达氏鲟精子的主要生物学特性

达氏鲟(Acipenser dabryanus)属淡水定居性鲟鱼类,为我国特有种,主要分布在长江上游干流及金沙江下游。长期人为过度捕捞及其生存环境的持续污染和水利工程的影响,使得达氏鲟自然种群资源遭到严重破坏,其配子质量的下降己经成为限制其规模化人工繁殖成功的关键因素之一,因此为解决达氏鲟规模化人工繁殖过程中存在的关键性技术点,作者从达氏鲟精液基本特征、精浆元素组成以及不同水体和Na+、K+对达氏鲟精子活力的影响、精子超微结构方面入手,对达氏鲟精子的生理生态特性进行了研究。结果显示,达氏鲟精子平均密度为1.52×109个/ml;精浆元素以Na+含量最高,其次是K+,之后为Ca2+、Mg2+、Cu2+、Zn2+,其中Na+、K+、Zn2+在达氏鲟精浆中的含量有极显著性差异(P0.01),Ca2+、Cu2+、Mg2+差异不明显;精子在江水中的活力最高;在Na+浓度为20 mmol/L时,精子活力最高,精子快速运动时间(FT)和寿命(LT)分别为(66.7±7.1)s和(177.0±14.9)s;达氏鲟精子对K+浓度变化较为敏感,在K+浓度为0.05 mmol/L时,精子FT和LT最长,分别为(109.0±16.1)s和(189.3±12.4)s,K+浓度超过0.05 mmol/L后精子FT和LT急速下降,当K+浓度达到0.5 mmol/L以上时,精子活力立即受到抑制;达氏鲟精子细胞核长(5.67±0.20)μm,鞭毛长(63.16±2.79)μm,全长为(70.35±2.92)μm。     

短波紫外线对日本三角涡虫的损伤及六种酶活力的影响

用短波紫外线(UV-C)照射处理日本三角涡虫,观察了涡虫的损伤状况,探讨了紫外线辐射对涡虫乳酸脱氢酶(LDH)、Na -K -ATP酶、Ca2 -Mg2 -ATP酶、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)和谷胱甘肽过氧比物酶(GSH-PX)活力的影响。结果表明,短波紫外线辐射对涡虫的损伤明显,照射的时间越长,涡虫在照射后出现死亡解体现象的时间就越短。随着紫外线照射时间的延长,LDH活力呈下降趋势;Na -K -ATP酶活力呈上升趋势而Ca2 -Mg2 -ATP酶活力呈下降趋势;SOD活力有明显的增加;CAT活力有增加趋势;GSH-PX活力波动较大,但总体仍呈增加趋势。不论是从紫外线对日本三角涡虫的损伤情况、还是对其六种酶活力的影响来看,日本三角涡虫是一种对紫外线辐射非常敏感的动物,在检测紫外线辐射及筛选抗紫外线辐射防护剂等方面具有潜在价值和应用前景。     

雷公藤制剂对雄性布氏田鼠的不育作用

采用强制给药和自由取食2种方式,研究了雷公藤制剂对发育期成体雄性布氏田鼠不育作用,给药剂量分别为80 mg/kg、120 mg/kg和160 mg/kg.结果表明,120 mg/kg和160 mg/kg剂量使睾丸脏器系数下降,但剂量组间差异不明显(F(3,21)=2.197, F0.05=3.07,P>0.05).不同剂量能够对附睾中精子数量和活力产生显著影响(F(3,21)=17.305, F0.05=3.07,P<0.05),且随药物剂量的升高精子密度与精子活力均呈下降趋势.精子畸形率亦随药物剂量的增加而增加.80 mg/kg及其更高的剂量使附睾出现萎缩并造成睾丸组织损伤.120 mg/kg剂量可减少布氏田鼠平均胎仔数及降低繁殖率.给药后的繁殖后代未发现畸形幼仔.雷公藤制剂的最佳使用剂量介于120～160 mg/kg.自由取食与连续强制给药两种方式对雄性布氏田鼠的作用差异不显著.     

