• The Korean Journal of Zoology
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• v.31 no.2
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• pp.81-86
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• 1988

The relationship between cumulus cell expansion, cocyte maturation and metabolic cooperativitiy was investigated by using mouse and pig cocyte-cumulus complexes in vitro. Cocyte germinal vesicle breakdown (GVBD) and cumulus expansion were manipulated with hormones or reagents which increase intracellular cAMP leveL Metabolic cooperativity between oocyte and cumulus cells was assessed by determination of the fraction of radiolabelled uridine marker that was transferred from the cumulus mass to the oocyte. Uptake of uddine marker by mouse and pig cumulus mass was increased by about fourfold of basal level with the stimulation of hormones (human choriononic gonadotrophin, HCG; follicle stimulating hormone, FSH) or cyclic AMP sttmulators (3-isobutyl-1-methylxanthine, IBMX; forskolin) during culture. However, the fraction of uridine that was transferred from the cumulus mass to the cocyte (transfer ratio) was gradually decreased during culture, irrespective with the presence of hormones or stimulators. The decrease of the transfer ratio was not correlated with the state of occyte whether they have GV or not, or with the degree of cumulus expansion. In mouse complexes, HCG induced more significant reducton of transfer ratio than other treatments. These results do not support the idea that modulations of metabolic cooperativity between cumulus cells and oocytes are important for the regulation of meiotic resumption in mammals.

• The Korean Journal of Mycology
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• v.26 no.3 s.86
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• pp.314-320
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• 1998

A 0.4 kb cDNA fragment was isolated from mRNA of UV or mechanical wound damaged Pleurotus sajor-caju by the differential display method. Expression of the gene corresponding to this cDNA fragment was highly induced by mechanical wound damage or UV treatment. This gene showed only basal level expression in mycelia, stipe, and cap under normal growth conditions. Sequencing analysis showed that this cDNA fragment contains partial open reading frame. Homology search using genbank database revealed that although this gene do not have homology with already reported wound induced genes, it has a significant sequence homology in defined region with the cdc2-related protein kinase gene which is known to be involved in negative regulation of meiotic maturation in Xenopus oocytes.

• Development and Reproduction
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• v.12 no.3
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• pp.231-241
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• 2008

The present study was designed to clarify aspects of the annual reproductive cycle of the fresh water mitten crab, Eriocheir japonicus inhabiting Sumjin river, and to determine the adequate conditions of temperature, photoperiod and salinity on the gonadal maturation and breeding of adult female crabs. Based on the seasonal changes of GSIs and gonadal histology of adult crabs, the annual reproductive cycle could be divided into 4 periods as follows: previtellogenesis period (from September to October) when GSIs were low value and ovaries had oocytes in previtellogenesis stage and meiotic prophase stage; maturing period (from November to March of the next year) when GSIs increased gradually and ovarian oocytes accumulated yolk globules; spawning period (from April to June) when high value of GSI of female was kept and ovigerous female appeared; resting period (from July to August) when GSIs decreased rapidly, and vitellogenic oocytes degenerated. Gonadal maturation was influenced by water temperature, but not photoperiod and salinity. GSI more increased in experimental regime of $18^{\circ}C$ than that of $10^{\circ}C$ regardless of photoperiod conditions (12L12D and 15L9D). However, in $26^{\circ}C$ regime of both photoperiod conditions, the value of GSI was not changed, and vitellogenic oocytes were not observed. Spawning was considerably influenced by water temperatures and salinities regardless of photoperiods. Vitellogenic female crabs did not spawn at $10^{\circ}C$ in any salinity conditions. However, at $18^{\circ}C$ and $26^{\circ}C$, over a half of rearing female crabs spawned in condition of 30% sea water (salinity 9.6%o) and 60% sea water (salinity 19.2%o), while no female spawned in freshwater condition (salinity 0.0%o).

• Journal of Embryo Transfer
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• v.14 no.1
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• pp.17-22
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• 1999

To investigate the maturational and development competece of porcine oocytes of different diameter groups, oocytes were obtained by aspiration from slaughterdhouse ovaries. After washing three times in NCSU23 medium, each cumulus-oocyte complex was transferred into a $8{mu}ell$ drop of the maturation medium (one oocyte per drop) under paraffin oil. The diameter without zona pellucida of oocytes was measured with micor-calibrator (Mikrometer, E. Leitz) on a screen connected to a VCR on an inverted microscope $(200\times)$. After being measured, the oocytes were divided into 6 groups according to their diameter size : <105, 105 to < 110, 110 to < 115, 115 to < 120, 120 to < 125 and > $125{\mu}{\textrm}{m}$, and in vitro maturation (IVM), fertillzation (IVF) and production (IVP) of oocytes / embryo was performed. The rates of in vitro maturation on oocytes in the greater 105 ${\mu}{\textrm}{m}$ size groups(91.8~100%) were significantly (P<0.05) higher than in the < 105 ${\mu}{\textrm}{m}$ group(66.7%). The rates of sperm penetration were significantly (P<0.05) low in < $105{\mu}{\textrm}{m}$ group (50.0%) than others groups (81.6~85.5%). But the plyspermic fertilization rate was significantly (P<0.05) higher in < $110{\mu}{\textrm}{m}$ oocytes groups than in the $110\leq{\mu}{\textrm}{m}$ size groups. The rates of cleavage and development to blastocysts rose as oocytes diameter increased, however, while oocytes over $120{\mu}{\textrm}{m}$ in diameter failed to develop to blastocysts. There results suggest that porcine oocytes have acquired full meiotic competece at a diameter of $105{\mu}{\textrm}{m}$ but not yet attended full development competence to blastocyst and that oocytes have acquired full development competence at a diameter of $110{\mu}{\textrm}{m}$.

