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1.
超低温冷冻对日本鳗鲡精子酶活性的影响 总被引：2，自引：1，他引：1
研究超低温冷冻保存(-196℃)对日本鳗鲡精子内总ATP酶、肌酸激酶(CK)、琥珀酸脱氢酶(SDH)、乳酸脱氢酶(LDH)、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、谷胱甘肽还原酶(GR)等酶活性的影响。运用试剂盒分别测定了冷冻前后日本鳗鲡精子内酶活性的变化。结果表明,经过超低温冷冻保存后,日本鳗鲡精子的活力下降,精子内GR活性显著升高(P<0.05),酶活性从冻前的358.52±45.65 U/L上升到646.30±70.30 U/L;其它几种酶的活性均显著下降(P<0.05),总ATP酶、CK和SDH的活性分别从冻前的3.14±0.61 U/ml、17.10±3.51 U/ml和32±5.94 U/ml下降到1.83±0.43 U/ml、7.33±1.74 U/ml和21±1.41 U/ml,LDH、SOD和CAT活性分别从冻前的2 266.67±313.25 U/L、220.47±32.94 U/ml和48.51±5.94U/ml下降到1 195.91±198.51 U/L、84.16±22.11 U/ml和21.8±4.14 U/ml。超低温冷冻对日本鳗鲡精子酶活性和精子活力均有较大影响。 相似文献
2.
超低温冷冻对斑尾刺虾虎鱼卵中酶活性的影响 总被引：1，自引：0，他引：1
研究了超低温冷冻保存(-196℃)对斑尾刺虾虎鱼成熟卵中总ATP酶、肌酸激酶(CK)、琥珀酸脱氢酶(SDH)、乳酸脱氢酶(LDH)、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、谷胱甘肽过氧化物酶(GSH-PX)和丙二醛(MDA)等酶活性的影响。结果表明,经过冷冻保存后,总ATP酶、CK、SDH和LDH的活性显著下降(P<0.05),同时可见添加抗冻剂组比未添加抗冻剂组下降幅度更大(P<0.05);SOD、CAT和GSH-PX的活性也显著下降(P<0.05),未添加抗冻剂组比添加抗冻剂组下降更明显(P<0.05);MDA的活性显著升高,且未添加抗冻剂组显著高于添加抗冻剂组(P<0.05)。超低温冷冻和是否添加抗冻剂都对斑尾刺虾虎鱼卵中酶活性有显著性影响。 相似文献
3.
【目的】本文旨在探究长期超低温冷冻保存中鞍带石斑鱼精子质膜、活力、超微结构及酶活性的变化，为阐明影响鞍带石斑鱼精子冷冻保存质量的相关机制提供理论依据。【方法】采集2022年鞍带石斑鱼鲜精及储存时间分别为23、49、61个月冷冻保存精液，用伊红-苯胺黑染色方法检测精子质膜完整性；用计算机辅助精子分析仪（CASA）检测精子运动参数；测量精浆和精子中琥珀酸脱氢酶（SDH）、过氧化氢酶（CAT）、谷胱甘肽还原酶（GR）、总超氧化物歧化酶（T-SOD）、谷胱甘肽过氧化物酶（GSH-PX）和肌酸激酶（CK）共六种酶活性的变化及三磷酸腺苷（ATP）浓度变化；用扫描电镜和透射电镜观察鲜精和冻精超微结构。【结果】伊红-苯胺黑染色检测结果发现，鲜精质膜完整性最高为83.43±2.73 %，经过超低温冷冻后，精子质膜完整性显著降低（P<0.05），且随着冷冻保存时间的延长而逐渐降低。CASA结果显示鲜精活力最高为90.47±3.34 %，经过超低温冷冻后精子活力显著降低（P<0.05），但长期保存23-61个月精子活力无显著性差异，精子活力保持在63.95±3.66 %-68.58±2.73 %，具有稳定的活力，且鲜精与冻精之间精子平均直线运动速度（VSL）、平均曲线运动速度（VCL）和平均路径速度（VAP）均没有显著差异（P>0.05）。精子超微结构显示，鲜精形态结构正常、线粒体排列结构规则、形态大小正常。经过超低温冷冻保存后，精子形态结构损伤明显，表现为精子头部质膜破损、细胞质外漏、细胞核膜破损、尾部鞭毛断裂或脱落等损伤；鞍带石斑鱼精浆和精子超低温冷冻前后六种酶活性的变化及ATP含量结果显示，经过超低温冷冻后，精子内SOD、GSH-PX和CAT三种酶及ATP含量均有显著性降低（P<0.05）。精浆中酶活力升高，除GR和CAT外，其余酶活性均差异显著（P<0.05）。【结论】长期超低温冷冻对鞍带石斑鱼精浆和精子酶活性、精子活力及精子超微结构均具有较显著影响，【意义】研究结果为鱼类精子冷冻损伤机理研究积累了丰富的数据，为鱼类精子长期冷冻保存提供了技术参考和评价指标。 相似文献
4.