几种不同浓度的离子及单糖对中华鲟精子活力的影响

本文研究在不同浓度Na 、K 、Ca2 、Mg2 、Cu2 及葡萄糖、果糖溶液中,中华鲟(Acipenser sinensisGray)精子活动情况。结果表明:Na 浓度为2‰时,中华鲟精子寿命时间(Life time,LT)最长,为347s,而Na 浓度为1‰时,中华鲟精子剧烈运动时间(Acute movement time,AT)最长,为98.67s,精子激活率在Na 浓度小于等于2‰时,为100%,随着Na 浓度的增加中华鲟精子激活率明显下降;与Na 溶液不同,在K 浓度为0.005‰时中华鲟精子活力最佳,其AT和LT最长,分别为80s和174s。而精子激活率随K 浓度增加而增强,在浓度为0.005‰时激活率最强为60%,然后快速下降;在Ca2 、Mg2 、Cu2 三种溶液中,随着三种溶液浓度的增加,中华鲟精子剧烈运动时间和寿命逐渐缩短,精子激活率逐步下降;与Ca2 、Mg2 、Cu2 三种溶液完全不同的是,随着葡萄糖和果糖两种单糖浓度的增加,中华鲟精子AT及精子LT逐渐延长。结果说明适量浓度的Na 、K 可以延长中华鲟精子AT及其LT,激活精子活力;而Ca2 、Mg2 、Cu2 三种溶液即使在较低浓度下都表现出对中华鲟精子AT、精子LT及其激活率明显的抑制作用;而葡萄糖和果糖则能有效地延长中华鲟精子AT和LT。     

氧离子辐射对体外人精子自发化学发光、运动性、顶体反应和存活率的影响(英文)

我们研究了不同剂量(0,0.25,0.5,1,2,4,8,16,32,64Gy)3.17MeV/u氧离子对体外人精子自发化学发光(SCL)、运动性,顶体反应(AR)和存活率的影响。结果表明,精子SCL随辐射剂量明显增强,最低有效剂量为0.5Gy;精子运动性在0.5—2Gy,AR在0.5—4Gy辐射组均明显高于其对照组,但均随辐射剂量的进一步增加而明显降低;在0.25—8Gy剂量之间,精子存活率与其对照组相比无明显变化,但当剂量进一步增加至16—64Gy时,存活率急剧下降。这些结果提示,低剂量重离子辐射能对精子运动性及AR等功能产生兴奋效应。而大剂量则明显抑制这些功能,甚至造成精子死亡。其作用机制可能与重离子辐射诱导的自由基反应有关。     

UV-B增加对玉米花粉抗氧化能力及授粉后籽粒发育的影响

在玉米散粉期,收集花粉进行UV-B辐射增强处理后进行人工授粉研究,结果表明,随着UV-B辐射时间的增加,玉米花粉的SOD、POD和CAT活性都呈明显下降的趋势,而MDA含量则呈相应上升的趋势;玉米每穗粒数与花粉抗氧化酶活性呈正相关、与MDA含量呈负相关关系,并随UV-B辐射时间延长呈下降趋势,且辐射超过2h后下降达到显著水平;玉米百粒重随UV-B辐射时间的延长明显下降,但辐射花粉1~2h对其无显著影响;UV-B辐射花粉对籽粒的可溶性糖、淀粉、脂肪及粗蛋白含量等营养成分无显著影响。     

泥鳅和大鳞副泥鳅卵雌核的紫外辐射灭活条件筛选

采用紫外线对泥鳅和大鳞副泥鳅卵雌性原核进行干法灭活以及在合成卵巢液、TC-199溶液、Ringer氏溶液和Holtfreter氏溶液中灭活,结果表明:当辐射剂量为90-180mJ/cm~2时,卵子在四种溶液中的单倍体诱导率均较高。合成卵巢液组的最佳辐射剂量为120-180mJ/cm~2;TC-199和Ringer氏溶液组的最佳辐射剂量为120mJ/cm~2;原液使用Holtfreter氏溶液效果欠佳,干法灭活效果最差。在上述各辐射剂量下,畸形苗经染色体组鉴定92％为雄核发育单倍体。综合评价,灭活效果依次是:人工合成卵巢液>TC-199溶液>Ringer氏溶液>Holtfreter氏溶液>干法。     

