微囊藻毒素对太湖翘嘴红鲌早期发育阶段的毒性研究

(24±1) ℃时采用半静止式生物毒性试验法分析了5个不同质量浓度(500、250、125、62.5、31.25 μg/L)微囊藻毒素(MC-LR)对太湖翘嘴红鲌早期发育阶段的影响.试验结果表明,微囊藻毒素对翘嘴红鲌受精过程影响较小,对胚胎发育的影响较大,胚胎发育后期对微囊藻毒素的敏感性大于胚胎发育前期,对初孵仔鱼的毒性小于对胚胎的毒性;但对早期发育的各个阶段均有一定的致死和致畸毒性作用,主要表现为胚胎发育延缓、引起各种畸形、胚胎死亡和仔鱼运动性差、死亡率增高等.同时计算了MC-LR对翘嘴红鲌的受精过程的EC50分别为998.66、1519.26 μg/L;对卵裂期胚胎、囊胚期胚胎、原肠期胚胎和仔鱼的LC50分别为243.65、181.10、215.99、367.15 μg/L;对卵裂期胚胎、囊胚期胚胎、原肠期胚胎和仔鱼致畸作用的EC50分别为201.07、180.03、176.38、287.98 μg/L.     

紫外线照射对金鱼孵化率的影响

以黑龙睛金鱼（Carassius auratus）受精卵和处于4～8细胞期、多细胞期、原肠期、心脏搏动期的胚胎为对象，研究不同剂量紫外线（UV）照射对金鱼胚胎发育的影响。发现用低剂量的UV照射可以提高金鱼受精卵或发育早期胚胎的孵化率。对发育到原肠胚以后的胚胎UV照射无上述刺激作用，而是随着照射剂量的升高，孵化率急剧下降。另外，实验结果显示，UV并未加速胚胎发育的进程。当UV照射时间小于180s时，受精卵或发育早期胚胎的畸形率没有明显增加。对出现上述结果的可能机理做了探讨并对UV用于育苗孵化和育种两方面的应用提出了建议。     

Zn2+和Cd2+对斑马鱼早期胚胎发育阶段的单一与联合毒性

研究了重金属Zn2+和Cd2+对斑马鱼(Danio rerio)早期胚胎发育阶段的单一与联合毒性效应。分别用不同质量浓度的Zn2+和Cd2+对受精后24 h(24 hpf)的受精卵进行染毒实验,统计了斑马鱼胚胎48 hpf的卵凝结和心律,72hpf的卵凝结、孵化率和畸形率等指标。结果显示,Zn2+和Cd2+对斑马鱼胚胎各不同发育阶段的影响不同,镉的毒性比锌大。斑马鱼胚胎对Zn2+和Cd2+最敏感的毒理学终点均为72 hpf。Zn2+和Cd2+对斑马鱼胚胎72 hpf的发育毒性比48 hpf的发育毒性大。Zn2+和Cd2+在毒性单位配比为1︰1时,对斑马鱼胚胎的联合毒性趋向于毒性剧增的协同作用。Zn2+和Cd2+在毒性单位配比为1:2时,对斑马鱼的联合毒性由48 hpf的协同作用转为72 hpf的拮抗作用。Zn2+和Cd2+在毒性单位配比为2︰1时,对斑马鱼的联合毒性在凝结时为协同作用,在孵化时为拮抗作用。结果表明,Zn2+和Cd2+在不同毒性单位配比下,对斑马鱼胚胎的联合毒性不完全相同。即使在同一毒性单位配比下,Zn2+和Cd2+对斑马鱼胚胎不同毒理学终点的联合毒性也不完全相同。此外还发现,联合作用不仅与化合物的剂量有关,还与染毒时间相关。     

重金属对鱼类早期发育的生态毒理效应

鱼类早期生活阶段对外界环境极为敏感,即使低浓度的污染物暴露也可能对其胚胎发育、仔鱼生长存活造成损害,导致种群数量和质量的降低。研究污染物对鱼类早期生活阶段的毒理效应和机制是揭示污染物对生物种群和水生生态系统潜在风险的有效途径。重金属对鱼类的影响在精、卵发生时即可显现。例如,抑制卵(精)母细胞成熟和精子活力降低,使受精率降低;影响受精卵吸水膨胀过程,改变胚胎发育速率、引起胚胎畸形、影响胚胎心率、改变孵化时间、降低孵化率;影响初孵仔鱼体长、造成仔鱼畸形、影响卵黄囊吸收和胚后仔鱼发育生长、摄食行为等。重金属对鱼类早期发育阶段的毒性效应主要取决于金属种类、暴露浓度和受试鱼种及其发育阶段。此外,外界环境条件等因素也会对重金属的毒性产生显著影响。本研究综合分析国内外关于重金属污染物对鱼类精卵、胚胎、仔鱼等早期发育过程生态毒理效应的研究进展,并结合相关报道阐述了其毒性机制。     