研究了超低温冷冻保存(-196℃)对俄罗斯鲟(Acipenser gueldenstaedti)精浆和精子中总三磷酸腺苷酶(AT-Pase)、琥珀酸脱氢酶(SDH)、乳酸脱氢酶(LDH)、肌酸激酶(CK)等酶活性的影响,并分别测定了冷冻前后俄罗斯鲟精浆和精子中酶的活性。结果显示,经过超低温冷冻保存后,俄罗斯鲟精子活力下降,精子内各酶活性均显著降低,添加冷冻保护液组精子中总ATPase、SDH、LDH和CK的活性分别从(198.47±14.43)U/mL、(30.00±2.65)U/mL、(6 982.29±24.32)U/L和(1.94±0.05)U/mL下降至(110.19±2.32)U/mL、(16.33±2.08)U/mL、(5 122.93±195.07)U/L和(1.49±0.14)U/mL。未添加抗冻剂组则分别下降至(2.25±0.33)U/mL、(11.67±0.58)U/mL、(4 488.04±78.33)U/L和(1.16±0.02)U/mL;精浆中酶的活性均显著升高,添加冷冻保护液组精浆总ATPase、SDH、LDH和CK活性分别从(12.70±0.57)U/mL、(7.50±0.71)U/mL、(2017.26±116.81)U/L和(2.93±0.59)U/mL升高至(92.49±5.18)U/mL、(13.33±0.58)U/mL、(3 688.97±172.67)U/L和(4.39±0.24)U/mL,未添加抗冻剂组则分别上升至(200.27±12.97)U/mL、(24.67±3.06)U/mL、(6 124.40±329.14)U/L和(5.20±0.16)U/mL。结果表明,超低温冷冻对俄罗斯鲟精浆和精子中酶活性及精子活力均有较大影响。 相似文献
5.
为了解抗冻剂对施氏鲟(Acipenser schrenckii)精子冷冻保存效果的影响及其冷冻损伤机理,比较了添加不同浓度海藻糖和蔗糖作为冷冻保护剂的精子稀释液处理后施氏鲟精子冷冻复苏的活力、快速运动时间和寿命。结果表明,添加60 mmol·L-1海藻糖1.0 mmol·L-1氯化钾的稀释液处理后,精子冻后快速运动时间和寿命与鲜精相比无显著差异(P>0.05);且活力较其他处理组有所提高,达到(26.67±3.32)%,但仍显著低于鲜精(P<0.05)。利用透射电镜和扫描电镜对施氏鲟精子超低温冷冻保存前后的超微结构进行观察,并对其能量代谢酶和抗氧化酶活进行测定和比较,结果表明,低温冻存造成施氏鲟精子的膜系统和细胞器(主要为线粒体和轴丝)损伤;能量代谢酶[总ATP酶、肌酸激酶(CK)、琥珀酸脱氢酶(SDH)、乳酸脱氢酶(LDH)]和抗氧化酶[过氧化氢酶(CAT)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-PX)]活性均显著下降(P<0.05),而抗氧化酶谷胱甘肽还原酶(GR)活性显著上升(P<0.05),表明... 相似文献
6.