不同流速下杂交鲟幼鱼游泳状态与活动代谢研究

为研究水流速度对杂交鲟幼鱼行为和代谢的影响,探讨游泳状态与活动代谢及相关游泳运动参数之间的关系,在26℃水温下,使用特制的鱼类游泳行为和活动代谢同步测定装置,测定了杂交鲟幼鱼在0.1、0.3、0.5 m/s三种流速和静水条件下的游泳状态、趋流率、摆尾频率和耗氧率。结果表明:随着流速的增大,杂交鲟幼鱼逆流前进和逆流静止游泳状态所占时间比例显著减少,而逆流后退所占时间比例显著增加,顺流而下时间比例有所上升。在0.0—0.3 m/s的流速范围内,杂交鲟幼鱼各个时段的平均趋流率、摆尾频率和耗氧率均随着流速的增加而增大,在0.3 m/s流速下分别达到100%﹑(2.53±0.34)Hz和(490.99±164.59)mg O2/(kg.h)。当流速增加至0.5 m/s时,在趋流率仍保持100%的情况下,其耗氧率相比0.3 m/s增加了21.86%,而摆尾频率却减小了6.70%。实验过程杂交鲟幼鱼趋流率与摆尾频率呈显著线性正相关,而摆尾频率与耗氧率在大部分时段却无相关性。随着时间的延长,各流速组杂交鲟幼鱼趋流率、摆尾频率和耗氧率呈现不同的变化趋势,其趋流率均相对稳定;但摆尾频率均随时间延长呈下降趋势,而耗氧率则在实验前9h随时间延长逐渐增加,随后趋于稳定。研究结果提示:杂交鲟幼鱼游泳状态的变化与流速有关,而反映运动强度大小的摆尾频率与活动代谢率的关系受到游泳状态的显著影响,同时也与运动代谢特征的时间变化有关。       

钠、钾、钙和葡萄糖对白斑狗鱼精子活力的影响

观察了白斑狗鱼精子在0～679．6kPa NaCl、KCl、葡萄糖溶液和0～339．8kPa CaCl2溶液中的活动情况。在NaCl、KCl、葡萄糖溶液中,白斑狗鱼精子快速运动时间和寿命的变化规律基本一致,精子活动最适渗透压介于339．8～453．0kPa。K^ 有延长精子寿命的作用。外源性葡萄糖可被精子利用,增强精子活力．延长精子寿命。56．7kPa CaCl2对精子活动有抑制作用,并引起精子聚集,该效应随着Ca^2 浓度升高而增强。     

环境因子及超低温冻存对黄姑鱼精子活力的影响

通过测定精子的激活率、运动时间及寿命研究了环境因子变化对黄姑鱼精子活力的影响及超低温冻存后黄姑鱼精子的活力。结果表明,黄姑鱼精子激活与运动的适宜盐度为25～35、适宜pH为7.5～8.5。在pH 8.0～8.5、盐度25条件下,精子激活率达(85.33±2.52)%,运动时间及寿命分别为(336±14.02)s及(405.33±12.22)s。精子激活与运动的适宜NaCl、KCl、MgCl₂及葡萄糖溶液浓度分别为300～500 mmol·L^-1、600 mmol·L^-1、800～1000 mmol·L^-1及900mmol·L^-1;精子在缺少HCO₃^-的人工海水中未能被激活;精子在无Ca²⁺或无Mg²⁺的人工海水中激活率均大于80%,但运动时间及寿命均有所缩短。以Cortland及HBSS溶液为稀释液、10%EG为抗冻剂冻存黄姑鱼精子,冻精激活率>80%,运动时间均超过200s。     

