data presented here indicate that some proteolytic enzymes play a role in the regulation of the maturation(meiotic arrest or reinitiation) in amphibians.

• Journal of Animal Reproduction and Biotechnology
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• v.34 no.3
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• pp.205-211
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• 2019

In this study, we investigated the possibility of using mouse embryonic stem cell conditioned medium (ESCM) and embryonic stem cell medium (ESM) for in vitro maturation in the efficient in vitro production of blastocysts from porcine follicular oocyte. Depending on the concentration of supplement of ESCM added to the NCSU-23 solution did not affect 2-cell development rates and blastocysts development. However, in particular, the survival rate (10 days of culture) of blastocyst was significantly higher than that of the control group as the additive concentration (30%) increased (p < 0.05). The survival rate of blastocysts showed a similar tendency even with addition of ESM (30%) alone. On the other hand, the duration of the addition of these additives during IVM (0-44 h) was that the IVM I period (0-22 h) were more effective than the IVM II period (22-44 h). Thus, the effect of these additives is probably due to the combination of the various physiologically active substances of ESCM or the appropriate amino acids and vitamins of ESM. In particular, these additives were more effective during the first half (IVM I) of in vitro maturation. In summary, optimization of ESCM or ESM supplementation may improve in vitro maturation of porcine oocyte and affect developmental competency. Therefore, if more efficient methods of adding ESCM or ESM to basal culture medium can be developed during in vitro maturation of porcine follicle oocytes, high quality blastocysts will be developed from low porcine follicular oocyte compared to other domestic animals.

• Journal of Embryo Transfer
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• v.11 no.2
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• pp.179-191
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• 1996

In-vitro culture has provided new inforrnation on mechanisms of oocytes rnaturation and results obtained in vitro have led to new questions. In porcine, follicular and oocyte size have the crucial importance for the oocytes maturation. The addition of hormones to the culture medium was found to accelerate and facilitate meiotic maturation. The presence of some factors in serum trigger the resumption of meiosis and support the maturation of oocytes in vitro. The maturation rate of porcine oocytes was also increased by supplementation of porcine follicular fluid to the culture medium. The growth factors can stimulate nuclear maturation and enhances cytoplasnic maturation of oocytes by interaction with gonadotropins. The maturation-promoting factor brings about GVBD and the subsequent maturational events in oocytes. However, cAMP can block the spontaneous meiotic maturation of oocytes in culture. The understanding of these influences is a prerequisite to enhancing in vitro maturation of porcine oocytes.

• Korean Journal of Animal Reproduction
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• v.22 no.3
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• pp.253-264
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• 1998

Porcine follicular oocyte, collected from antral follicles (2~5 mm in diameter) of gilt ovaries were matured in vitro porcine follicular fluid (pFF) with PMSG (pFF-PMSG) buffer with at 37$^{\circ}C$ under 5% CO2 in air their ability of maturation promoting factor (MPF), of GV and GVBD formation was examined followed during time after in vitro culture. Formation of second metaphase was observed in 57.6% and 71.2% of matured in with pFF-PMSG buffer to 45 and 50 hours after invitro. Porcine oocytes cultured in pFF-PMSG for various periods of up to 30 hours were stained with Hoechst-33342 and classified according to maturation before assaying. Histone H1 kinase (H1K) activity was assayed during meiotic maturation in porcine oocytes matured in pFF-PMSG buffer in vitro. In oocytes matured in pFF-PMSG, H1K activity was at the 30 hours after culture and increased about 15 fold than at the germinal vesicle stage with before at the cultured in vitro. This pattern is similar to those reported in non-mammalian species and su, pp.rts the concepts that H1K is ubiquitous in eukaryotes and controls the meiotic cell cycle in mammals. These results suggest that the maturation pFF-PMSG buffer used influences the fluctuation pattern of H1K activity and biological characteristics of porcine oocytes cultured in vitro.

• International Journal of Oral Biology
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• v.30 no.4
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• pp.131-134
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• 2005

In the present study, performances of several in vitro maturation (IVM) systems for porcine follicular oocytes were evaluated, and an efficient chemically defined IVM system for porcine oocytes was proposed. The proposed one-step culture system supplemented with polyvinylalcohol (PVA) gave competitive efficiencies in terms of oocyte maturation and blastocyst development after parthenogenetic activation and in vitro culture, compared with the conventional two-step culture system by a supplementation of porcine follicular fluid (pFF). Additionally, it is identified that the proposed chemically defined one-step culture system yielded the comparable level of blastocyst production to the conventional maturation system in porcine somatic cell nuclear transfer (SCNT). Therefore, one can eliminate un-expected effects accompanied by supplementation of pFF. No medium replacement during whole maturation period is an additional benefit by applying this new system. Thus, these data support that the developed PVA supplemented chemically defined one-step IVM system for porcine follicular oocyte might be used in porcine SCNT program.

• Clinical and Experimental Reproductive Medicine
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• v.23 no.3
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• pp.357-366
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• 1996

The present experiments aimed to investigate the metabolism of calcium during oocyte maturation in rat. The concentration of free calcium and calmodulin in oocytes was measured respectively by using of fluo-3/AM and FITC with microscope fluorescence spectrometer. The ultrastructural localization of calcium precipitates in oocytes was observed with the transmission electron microscope. Cumulus-free immature oocytes(GV-oocyte) were cultured in vitro through 15 hours. The free calcium concentration in GV oocyte was $55.9{\pm}3.5nM$. In calcium-containing medium, the free calcium concentration was increased in germinal vesicle breakdown(GVBD) oocyte($64.2{\pm}7.3nM$). In normal medium after calcium chelator treatment ($10{\mu}M$ BAPTA/AM), the free calcium contents were slightly lower than those in control group. In calcium-free medium, the free calcium content was drastically increased in GVBD($72.7{\pm}3.4nM$) and metaphase I - anaphase I ($88.0{\pm}3.4nM$) oocyte. In maturation rate of oocytes, GVBD rate was high in control group($82.9{\pm}6.55%$) and calcium chelator treatment group($91.2{\pm}4.4%$), but in calcium-free medium group, it was low and then the oocyte was degenerated without polar body formation. Relative content of calmodulin in oocyte was significantly(P<0.001) increased in metaphase I - anaphase I than in GV and GVBD oocyte. The calcium precipitates were observed in mitochondria and cytoplasm of GV oocyte but that were not observed in mitochondria of GVBD and metaphase I - anaphase I oocyte. And then the calcium precipitates reappeared in mitochondria of metaphase II oocyte. The above results indicate that changes in free calcium and calmodulin concentration of oocyte occur according to the maturational stages and the extracellular calcium is required during oocyte maturation. Also change of calcium localization in oocyte occurs according to the maturational stages.