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KMID : 0359320020420010035
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Korean Journal of Veterinary Research
2002 Volume.42 No. 1 p.35 ~ p.43
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Calcium current on cryopreservation in mouse oocytes
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강다원/Kang, Da Won
김은심/최창용/박재용/한재희/홍성근/Kim, Eun Sim/Choe, Chang Yong/Park, Jae Yong/Han, Jae Hee/Hong, Seong Geun
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Abstract
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Cryopreservation is commonly used for an efficient utilization of semen, oocytes and embryos but has disadvantage in the survival, development of the post-thawed eggs. The high risk in the survival, development of eggs after thawing is thought to be caused by inappropriate internal regulation of Ca^2+ and/or formation of intracellular ice crystals. In this experiment, we tested whether the Ca^2+ current (iCa), a decisive factor to Ca^2+ entry, was altered in post-thawed oocytes by using whole cell voltage clamp technique.
The quality and survival rates of the oocytes derived from both fresh and frozen groups were examined by morphology and FDA-test. Vitrified oocytes (VOs) were incubated for 4 hr after thawing and then donated to this experiment. Ethyleneglycol-ficoll-galactose (EFG) was used as a cryoprotectant for vitrification. The membrane potential was held at -80 mV and step depolarizations of 250 ㎳ were applied from -50 mV to 50 mV in 10 mV increments.
The survival rates showed a higher in VOs vitrified with EFG containing Ca^2+ than in VOs vitrified with EFG under the Ca^2+ -free condition (82.0% vs 14%). In group with/without Ca^2+, the survival rates were significantly (P＜0.01) difference. In the fresh metaphase Ⅱ oocytes (FOs), current-voltage (Ⅰ-Ⅴ) relationship showed that iCa began to activate at -40 mV and reached its maximum at -10 mV. With same voltage pulses, inward currents were elicited in VOs. Ⅰ-Ⅴ relationships observed in VOs were similar to those in FOs. Time constants of activation and inactivation of the inward current shown in VOs were not different to those in FOs. This accordance in Ⅰ-Ⅴ relations and time constants in FOs with those in VOs indicates that the inward currents in FOs are unaltered by vitrification and thawing. Therefore, vitrification with EFG does not play as a factor to deteriorate Ca^2+ entry across the membrane of the oocytes.
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