氧离子辐射对体外人精子自化学发光,运动性,顶体反应和存活率的影响

Effects of 16O+6 ion irradiation with different doses on human sperm spontaneous chemiluminescence (SCL), motility, acrosome reaction (AR) and viability were examined. Spermatozoa were irradiated with 0, 0.25, 0.5, 1, 2, 4, 8, 16, 32, or 64 Gy 16O+6 ion beam at the energy of 3.17 MeV/u. After irradiation, samples were analyzed by SCL measurement at 1, 2 and 3 h of incubation; motility was determined by the transmembrane migration method within 2 h of incubation; the percentage of AR and viability was evaluated by the triple-stain technique at 3.5 h of incubation. The results showed: sperm SCL was significantly increased with irradiation doses and the lowest effective dose was 0.5 Gy; compared with controls, the transmembrane migration ratio of spermatozoa progressively elevated with irradiation doses at 0.5, 1, and 2 Gy; the percentage of sperm AR markedly increased in 0.5-4 Gy irradiation and the optimal dose was 2 Gy, and then significant decreased with further increase of irradiation doses; the viability had no significant change within 0.25-8 Gy, but was progressively decreased at 16, 32 and 64 Gy. These data suggested that heavy ion at low doses increased motility and AR, whereas had deleterious effects at higher doses, which are associated with free radical reactions induced by heavy ion irradiation.     

不同剂量x射线对大鼠精子CRISP2mRNA表达水平的影响

目的：观察不同剂量x射线对大鼠精子CRISP2mRNA表达水平的影响,探讨其在电离辐射所致大鼠精子功能改变中的作用。方法：用吸收剂量为1、2、4、和6Gy的x射线分别照射活体SD大鼠的外生殖系统1…4812、24h后,用PCR技术检测精子CRISP2基因mRNA表达水平;用光学显微镜观察精予活力。以未照射组为对照。结果：4、6GyX射线照射不同时间（1、4、8、12、24h时）后大鼠精子的CRISP2mRNA相对表达量均较对照组显著下降（P．〈0．05）,其中6Gb,照射24小时后相对表达量最低（P〈0．01）,而4Gy照射组与6Gy照射组相比较差异无统计学意义（P〉0．05）;2Gyx射线照射8h后CRISP2mRNA相对表达量下降有统计学意义（P〈0．05）;2GyX射线照射1、4h后及1GyX射线照射不同时间（1、4、8、12、24la）后大鼠精子的CRISP2mRNA相对表达量较对照组下降,但差异无统计学意义（P〉O．05）。1、2GyX射线照射不同时间（1、4、8、12、24小时）及4GyX射线照射（1、4、8h）后,精子活力与正常对照组相比无明显改变（P〉0．05）;4GyX射线照射12、24h后大鼠精子活力显著低于正常对照;6GyX射线照射不同时间（1、4、8、12、24h）后,精子活力明显低于对照组（P〈0．05）。结论：不同剂量X射线照射不同时间可导致SD大鼠精子活力下降,这可能与其下调CRISP2基因的mRNA表达水平有关。     

Improvement of stored turkey semen quality as a result of He-Ne laser irradiation

Two experiments were carried out to evaluate the effects of He-Ne laser irradiation at various energy doses on the quality of stored turkey semen. Four semen pools were used in Experiment 1. Each pool was divided into 10 aliquots, nine of which were irradiated with energy doses ranging from 0.144 to 10.8 J/cm2 while the tenth one was not irradiated (control). Each sample was evaluated for motility immediately after irradiation, 24 and 48 h later. Energy doses ranging from 3.24 to 5.4 J/cm2 had higher (P <0.01) sperm motility index (SMI) value compared to the control and samples irradiated with lower and higher laser doses. The energy dose of 3.96 J/cm2 was selected for Experiment 2 to obtain further insight on its effects on turkey sperm preservation for up to 60 h. Each pool of four semen was divided into two aliquots: one represented the control and the other one was irradiated with He-Ne laser at an energy dose of 3.96 J/cm2. Each sample was evaluated for motility and viability immediately after irradiation and then at 12 h intervals up to 60 h. The cell energy charge was also measured by HPLC. Exposure to 3.96 J/cm2 increased the SMI and viability of turkey semen stored for 60 h compared to the control (P <0.05). The cell energy charge of irradiated samples was 200% higher than in the control. Laser irradiation increased the longevity of stored turkey spermatozoa, and might be a useful technique to enhance semen quality in long-term storage.     

