金乌贼精子释放及精子活力的研究
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摘要
以金乌贼(Sepia esculenta)精荚为研究对象,探究了组织蛋白提取液对其中精子释放的影响。实验将活体解剖取出的成熟精荚置于5种组织蛋白提取液中,于19和4℃条件下观察并记录精荚保持完整形态时间及其中精子活力水平。并于19~24℃6个水温梯度下探讨了维持精子活力的最适温度范围。研究表明,水温19时,卵巢蛋白提取液可促进精荚中精子释放(9.5h),而副性腺蛋白、精荚囊蛋白、精巢蛋白和缠卵腺蛋白提取液均对精子释放的影响较小,离体后的精荚可保持完整形态1~2d。水温4℃时,放置于海水中的精荚,其完整形态和部分精子活力可维持较长时间(25和22d),而其他组织蛋白液中保存的精荚2~3d后即无完整个体,精荚中的精子1d后亦无活性。在19~24℃水温范围内,精子激烈运动时间和寿命均随水温升高呈先升高后降低的趋势。在20、21和22℃海水中,精子激烈运动时间分别为(93.33±13.33)、(116.67±6.67)和(93.33±6.67)min,组间差异不显著(P>0.05),但高于23和24℃实验组,并显著高于19℃组(43.33±12.02)min(P<0.05),在水温21℃时精子寿命达到最大值,为(206.67±6.67)min。通过拟合精子激烈运动时间y和温度x的关系曲线,得出当x=21.49℃时,ymax21.49=100.47min。在90%置信区间内,精子最大激烈运动时间的温度范围为20.22~22.76℃。研究结果表明,卵巢蛋白提取液可促进金乌贼精荚中精子释放,保持精子活力的适宜温度为20.22~22.76℃。
The present study was conducted to explore the effect of tissue proteins on the spermiation of Sepiaesculenta.Mature spermatophores were dissected from live adults and put in five tissue proteins.The time of maintaining integrated spermatophores and sperm motility were recorded at 19℃and 4℃.Optimum temperature range was also detected,which was from 19℃to 24℃.Results showed that spermiation was promoted by ovary protein earlier(9.5 h)than other proteins at 19 ℃.Accessory gland,spermatophore,testis and nidamental gland protein had little impact on spermiation(1~4 d).When the water temperature was maintained at 4℃,the morphological integrity of spermatophores stored in sea water and the motility of a portion of sperms(22~25 d)were higher than those of other treatment groups(2~3 d,1 d).Sperm lifespan and acute movement time increased first and decreased then as the temperature increased from 19 ℃to 24 ℃.There was no significant difference among acute movement time of sperms in treatment groups of 20℃,21℃and 22℃((93.33±13.33)min,(116.67±6.67)min and(93.33±6.67)min,respectively)(P>0.05),which was higher than that at 23℃and 24℃.However,they were significantly higher than that at 19 ℃((43.33±12.02)min)(P<0.05).The longest lifespan was206.67±6.67 min when the temperature was 21℃.The optimum temperature and corresponding maximum acute movement time were xopt=21.49℃and ymax21.49=100.47 min through curve fitting the acute movement time(y,in min)and temperature(x,in℃).When the most acute movement was maintained,the 90%confidence interval for temperature was between 20.22℃and 22.76℃.In conclusion,spermiation was promoted by ovary protein and temperatures ranging from 20.22℃to 22.76℃ were the optimum temperatures for sperm motility.
The present study was conducted to explore the effect of tissue proteins on the spermiation of Sepiaesculenta.Mature spermatophores were dissected from live adults and put in five tissue proteins.The time of maintaining integrated spermatophores and sperm motility were recorded at 19℃and 4℃.Optimum temperature range was also detected,which was from 19℃to 24℃.Results showed that spermiation was promoted by ovary protein earlier(9.5 h)than other proteins at 19 ℃.Accessory gland,spermatophore,testis and nidamental gland protein had little impact on spermiation(1~4 d).When the water temperature was maintained at 4℃,the morphological integrity of spermatophores stored in sea water and the motility of a portion of sperms(22~25 d)were higher than those of other treatment groups(2~3 d,1 d).Sperm lifespan and acute movement time increased first and decreased then as the temperature increased from 19 ℃to 24 ℃.There was no significant difference among acute movement time of sperms in treatment groups of 20℃,21℃and 22℃((93.33±13.33)min,(116.67±6.67)min and(93.33±6.67)min,respectively)(P>0.05),which was higher than that at 23℃and 24℃.However,they were significantly higher than that at 19 ℃((43.33±12.02)min)(P<0.05).The longest lifespan was206.67±6.67 min when the temperature was 21℃.The optimum temperature and corresponding maximum acute movement time were xopt=21.49℃and ymax21.49=100.47 min through curve fitting the acute movement time(y,in min)and temperature(x,in℃).When the most acute movement was maintained,the 90%confidence interval for temperature was between 20.22℃and 22.76℃.In conclusion,spermiation was promoted by ovary protein and temperatures ranging from 20.22℃to 22.76℃ were the optimum temperatures for sperm motility.