• Korean Journal of Veterinary Research
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• v.34 no.4
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• pp.735-744
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• 1994

Catheters were placed into the external jugular veins of immature female rats. On the following day (day 28 of age), the animals were injected subcutaneously with pregnant mare serm gonadotropin(PMSG): 4IU(control) or 20IU(superovulation). Each animal was sequentially bled at Ohr and 12hr and subsequently at 6hr intervals until sacrifice at 72hr after PMSG. The superovulatory dose of PMSG significantly(P<0.05) increased the ovulatory response by 4.0 fold above controls. On the other hand, superovulated oocytes displayed considerably different stages of meiotic maturation: prophase I (14.7%), anaphase I (36.2%), telophase I (10.3%), metaphase I/II (32.4%), while in control rats a majority of the oocytes examined(94.0%) consistently showed a metaphase II configuration. Serum luteinizing hormone(LH) levels were determined by RIA. Both groups exhibited a similar time relationship with two distinct peaks: an initial slight rise at 0-18hr and a second sharp rise at 54-60hr. However, there was a marked change in the magnitude of LH levels between the two groups. In superovulated animals, prior to the second peak, overall LH levels were significantly(P<0.001) higher than controls. In contrast, at the peak occurring at 60hr, LH concentrations were significantly(P<0.001) reduced by 54% below that of control. Additionally, a maximum increase of mean ${\Delta}LH$ between two peaks was much less in superovulated as compared to control rats. The initial prolonged elevation of serum LH before 54hr in superovulated rats was found to result from actual cross-reaction of the injected PMSG with LH antibody in the assay, while a precipitous second elevation between 54hr and 60hr resulted primarily from an endogenous LH surge. This study clearly defines time-course features of serum LH in PMSG-treated rats. The overall results indicate that, following superovulatory treatment with PMSG, the increased ovulatory response is primarily associated with PMSG-derived intrinsic gonadotropin, and that the recovery of immature or asynchronously mature oocytes at ovulation may reult from the circulatory alteration of LH activity characterized by an initial prolonged elevation of serum LH and its subsequent attenuation.

• Reproductive and Developmental Biology
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• v.30 no.2
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• pp.99-105
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• 2006

The growing oocytes become progressively capable of resuming meiosis, and full meiotic competence appear when they are about 80% of the size of fully grown oocytes. As hormonal influences vary at different stages of reproductive cycle, the size of oocytes may vary according to the reproductive stages. The present study was designed to compare the diameter between the ovulated and freshly collected immature canine oocytes. The ovulated oocytes were collected 72 hr after ovulation by oviductal tube flushing by laparotomy under general anesthesia. Immature oocytes were collected by ovarian slicing method. Diameter of all oocytes was measured directly using epiflurescence microscope with a calibrated micro-eyepiece micrometer at ${\times}200$ magnification. The thickness of zona pellucida and diameter of cytoplasm were measured separately and recorded. A total of 2209 zona intact oocytes were collected, among them 628 from anestrus, 675 from follicular, 838 from luteal and 68 by fallopian tubes flushing methods. The average number of oocytes was 104.7, 168.8, 119.7 and 11.3 for anestrus, follicular, luteal and fallopian tubes flushing methods, respectively. The average diameters of the ooplasm and oocyte were significantly varied in different reproductive stages as well as with ovulated oocytes (P<0.05). The average diameter of ooplasm and oocyte was 115.6 and 127.7, 143.0 and 162.0, 134.6 and 150.6, 159.6 and 185.6 for anestrus, follicular, luteal and ovulated oocytes, respectively. Highest number of oocytes with larger diameter could be collected from the follicular and luteal stages. In conclusion, the follicular and luteal ovaries are the best sources of oocytes for canine IVM.