Swimming of Xenopus laevis sperm exhibits multiple gears and its duration is extended by egg jelly constituents

The motility of Xenopus sperm is initiated by the osmotic shock experienced when these cells are ejaculated into low-salinity pond water. Motility is brief and is required for the sperm to penetrate the jelly layers and fertilize the egg. In this study we demonstrate that extracts of egg jelly contain factors that extend the period of sperm motility as well as providing a chemoattractant activity as previously reported. Both activities are partially dependent on extracellular calcium. Time-lapse and video microscopy show that after activation of motility the number of motile sperm decreases rapidly, with a half-time of about 2 min. Addition of 10% v/v egg jelly extract ("egg water") increased the number of motile sperm 2-fold over controls at 20 s and about 4- to 10-fold over controls at 10 min after initiation of motility. Extension of motility lifetime was not mediated by a nonspecific protein or by allurin, the egg-water protein that has chemoattractant activity. The helical path of Xenopus sperm exhibited tight coupling between rotational and forward velocities in egg jelly, but coupling changed rapidly from moment to moment in low-salinity buffer. Our observations suggest that jelly-derived factors regulate both the longevity and directionality of sperm propulsion.     

Automated analysis of rabbit sperm motility and the effect of chemicals on sperm motion parameters.

Appropriate software settings and optimum procedures were determined for the measurement of the motion parameters of rabbit spermatozoa by the CellSoft (Cryo Resources Ltd., Montgomery, NY) computer-assisted digital image analysis system. The system was used to follow motion parameter changes occurring in spermatozoa incubated for 6 hr with or without exposure to chemicals. Mean amplitude of lateral head displacement (AALH) increased over the 6 hr period, while curvilinear velocity (Vc) first increased and then decreased. Values for linearity (Lin), or beat cross frequency (BCF), were unchanged. The majority of spermatozoa progressed linearly, with rapid rotation of the sperm head, but subpopulations of spermatozoa with different swimming patterns appeared after 1-3 hr of incubation. Percentage motile sperm and Vc were most sensitive to the action of the compounds (pyrogallol, hydroquinone, ammonium oxalate, triethyl phosphite, and pinocolyl alcohol), while BCF was least affected. The decline in percentage of motile sperm was dependent on duration of exposure and chemical concentration. Mean Vc of the sperm population decreased rapidly upon chemical exposure and remained at a low value until motility ceased. The initial decrease in Vc was dependent on the concentration of the added compound. Motion-based indices--motility concentration (MCI50), motility time (MTI50), and velocity (VI)--were defined and used as toxicological endpoints. The rank order of these indices, the end point of the neutral red in vitro assay for cytotoxicity, and LD50 values for the five compounds were the same, suggesting that chemical inhibition of sperm motility may be useful as a method for the in vitro assessment of chemical cytotoxicity.     

Labelling of living mammalian spermatozoa with the fluorescent thiol alkylating agent, monobromobimane (MB): immobilization upon exposure to ultraviolet light and analysis of acrosomal status

Living spermatozoa of seven mammalian species were treated with the thiol-alkylating fluorescent labelling compound, monobromobimane (MBBR). MB-labelling alone had no effect on sperm motility, nor on the time course or ability of golden hamster spermatozoa to undergo the acrosome reaction when capacitated in vitro. Exposure of MB-labelled spermatozoa to ultraviolet (UV) light and excitation of the MB fluorochrome resulted in virtually immediate immobilization of the spermatozoa without affecting acrosomal status. UV exposure of unlabelled spermatozoa for up to 30 sec had no effect upon motility. Immobilization of MB-labelled spermatozoa depended on the midpiece being irradiated, as irradiation of the head alone, or of the more distal parts of the principal piece, had little or no effect upon motility. Labelling with MB followed by immobilization of individually selected spermatozoa was most useful for detailing the course and site of occurrence of the acrosome reaction during penetration of the cumulus oophorus by golden hamster spermatozoa in vitro. In these often hyperactivated spermatozoa, precise determination of the acrosomal status could not often otherwise be made due to the difficulty in visualizing the acrosomal region of a vigorously thrashing, hyperactivated spermatozoon. This technique should prove valuable in a variety of studies on sperm motility, capacitation and fertilization, and could also be extended to other cell systems.     