引文
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[9]孟祥黔,张德福.猪精液超低温保存技术及其对养猪业未来的重要作用[J].国外畜牧学:猪与禽,2009(3):42-44.Meng X Q,Zhang D F.The important role of cryopreservation of pig sperm in pig industry[J].Foreign Animal Husbandry:Pigs and Poultry,2009(3):42-44.
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[12]颜跃弟,傅洪拓,乔慧,等.酶解制备日本沼虾游离精子方法的研究[J].中国农学通报,2015,31(2):107-111.Yan YD,Fu HT,Qi H,et al.Trypsinase digestion in acquiring free sperms from Macrobrachium nipponensis[J].Chinese Agricultural Science Bulletin,2015,31(2):107-111.
[13] Takami Y.Mating behavior,insemination and sperm transfer in the ground beetle Carabus insulicola[J].Zoological Science,2002,19(9):1067-1073.
[14] Uma K,Subramoniam T.Histochemical characteristics of spermatophore layers of Scylla serrata(Forskal)(Decapoda:Portunidae)[J].International Journal of Invertebrate Reproduction,1979,1(1):31-40.
[15]袁路,蔡生力.温度,盐度对凡纳滨对虾精荚再生和精子质量的影响[J].水产学报,2006,30(1):63-68.Yuan L,Cai S L.Effect of temperature and salinity on spermatophore regeneration and sperm quality in Litopenaeus vannamei[J].Journal of Fisheries of China,2006,30(1):63-68.
[16]赵会宏,刘晓春,林浩然,等.斜带石斑鱼精子超微结构及盐度,温度,pH对精子活力及寿命的影响[J].中国水产科学,2003,10(4):286-292.Zhao H H,Liu X C,Lin H R,et al.Ultrastructure of spermatozoa and effects of salinity,temperature and pH on spermatozoa motility in Epiniphelus coioides[J].Journal of Fishery Sciences of China,2003,10(4):286-292.
[17]刘鉴毅,黄晓荣,章龙珍.盐度,温度和pH对2种篮子鱼精子活性的影响[J].海洋渔业,2014,36(6):523.Liu J Y,Huang X R,Zhang L Z.Effects of salinity,temperature and pH on spermatozoa motility in two kinds of rabbitfish[J].Marine Fisheries,2014,36(6):523.
[18]谢恩义,何学福.瓣结鱼的繁殖习性以及精子的活力与寿命[J].动物学杂志,1999,34(2):5-8.Xie E Y,He X F.Reproductive habit in Folifer brevifilis and sperm motility[J].Chinese Journal of Zoology,1999,34(2):5-8.
[19]张涛,颜世伟,章龙珍,等.紫外线辐射对西伯利亚鲟精子活力和寿命的影响[J].应用生态学报,2011,22(8):2179-2183.Zhang T,Yan S W,Zhang L Z,et al.Impacts of ultraviolet irradiation on the sperm motility and longevity of Acipenser baerii[J].Chinese Journal of Applied Ecology,2011,22(8):2179-2183.
[20]陈昭廷,李琪,陈四清,等.海月水母精巢发育及排精过程的观察[J].动物学杂志,2015,50(1):131-140.Chen Z T,Li Q,Chen S Q,et al.The testis development and spermiation process in moon jellyfish[J].Chinese Journal of Zoology,2015,50(1):131-140.
[21]苏新有.玛丽鱼(Poecilia latipinna)精子的超低温冷冻保存[D].汕头:汕头大学,2007.Su XY.Cryopreservation of Molly Fish(Poecilia latipinna)Sperm[D].Shantou:Shantou University,2007.
[2]韩松,郑小东,李琪,等.金乌贼(Sepia esculenta)精子超微结构和低温损伤电镜观察[J].海洋与湖沼,2010(4):483-488.Han S,Zheng X D,Li Q,et al.Ultrastructure of Sepia esculenta(Cephalopoda:Sepiida)spermatozoa and its damage by low temperature[J].Oceanologia et Limnologia Sinica,2010(4):483-488.
[3]宋训民.金乌贼(Sepia ecsulenta Hoyle)精荚器的结构和精荚的生成[J].山东大学学报,1963(3):78-91.Song X M.The structure of spermatheca and generation of spermatophore in Sepia ecsulenta Hoyle[J].Journal of Shandong University,1963(3):78-91.
[4]曲迪.中华绒螯蟹精荚破裂以及精子顶体酶活力研究[D].上海:华东师范大学,2007:1-38.Qu D.Studies on Spermathophore Dissolution and Sperm Acrosin Activity of Eriocheir sinensis[D].Shanghai:East China Normal University,2007:1-38.
[5]李嘉泳.金乌贼(Sepia esculenta Hoyle)在黄渤海的结群生殖和洄游[J].山东海洋学院学报,1963,5(2):69-108.Li J Y.Breeding and migration in the Bohai Sea and the Yellow Sea of Sepia esculenta Hoyle[J].Journal of Shandong College of Oceanography,1963,5(2):69-108.