• The Korean Journal of Zoology
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• v.34 no.2
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• pp.181-187
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• 1991

We previously showed that transient exposure of Rana dybowskii follicles to exogenous cAMP in vitro could induce meiotic maturation. The present experiments were carried out to acertain whether the exogenous cAMP penetrate into the follicles. Isolated follicles were precultured in the medium containing cAMP (2.5 mM) for 6 hours and then cultured further in plain medium for 18 hours. The change of intrafollicular cAMP levels during the culture period were examined by utilizing cAMP radioimmunoassay (RIA). The intrafollicular levels of cAMP increased about thirty times of the basal level (about 3 p mole/follicle) in two hours and reached a peak in six hours (170 p mole/follicle) during the preculture period. However, when the follicles were transferred to plain medium, the levels decreased markedly in six hours to very low levels (about 10 p mole/ follicle), and kept the same levels thereafter. But the levels did not decrease to the basal levels. The increase and decrease of the intrafollicular cAMP was not affected by the presence of isobutyl methylxanthine (IBMX) or progesterone. The data suggest that exogenous cAMP pene-trate into the follicles and the cAMP accumulated by the follicles are degraded very rapidly.

• Korean Journal of Poultry Science
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• v.23 no.1
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• pp.1-7
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• 1996

In order to produce polyploid quail, the patterns of spermatogenesis and induction of diploid spermatozoa were analyzed by administration of spindle fiber inhibitor agent. Colcemid at the dose level of 37 $\mu\textrm{g}$ /100 g BW was Injected intraperitoneally to 50 Japanese quail males for 3 consecutive days. Five to 20 days after the first colcemid injection, the metaphase spreads from mitotic spermatogonia, primary spermatocyte and secondary spermatocyte were observed. By cytogenetic analysis, 9.4％ of spermatogonia and spermatocyte cells in germ cells from the treated males was found to be polyploid cells. As compared with colcemld treated, the males with non-treated colcemid had only 2.3％ polyploid cells in germ cells. The induction of diploid germ cells was highest in 10 days after the first colcemid injection and was lowest in 5 days after the first colcemid injection. These results suggested that between 10 to 15 days before maturation of the spermatozoa, the male germ cells were most sensitive to colcemid treatment. Spindle fiber inhibitor agent was also more sensitive to mitotic division of spermatogonia than meiotic division of primary and secondary spermatocyte.

• Clinical and Experimental Reproductive Medicine
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• v.26 no.3
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• pp.407-417
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• 1999

Objective: Mammalian follicle cells are the most important somatic cells which help oocytes grow, mature and ovulate and thus are believed to provide oocytes with various functional and structural components. In the present study we have examined whether cumulus or granulosa cells might playa role in establishing the plasma membrane structure of mouse oocytes during meiotic maturation. Design: In particular the differential resistances of mouse oocytes against chymotrypsin treatment were examined following culture with or without cumulus or granulosa cells, or in these cell-conditioned media. Results: When mouse denuded oocytes, freed from their surrounding cumulus cells, were cultured in vitro for $17{\sim}18hr$ and then treated with 1% chymotrypsin, half of the oocytes underwent degeneration within 37.5 min ($t_{50}=37.5{\pm}7.5min$) after the treatment. In contrast cumulus-enclosed oocytes showed $t_{50}=207.0$. Similarly, when oocytes were co-cultured with cumulus cells which were not associated with the oocytes but present in the same medium, the $t_{50}$ of co-cultured oocytes was $177.5{\pm}13.1min$. Furthermore, when oocytes were cultured in the cumulus cell-conditioned medium, $t_{50}$ of these oocytes was $190.0{\pm}10.8min$ whereas $t_{50}$ of the oocytes cultured in M16 alone was $25.5{\pm}2.9min$. Granulosa cell-conditioned medium also increased the resistance of oocytes against chymotrypsin treatment such that $t_{50}$ of oocytes cultured in granulosa cell-conditioned medium was $152.5{\pm}19.0min$ while that of oocytes cultured in M16 alone was $70.0{\pm}8.2min$. To see what molecular components of follicle cell-conditioned medium are involved in the above effects, the granulosa cell-conditioned medium was separated into two fractions by using Microcon-10 membrane filter having a 10 kDa cut-off range. When denuded oocytes were cultured in medium containing the retentate, $t_{50}$ of the oocytes was $70.0{\pm}10.5min$. In contrast, $t_{50}$ of the denuded oocytes cultured in medium containing the filtrate was $142.0{\pm}26.5min$. $T_{50}$ of denuded oocytes cultured in medium containing both retentate and filtrate was $188.0{\pm}13.6min$. However, $t_{50}$ of denuded oocytes cultured in M16 alone was $70.0{\pm}11.0min$ and that of oocytes cultured in whole granulosa cell-conditioned medium was $156.0{\pm}27.9min$. When surface membrane proteins of oocytes were electrophoretically analyzed, no difference was found between the protein profiles of oocytes cultured in M16 alone and of those cultured in the filtrate. Conclusions: Based upon these results, it is concluded that mouse follicle cells secrete a factor(s) which enhance the resistance of mouse oocytes against a proteolytic enzyme treatment. The factor appears to be a small molecules having a molecular weight less than 10 kDa.