The Effect of Low-Level Laser Irradiation on Sperm Motility,and Integrity of the Plasma Membrane and Acrosome in Cryopreserved Bovine Sperm

Background and Objective

Freezing changes sperm integrity remarkably. Cryopreservation involves cooling, freezing, and thawing and all these contribute to structural damage in sperm, resulting in reduced fertility potential. Low-level laser irradiation (LLLI) could increase energy supply to the cell and cause reactive oxygen species reduction (ROS), contributing to the restoration of oxygen consumption and adenosine triphosphate synthesis (ATP) in the mitochondria. Our goal was to analyze the effects of low-level laser irradiation on sperm motility and integrity of the plasma membrane and acrosome in cryopreserved bovine sperm.

Study Design/Materials and Methods

We analyzed 09 samples of bull semen (Bos taurus indicus), divided into three groups: a control group without laser irradiation, a 4J group subjected to a laser irradiation dose of 4 joules, and a 6J group subjected to dose of 6 joules. Samples were divided for the analysis of cell viability and acrosomal membrane integrity using flow cytometry; another portion was used for motion analysis. Irradiation was performed in petri dishes of 30 mm containing 3 ml of semen by an aluminum gallium indium phosphide laser diode with a wavelength of 660 nm, 30 mW power, and energy of 4 and 6 joules for 80 and 120 seconds respectively. Subsequently, the irradiated and control semen samples were subjected to cryopreservation and analyzed by flow cytometry (7AAD and FITC-PSA) using the ISAS - Integrated Semen Analysis System.

Results

Flow cytometry showed an increase in the percentage of live sperm cells and acrosome integrity in relation to control cells when subjected to irradiation of low-power laser in two different doses of 4 and 6 joules (p < 0.05). In the analysis of straightness, percentage of cell movement, and motility, a dose of 4 joules was more effective (p < 0.05).

Conclusion

We conclude that LLLI may exert beneficial effects in the preservation of live sperm. A dose of 4 joules prior to cryopreservation was more effective than a dose of 6 joules in preserving sperm motility.     

Reproductive potential of gamma-irradiated males of the meadow grasshopper Chorthippus parallelus (Zetterstedt) (Orth., Acrididae)

Males of the meadow grasshopper Chorthippus parallelus were treated with five doses ranging from 0 (control) to 140 Gy of gamma irradiation. There was a negative correlation between irradiation dose and post-treatment longevity. Individuals irradiated with the lowest dose lived, on average, longer than the controls. This was presumably not attributable to an energy allocation from tissue since at the time of death there were no dry weight differences. There was a negative correlation between irradiation dose and number of faecal pellets excreted in a period 6–9 days after treatment but not before. There were no negative correlations between irradiation dose and the time required to copulate, copulation duration and ejaculate size. The proportion of tail-less sperm did not differ between the treatments.     

Bower‐building behaviour is associated with increased sperm longevity in Tanganyikan cichlids

We investigated the evolutionary relationship between spawning behaviour and sperm motility traits among Tanganyikan mouth‐brooding cichlid species that have developed diverse mating behaviours and male sexual traits. Mouth‐brooding behaviour is common among these fish, but different species demonstrate a range of spawning behaviours, bower construction, male sexual traits and timing of gamete release. We observed spawning behaviours and compared sperm motility traits of 28 Tanganyikan mouth‐brooding cichlids to elucidate the evolutionary correlations between these traits. Sperm longevity was considerably longer in bower‐building species that construct crater‐shaped spawning sites compared with species that do not build bowers. Male bower builders released sperm in the pit of the bower prior to spawning, and the time from ejaculation to fertilization was longer. Conversely, most mouth‐brooding cichlids deposited semen directly into the female buccal cavity, and spawned eggs were immediately picked up to be placed inside the cavity; thus, the time from ejaculation to fertilization was short. These observations suggest that increased sperm longevity is favoured in bower builders. Comparative phylogenetic analyses suggested that bower‐building behaviour and greater time from ejaculation to fertilization are associated with the extension of sperm longevity, whereas sperm competition rank does not play a major role. In addition, bower‐building behaviour preceded the emergence of increased sperm longevity. These results indicate that the extension of sperm longevity as a result of the emergence of bower builders may have acted as an evolutionary attractor for sperm longevity.