Journal
ANIMAL SCIENCE JOURNAL
Volume 82, Issue 3, Pages 412-419Publisher
WILEY-BLACKWELL
DOI: 10.1111/j.1740-0929.2010.00856.x
Keywords
artificial insemination; cryopreservation; EGTA; pig; sperm
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Funding
- [12-068]
- [12-104]
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In our previous study, seminal plasma effectively suppressed the induction of sperm to capacitation-like status and acrosome loss during the thawing process. However, because boar seminal plasma is contaminated with various kinds of bacteria and/or viruses, it is necessary to develop a thawing solution without animal-derived materials. In this study, we focused on the increase of intracellular Ca2+ ([Ca2+](i)) in sperm after thawing and the negative effects of sperm qualities. After thawing, the fluorescent intensity of [Ca2+](i) indicator, Fluo-3/AM, and the level of phosphorylated tyrosine residue of protein were increased in the sperm. Next, we investigated whether the addition of Ca2+ chelators (ethylenediaminetetraacetic acid (EDTA) and ethyleneglycoltetraacetic acid (EGTA)) improved post-thawed sperm motility. When the frozen-thawed sperm were treated with 6 mmol/L EDTA + 6 mmol/L EGTA, sperm motility was significantly increased as compared with control (6 mmol/L EDTA alone) at all incubation periods (P < 0.05). The combinational treatment significantly suppressed the elevation of [Ca2+](i) and the tyrosine phosphorylation, which improved the acrosomal status and fertilizing ability in vitro. Furthermore, when the thawing method was applied for artificial insemination, the fertilization rate was significantly higher than control (P < 0.05, 33% vs. 82%). Thus, we conclude that the addition of EDTA + EGTA to thawing solution is a beneficial tool for artificial insemination using frozen-thawed boar sperm.
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