Cryopreservation of dolphin spermatozoa

海豚精子的冷冻保存

• 批准号: 15580163
• 负责人: YOSHIOKA Motoi
• 金额: $ 1.47万
• 依托单位: Mie University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15580163/
• 关键词: dolphin; sperm; cryopreservation; artificial insemination; straw method; cetacean; marine mammals; reproductive physiology

In order to develop an effective and simple method for cryopreserving dolphin semen processed immediately after collection by the hand-manipulation method, we conducted some freezing and thawing experiments by straw method for fresh semen samples obtained from two bottlenose dolphins (Tursiops truncatus) and one Pacific white-sided dolphins (Lagenorhynchus obliquidens) in captivity. For both dolphin species, post-thaw sperm with total motility about 60% or more, percentage progressive motility over 95% and kinetic rating 4 to 5 (rapid forward progressive movement) were obtained by the straw method, with semen extender containing 2.9% trisodium citrate hydrate, 8% glycerol, 20% egg-yolk and antibiotics. Fresh semen was diluted twice with the semen extender to the final dilution 1:2 (semen : extender, v/v) and precryopreserved at 4 degrees C (refrigerator;1-1.5 hr) at the first dilution and then at 2 degree C (ice-water bath;0.25-1hr) at the second dilution. About 300-400 micro litters of extended semen put into 0.5mL-straw were frozen in the liquid nitrogen vapor for 15 min. and then straws were plunged into liquid nitrogen for complete freezing. Rapid thawing in the water bath at 35-36 degrees C led into better post-thaw semen characteristics. Post-thaw sperm frozen by the present straw method was used for the artificial insemination(AI) to a female bottlenose dolphin in September 2003,and a male calf was born as the second AI dolphin baby (the first with frozen-thawed sperm) in Japan in September 2004,

为了探索一种简单有效的海豚精液冷冻保存方法，对人工饲养的两条宽吻海豚(Tursioptruncatus)和一条太平洋白边海豚(Lagnowhynchus Obliquidens)的新鲜精液进行了吸管法冷冻和解冻试验。两种海豚精液解冻后，用含2.9%柠檬酸三钠、8%甘油、20%蛋黄和抗生素的吸管法精液解冻后，精子的总活动率在60%左右，前向运动百分率在95%以上，运动能力分级为4~5级(快速前进运动)。新鲜精液用精液稀释液稀释两次，最终稀释度为1：2(精液：稀释液，v/v)，第一次稀释液在4℃(冰箱；1-1.5小时)预冻，第二次稀释液在2℃(冰水浴；0.25-1小时)预冻。将大约300-400微升的延伸精液放入0.5ml的吸管中，在液氮蒸气中冷冻15分钟。然后将稻草放入液氮中完全冷冻。在35-36摄氏度的水浴中快速解冻可以获得更好的解冻后精液特性。2003年9月，采用目前管道法冷冻的解冻精子对雌性宽吻海豚进行人工授精，2004年9月日本出生了第二只雄性海豚幼崽(第一只冷冻解冻精子)，

项目成果

Artificial insemination using frozen-thawed semen in the Pacific white-sided dolphin (Lagenorhynchus obliquidens).

使用太平洋白海豚（Lagenorhynchus obliquidens）冻融精液进行人工授精。

• 发表时间: 2003
• 期刊: Proceedings for the 34th Annual Conference of the International Association for Aquatice Animal Medicine, May 9-14,2003, Waikoloa, Hawaii
• 作者: Robeck.T.R.;Greenwell;M.;Boehm;J.R.;Yoshioka;M.;Tobayama;T.;Steinman;K.;Monfort;S.L.
• 通讯作者: S.L.

Estrous cycle characterization and artificial insemination using frozen-thawed spermatozoa in the bottlenose dolphin

宽吻海豚的发情周期特征和冻融精子人工授精

• 发表时间: 2005
• 期刊: Reproduction (In press)
• 作者: Robeck T.R.;Steinman;K.J.;Yoshioka;M.ほか
• 通讯作者: M.ほか

Estrous cycle characterization and artificial insemination using frozen-thawed spermatozoa in the bottlenose dolphin (Tursiops truncatus).

使用宽吻海豚（Tursiops truncatus）冻融精子进行发情周期特征和人工授精。

• 发表时间: 2005
• 期刊: Reproductin 129
• 作者: Robeck T.R.;Steinman;K.J.;Yoshioka;M.;Jensen;E.;O'Brien;J.K.;Katsumata;E.;Gili;C.;McBain;J.F.;Sweeney;J.;Monfort;S.L.
• 通讯作者: S.L.

Estrous cycle characterization and artificial insemination using, frozen-thawed spermatozoa in the bottlenose dolphin

宽吻海豚的发情周期特征和使用冻融精子的人工授精

• 发表时间: 2005
• 期刊: Reproduction 129
• 作者: Robeck T.R.;Steinman;K.J.;Yoshioka;M.ほか
• 通讯作者: M.ほか

Artificial insemination using frozen-thawed semen in the Pacific white-sided dolphin (Lagenorhynchus obliquidens)

使用太平洋白海豚（Lagenorhynchus obliquidens）冻融精液进行人工授精

• 发表时间: 2003
• 期刊: Proceedings for the 34th Annual Conference of the International Association for Aquatice Animal Medicine
• 作者: Robeck;T.R.;Greenwell;M.;Boehm;J.R.;Yoshioka;M.ほか
• 通讯作者: M.ほか