研究了超低温冷冻保存(-196℃)对脊尾白虾精子内琥珀酸脱氢酶(SDH)、乳酸脱氢酶(LDH)、Na+/K+-ATP酶、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、谷胱甘肽还原酶(GR)与顶体酶活性的影响,以期为提高脊尾白虾精子超低温冷冻效果提供理论依据.设置对照组(未添加抗冻剂)、3个实验组[分别添加DMSO (V/V) 10.0％、12.5％、15.0％],于冷冻0、1、3、5、7、15d取样测定各酶活性.结果表明,经冷冻后,除GR外,其他所测酶活性均出现显著下降(P＜0.05),且以对照组酶活性下降幅度最大.GR活性在冷冻7d内显著升高,且对照组明显高于实验组(P＜0.05),在冷冻15d时又出现下降.添加15.0％ DMSO组所测酶活性均高于同期其它各组,表明15.0％ DMSO对精子内酶的保护作用较好.冷冻15 d后15.0％ DMSO组的SDH、LDH和Na+/K+-ATP酶活性由(28.500±1.453) U/mL、(1290.836±27.603) U/L和(2.605-0.232) μmol/(mg·h)分别降至(15.300±0.950) U/mL、(363.713-13.943) U/L和(0.542-0.186) μmol/(mg.h);SOD和CAT活性由(106.497±7.217) U/mL、(383.632±4.731)U/g分别降至(17.036 ±0.321)U/mL、(166.940±1.910) U/g;顶体酶活性从(3.521±0.010)μIU/106降至(1.212±0.043)μIU/106;而GR活性由(217.042±6.962) U/L上升至(302.787±24.558)U/L.从冷冻后各酶下降幅度来看,超低温冷冻对SOD活性的影响最大,其次是Na+/K+-ATP酶. 相似文献
7.
超低温保存对罗氏沼虾胚胎几种酶活性的影响 总被引：1，自引：0，他引：1
研究了超低温冷冻保存(-196℃)对罗氏沼虾胚胎内总三磷酸腺苷酶(ATPase)、肌酸激酶(CK)、琥珀酸脱氢酶(SDH)、乳酸脱氢酶(LDH)、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、谷胱甘肽过氧化物酶(GSH-Px)、丙二醛(MDA)等酶活性的影响。运用试剂盒分别测定了冷冻前后罗氏沼虾胚胎内酶活性的变化。结果表明,经过超低温冷冻保存后,胚胎内7种酶的活性均显著下降(P0.05),其中总ATPase、CK、LDH和SDH的活性分别从冻前的1.322±0.162 U/mg prot、0.404±0.015 U/mg prot、352.225±23.214 U/gprot和2.067±0.139 U/mg prot下降到0.087±0.003 U/mg prot、0.010±0.002 U/mg prot、5.890±0.658 U/gprot和0.552±0.138 U/mg prot;SOD、CAT和GSH-Px的活性分别从冻前的19.217±0.677 U/mg prot、3.587±0.233 U/mg prot和7.626±1.106 U/(min.mg)下降至3.579±0.234 U/mg prot、1.773±0.227 U/mg prot和1.524±0.096 U/(min.mg);MDA的活性从1.015±0.038 n mol/mg prot上升到20.937±0.320 n mol/mgprot,超低温冷冻对罗氏沼虾胚胎酶活性有显著性影响。 相似文献
8.
人工养殖西伯利亚鲟精子超低温冷冻保存研究 总被引：7，自引：6，他引：7
研究了人工养殖西伯利亚鲟精子的生物学特征及超低温冷冻保存方法。西伯利亚鲟的产精量为113.67±39.86 ml,精子密度为(6.49±3.10)×108/ml,精子活力为(85.4±9.5)%,精子寿命为353±23 s。精子密度与精子快速运动时间、精子寿命之间均存在线性相关,用方程分别表示为:y=1.0384x+1.5089(R2=0.7325);y=2.9069x+74.289(R2=0.6967)。结果表明精子密度可作为一项精子质量评价的标准。通过比较西伯利亚鲟精子在不同稀释液、不同抗冻剂和抗冻剂浓度、降温速率、解冻温度下的保存效果,结果表明:配方2作为稀释液,18%甲醇作为抗冻剂,二步法超低温(-196℃)冷冻保存精子,40℃水浴解冻取得最好的冻后活力,解冻后活力为(51.8±5.8)%。西伯利亚鲟授精的最佳精卵比为106∶1。在此精卵比下用冻精授精分别得到了(72.3±3)%的受精率和(52.9±4.1)%的孵化率,其中受精率与鲜精没有显著性差异,孵化率与鲜精有显著差异(P<0.05)。 相似文献
9.
