• Korean Journal of Animal Reproduction
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• v.14 no.4
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• pp.237-244
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• 1990

These studies were carried out to investigate the effects of the follicles size, hormone supplementation, semen types and capacitation methods on in vitro maturation and fertilization of bovine follicular oocytes. The ovaries were obtained from slaughtered Korean Native cows. The follicular oocytes surrounded with cumulus cells were recovered by aspirating follicular fluid from the visible follicles of diameter 3~5mm. The follicular oocytes were cultured in TCM-199 medium containing hormones, FCS, ECS, BFF and MCC for 24~48hrs. in a incubator with 5% CO2 in air at 38.5$^{\circ}C$ and then matured oocytes were again cultured for 18~20hrs. with motile capacitated sperm in the TCF(Tyroide calcium-free) solution containing 200$\mu\textrm{g}$/ml of heparin. The results obtained in these experiments were summarized as follows : 1. The oocytes classified as "A, B, C, D and Degenerative" depending morphological integrity and those were 61.4%, 12.1%, 19.2%, 4.2% and 3.0% of the total oocytes recovered, respectively. The maturation and fertilization rate of the A, B, C class follicular oocytes, cultured in the TCM-199 medium supplemented with 10% FCS were 89.1%, 78.0%, 52.6% and 78.1%, 66.1%, 33.3%, respectively. 2. The average number of the follicular oocytes recovered from follicles size, 1~2mm, 3~5mm and above 5mm in dimeter were 67, 98 and 63, respectively. The maturation and fertilization rate of the follicular oocytes, cultured in the TCM-199 medium were 56.7%, 82.5%, 46.0% and 44.8%, 71.4%, 28.6%, respectively. 3. The maturation and fertilization rate of follicular oocytes, cultured in the TCM-199 medium supplemented with 5%, 10%, 15%, 20% FCS and hCG, HCG, $\beta$-estradiol were 76.0%~82.3% and 26.2%~70.0%, and those values were higher the supplementation of the hormone than the non-supplementation. 4. The fertilization and cleavage rate of the follicular oocytes, inseminated with spermatozoas of epididymis cauda, neat and frozen semen were 63.3%, 73.3%, 70.0% and 32.7%, 37.8%, 38.3, respectively. 5. The fertilization and cleavage rate of follicular oocytes, fertilized with capacitated spermatozoas by heparin, BFF and HIS methods were 70.0%, 53.8%, 34.2% and 38.3%, 23.1%, 17.1%, respectively. And the fertilization and cleavage rate were higher method of heparin than other methods.r methods.

• Journal of Embryo Transfer
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• v.16 no.2
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• pp.79-89
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• 2001

Steroid hormones control the expression of many cellular regulators, and a role thor estrogen in mouse oocytes has been well documented. The preovulatory $E_2$increment is generally accepted as the endocrine process regulating induction of in vivo oocyte maturation To address whether the activity of the T-type $Ca^{2+}$ channel is altered by 17 beta-estradiol ( $E_2$), we examined the actions of $E_2$on the calcium channel of mouse oocytes and early embryos. Oocrtes were collected from the oviduct of mice treated with pregnant mare's serum gonadotropin (PMSG) and human choronic gonadotropin (hCG). Whole cell voltage clamp technique and confocal microscopy were used to examine that $E_2$increase intracellular $Ca^{2+}$ concentration ([C $a^{2+}$]$_{i}$ ) via voltage dependent $Ca^{2+}$ channel (VDC) and estrogen receptor (FSR), and $E_2$concentration by the use of radioimmunoassay (RIA) were examined in mouse. The results obtained were as follows: The peak of $Ca^{2+}$ current induced by $E_2$increased 122% to 1.50$\pm$0.03 nA from 1.23$\pm$0.21 nA (n=15) in the presence of 5 mM extracellular $Ca^{2+}$ concentration ([C $a^{2+}$]$_{o}$ ). The increased $Ca^{2+}$ current was temporally associated with $Ca^{2+}$ transients. The intracellular $Ca^{2+}$ level increased 207%~30 s following the addition of 1${\mu}{\textrm}{m}$ $E_2$(relative fluorescence intensity: 836.4$\pm$131.2 for control, n=10, 1736.4$\pm$192.0 in the presence of $E_2$, n=10). $E_2$increased amplitude of $Ca^{2+}$ current and [C $a^{2+}$]$_{i}$ . $E_2$-induced $Ca^{2+}$ current and $E_2$concentration in blood were showed difference on the stage of embryo. These results suggest that $E_2$modulate $Ca^{2+}$ channel to increase $Ca^{2+}$ influx.$Ca^{2+}$ influx.