[6]何璐,虞泓,袁理春,等.生物温度适应的分子生态学研究进展[J].亚热带农业研究,2014,10(3):205-211.He L,Yu H,Yuan L C,et al.Advances in molecular ecology of biological temperature adaptation[J].Subtropical Agriculture Research,2014,10(3):205-211.
[7]刘石林,茹小尚,徐勤增,等.高温胁迫对刺参耐高温群体和普通群体主要免疫酶活力的影响[J].中国水产科学,2016(2):344-351.Liu S L,Ru X S,Xu Q Z,et al.Effects of high-temperature stress on several immune enzyme activities of Apostichopus japonicus thermotolerant and normal species[J].Journal of Fishery Sciences of China,2016(2):344-351.
[8] Drobnis E Z,Crowe L M,Berger T,et al.Cold shock damage is due to lipid phase transitions in cell membranes:A demonstration using sperm as a model[J].Journal of Experimental Zoology Part A:Ecological Genetics and Physiology,1993,265(4):432-437.
[9]孟祥黔,张德福.猪精液超低温保存技术及其对养猪业未来的重要作用[J].国外畜牧学:猪与禽,2009(3):42-44.Meng X Q,Zhang D F.The important role of cryopreservation of pig sperm in pig industry[J].Foreign Animal Husbandry:Pigs and Poultry,2009(3):42-44.
[10] Cowey C B,Cooke D J,Matty A J,et al.Effects of quantity and quality of dietary protein on certain enzyme activities in rainbow trout[J].The Journal of Nutrition,1981,111(2):336-345.
[11] Wang J,Fang D A,Wang Y,et al.Cathepsin A protein from the accessory sex gland of the Chinese mitten crab(Eriocheir sinensis)plays a key role in spermatophore digestion[J].Journal of Insect Physiology,2013,59(9):953-960.
[12]颜跃弟,傅洪拓,乔慧,等.酶解制备日本沼虾游离精子方法的研究[J].中国农学通报,2015,31(2):107-111.Yan YD,Fu HT,Qi H,et al.Trypsinase digestion in acquiring free sperms from Macrobrachium nipponensis[J].Chinese Agricultural Science Bulletin,2015,31(2):107-111.
[13] Takami Y.Mating behavior,insemination and sperm transfer in the ground beetle Carabus insulicola[J].Zoological Science,2002,19(9):1067-1073.
[14] Uma K,Subramoniam T.Histochemical characteristics of spermatophore layers of Scylla serrata(Forskal)(Decapoda:Portunidae)[J].International Journal of Invertebrate Reproduction,1979,1(1):31-40.
[15]袁路,蔡生力.温度,盐度对凡纳滨对虾精荚再生和精子质量的影响[J].水产学报,2006,30(1):63-68.Yuan L,Cai S L.Effect of temperature and salinity on spermatophore regeneration and sperm quality in Litopenaeus vannamei[J].Journal of Fisheries of China,2006,30(1):63-68.
[16]赵会宏,刘晓春,林浩然,等.斜带石斑鱼精子超微结构及盐度,温度,pH对精子活力及寿命的影响[J].中国水产科学,2003,10(4):286-292.Zhao H H,Liu X C,Lin H R,et al.Ultrastructure of spermatozoa and effects of salinity,temperature and pH on spermatozoa motility in Epiniphelus coioides[J].Journal of Fishery Sciences of China,2003,10(4):286-292.
[17]刘鉴毅,黄晓荣,章龙珍.盐度,温度和pH对2种篮子鱼精子活性的影响[J].海洋渔业,2014,36(6):523.Liu J Y,Huang X R,Zhang L Z.Effects of salinity,temperature and pH on spermatozoa motility in two kinds of rabbitfish[J].Marine Fisheries,2014,36(6):523.
[18]谢恩义,何学福.瓣结鱼的繁殖习性以及精子的活力与寿命[J].动物学杂志,1999,34(2):5-8.Xie E Y,He X F.Reproductive habit in Folifer brevifilis and sperm motility[J].Chinese Journal of Zoology,1999,34(2):5-8.
[19]张涛,颜世伟,章龙珍,等.紫外线辐射对西伯利亚鲟精子活力和寿命的影响[J].应用生态学报,2011,22(8):2179-2183.Zhang T,Yan S W,Zhang L Z,et al.Impacts of ultraviolet irradiation on the sperm motility and longevity of Acipenser baerii[J].Chinese Journal of Applied Ecology,2011,22(8):2179-2183.
[20]陈昭廷,李琪,陈四清,等.海月水母精巢发育及排精过程的观察[J].动物学杂志,2015,50(1):131-140.Chen Z T,Li Q,Chen S Q,et al.The testis development and spermiation process in moon jellyfish[J].Chinese Journal of Zoology,2015,50(1):131-140.
[21]苏新有.玛丽鱼(Poecilia latipinna)精子的超低温冷冻保存[D].汕头:汕头大学,2007.Su XY.Cryopreservation of Molly Fish(Poecilia latipinna)Sperm[D].Shantou:Shantou University,2007.