为研究不同浓度溶解氧(DO)对团头鲂(Megalobrama amblycephala)耐低氧新品系F5代鳃组织形态以及各组织酶活性的影响,本研究将团头鲂在低氧[DO为(1.7±0.2) mg/L]和高氧[DO为(19.3± 0.5) mg/L]条件下分别处理0、4和7 d,恢复常氧[DO为(7.8±0.3) mg/L] 7 d后,通过石蜡切片观察鳃组织的形态,并测定了鳃、肝胰腺、肠道、肌肉中过氧化氢酶(CAT)、琥珀酸脱氢酶(SDH)及乳酸脱氢酶(LDH)活性和丙二醛(MDA)含量。石蜡切片结果显示,随着低氧处理时间的延长,团头鲂新品系鳃丝的层间细胞团体积减小,鳃小片表面积增加,恢复常氧7 d后又有所恢复。高氧条件下,层间细胞团体积增大,鳃小片表面积减小,恢复常氧7 d后也会恢复。酶活性检测结果显示,在低氧和高氧2种条件下,随着处理时间的延长,CAT活性和MDA含量在各组织中的变化无明显规律,但均有显著差异(P<0.05)。低氧条件下,LDH活性显著增高,SDH活性显著降低(P<0.05)。高氧条件下,LDH活性显著降低,SDH活性显著增高(P<0.05)。本研究可为溶解氧对团头鲂的鳃组织以及各组织酶活性的影响提供基础资料,并为团头鲂新品系的养殖与选育奠定基础。 相似文献
10.
日本黄姑鱼精子生理特性及超低温冷冻保存研究 总被引：5，自引：1，他引：4
对日本黄姑鱼精子部分生理特性及其超低温冷冻保存技术进行了研究。结果表明,日本黄姑鱼精液pH值为7.0~7.5,精子密度为(11.23±2.78)×109/m l,精子寿命(229.33±17.16)s。在盐度为30~35,pH为7.5~8.5时,精子的活力最高,分别达到(88.33±2.89)%和(88±4.33)%。精子在室温(25℃)条件下,可存活24 h;在低温(4℃)条件下可以存活36 h。以D液作为稀释液,20%的乙二醇作为抗冻剂,利用快速降温法对精子进行超低温冷冻保存,38℃水浴快速解冻,解冻后的精子获得最高的活力为(47±5.78)%。 相似文献
11.
Abstract Four parameters were examined in order to define sperm quality in turbot Scophthalmus maximus L., sperm: (1) sperm motility, measured by direct counts of the number of active spermatozoa, expressed as % of total spermatozoa; (2) retention of motility after activation, measured by direct counts, 0–60min after activation, expressed as a % of the initial level of activity; (3) resistance to thermal stress, measured as change in retention of motility, and (4) adenosine phosphate (ATP) concentration, determined for samples of non-activated sperm. The proportion of motile spermatozoa at activation ranged from 34·8% to 97·6% (mean 76·3%) for the individual males tested. Turbot sperm retained on average 52% (range 27–90%) of its initial activity one hour after activation. Sperm samples which were stressed by cooling to –27°C retained only 8·6% (range 0–25%) of initial activity, compared to control samples which retained 49% (range 38–63%) of initial activity. The retention of motility after activation was not significantly related to the initial motility or the levels of ATP. Concentrations of ATP in turbot sperm (mean 0·46mg ATP/106 spermatozoa, equivalent to 9·2nmol ATP/108 spermatozoa) were comparable to those measured in mammals. 相似文献
12.
Padmanav Routray Surjya Narayan Dash Chidananda Dash Priyabrat Swain Sampad Kumar Sarkar Niranjan Sarangi 《Aquaculture Research》2008,39(15):1597-1605
The effects of extender composition, cryoprotectant concentration and freezing and thawing on the fertilization efficiency of cryopreserved spermatozoa of Puntius gonionotus were evaluated. Computer‐aided motility analysis of semen was conducted to check the suitability of spermatozoa for cryopreservation after mixing with different extenders and cryoprotective agents (CPAs). Extender‐4 with an osmolality 260 mOsmol kg−1and pH 7.6 was used for the cryopreservation study. Among the CPAs, dimethyl sulphoxide (DMSO) was least toxic and more than 60% fertilization was achieved when used at 1.4 M at 0 °C for 10 and 30 min, whereas the toxicity of all CPAs to spermatozoa was evident when tested at 30 °C. Semen frozen at −16 °C min−1 with 1.4 M DMSO showed 70% fertilization, which was significantly higher (P<0.05) than other freezing rates. Samples thawed at 35 °C water showed a fertilization rate comparable with that of fresh semen. Computer‐assisted semen analysis of fresh and frozen semen after thawing showed variations in different types of motility in spermatozoa and in their class. There was no significant difference in motility before or after cryopreservation; however, significant differences could be observed in the average path velocity (VAP), straight line velocity (VSL) and curve linear velocity (VCL). Semen of silver barb could be cryopreserved with extender‐4 by addition of 1.4 M DMSO to a final cryopreservation medium (MED 2) cooled at a rate of −16 °C min−1, stored in liquid nitrogen (−196 °C) and utilized after thawing at 35±2 °C. 相似文献
13.
《水生生物资源》1998,11(6):387-394
A sperm cryopreservation protocol adapted from turbot, was tested on sea bass using either 250-μL straws or 1.5-mL cryovials. A dilution to 1/3 in Mounib s extender and a cooling rate of −65 °C·min−1 allowed frozen sperm to recover an initial motility similar to that of fresh sperm at thawing; however, significant differences in motility (P < 0.001, n = 10 fish semen) were observed at further post-activation times, the motility decrease being faster in thawed sperm. At the experimental scale, triplicate inseminations of 2-mL aliquots (approximately 2 000 eggs) showed a significant fertility decay of thawed sperm compared to that of fresh sperm (P < 0.01, n = 12 fish semen) when a discriminating 35·103 spermatozoa to egg ratio was applied. When 70·103 and 200·103 spermatozoa per egg were provided in the same experimental conditions, no significant difference appeared between the fertilisation rates of fresh and thawed sperm. In order to validate the procedure for production or cryobank purpose, a scaled-up protocol was established. Two and 50 mL batches of eggs (approximately 2·103 and 50·103 eggs, respectively) were inseminated in triplicate using either fresh or thawed individual sperms of 5 males with 200·103 spermatozoa per egg. The mean fertility decreased by 23.5 % due to cryopreservation. This decline was explained by the loss of fertility of only one sperm, and only in large-volume conditions, probably due to the delay of use after thawing. 相似文献
14.
A. Shaliutina-Kolešová I. Gazo J. Cosson O. Linhart 《Fish physiology and biochemistry》2014,40(6):1771-1781
The protective influence of seminal plasma and the antioxidants catalase (CAT), superoxide dismutase (SOD), and glutathione (GTH) on quality parameters, oxidative stress indices, and antioxidant activity was studied in common carp (Cyprinus carpio) spermatozoa exposed to the xanthine–xanthine oxidase (X–XO) system. Fish spermatozoa were incubated for 5 and 20 min at 4 °C with X–XO concentrations of 1 mM X–0.1 U/mL, 0.6 mM X–0.05 U/mL, 0.3 mM X–0.025 U/mL, and 0.1 mM X–0.0125 U/mL. A dose-dependent reduction in spermatozoa motility and velocity was observed at concentrations of 0.1 mM X–0.0125 U/mL to 1 mM X–0.1 U/mL XO. Increase in spermatozoa motility parameters was recorded following treatment with antioxidants and seminal plasma. The level of the oxidative stress indices lipid peroxidation (LPO) and carbonyl derivatives of proteins (CP) was significantly reduced after addition of CAT, SOD, or GTH along with seminal plasma. Significant differences in SOD, glutathione reductase, and glutathione peroxidase activity were seen in spermatozoa incubated with, compared to that without, seminal plasma at all studied X–XO concentrations. The data demonstrated that CAT, SOD, or GTH in combination with SP can reduce reactive oxygen species stress in fish spermatozoa and improve spermatozoa quality. 相似文献
15.
Teruyoshi Narita Takayuki Kawamoto Kiyoshi Isowa Hideo Aoki Masahiro Hayashi Hiromi Ohta Akira Komaru 《Fisheries Science》2008,74(5):1069-1074
To clarify factors reducing the motility and fertility of cryopreserved spermatozoa of the Japanese pearl oyster Pinctada fucata martensii, the structure of spermatozoa before and after cryopreservation was observed by scanning electron microscopy. Testicular
spermatozoa were diluted with cryopreservation diluent (10% methanol+18% fetal bovine serum+72% sea water), and dispensed
into 0.25-mL straws. The straws were cooled at a rate of approximately −20 °C/min to −50°C, and subsequently immersed in liquid
nitrogen. Percentage motility of spermatozoa before cryopreservation was 69.9±4.2%, and that of cryopreserved spermatozoa
was 24.0±1.8%, respectively. In cryopreserved spermatozoa, the percentage that lacked or had a deformed flagellum was 56.6±3.9%,
while in fresh spermatozoa this was 8.7±2.0%. In cryopreserved spermatozoa, the percentage of deformed acrosomes was 76.6±5.2%,
while in fresh spermatozoa this was only 0.9±0.3%. Cryopreserved spermatozoa with a normal acrosome and flagellum were only
15.4±3.5% of those in fresh spermatozoa. These results indicate that lesion of the flagellum and deformation of the acrosome
occurred through the cryopreservation procedure, and both types of damage lead to loss of the motility and fertility in thawed
spermatozoa. 相似文献