苯硫脲对斑马鱼黑色素生成及早期发育的影响

为探索苯硫脲抑制斑马鱼黑色素生成的最佳浓度及苯硫脲对斑马鱼早期发育的影响,用0.05～2.O mmol/L苯硫脲处理28-体节期斑马鱼胚胎,用1.0～15 mmol/L苯硫脲处理受精后2、10、24 h的斑马鱼胚胎.结果表明,在不同的发育时期,抑制黑色素生成的苯硫脲最佳浓度是不同的;且发育时期越早,斑马鱼对苯硫脲毒性越敏感;苯硫脲低于0.2 mmol/L时对斑马鱼孵化、存活和早期发育无显著影响;0.2 mmol/L苯硫脲导致斑马鱼孵化率降低;0.5～2.0 mmol/L苯硫脲导致斑马鱼的孵化率和存活率均降低,并引起斑马鱼畸形.     

全氟辛烷磺酸对斑马鱼胚胎的急性毒性与致畸效应

为验证全氟辛烷磺酸对鱼类胚胎的毒性作用，采用静态染毒法研究不同质量浓度全氟辛烷磺酸对斑马鱼胚胎的急性毒性和致畸作用。试验结果显示，全氟辛烷磺酸对斑马鱼胚胎24、48、72、96 h的半致死质量浓度分别为7.81、6.58、3.44、2.67μg/mL,安全质量浓度为0.27μg/mL,对斑马鱼胚胎的毒性为高毒。全氟辛烷磺酸质量浓度越高，胚胎孵化率越低、死亡率越高，并伴随明显的心率增加、行动迟缓等现象。在全氟辛烷磺酸胁迫下，斑马鱼胚胎发育不同时期会出现不同的畸形现象：囊胚期(2.7～6.0 h)出现卵凝结，卵黄细胞隆起不明显；原肠胚期(6.6～10.6 h)出现卵黄外突、原肠外突和卵凝结；分裂期(12.1～23.8 h)胚胎尾部畸变和卵黄囊异常；成形期(23.8～52.1 h)胚胎出现心肌肿大、心率不稳、后静脉血流减慢或停止、脊椎和尾部弯曲、色素沉积减少等现象；孵化期(54～80 h)幼鱼出现明显的尾部萎缩和曲折、色素沉积减少、脊椎弯曲、心包腔肿大等现象。当全氟辛烷磺酸质量浓度为1.00～1.58μg/mL时，其致畸作用相对较弱；质量浓度在2.51～10.00μg/mL时，会诱导胚胎出现...     

紫外线照射对中华锯齿米虾胚胎发育的影响

自抱卵的中华锯齿米虾雌虾腹部剥离取卵裂期、囊胚期、原肠胚期和无节幼体Ⅰ期的胚胎置于培养皿中,暴露在紫外线下,照射剂量分别为0、1、2、4、8、16 W·s/cm^2,人工振荡培养箱中以速度55 r/min、温度25℃、湿度60%培养。每隔24 h在解剖镜下观察胚胎发育至48 h后进行数据分析,研究紫外线照射剂量对中华锯齿米虾胚胎发育的影响。结果表明,胚胎的存活率对紫外线具有明显的剂量依赖关系。随着紫外线照射剂量的增加,胚胎发育延缓。紫外线暴露后48 h的半数致死量分别为:卵裂期11.5 W·s/cm^2、囊胚期8.6 W·s/cm^2、原肠胚期12.0 W·s/cm^2、无节幼体Ⅰ期14.2 W·s/cm^2。相同的暴露剂量下,暴露时间对胚胎发育也有明显影响。     

肌间刺缺失突变对斑马鱼胚胎发育过程中肌肉发育的影响

为阐明肌间刺缺失对斑马鱼(Danio rerio)胚胎发育时期肌肉发育的影响,采用qRT-PCR的方法,检测分析了斑马鱼不同肌间刺表型间5个肌肉特异性基因(mef2、myf5、myod、myog和sox6)在囊胚期(3hpf,hourspost fertilization)、原肠胚期(6 hpf)、体节期(12 hpf)、咽囊期(24 hpf)和孵化期(72 hpf)5个发育时期的表达情况,对比分析了肌间刺缺失对斑马鱼胚胎发育过程中肌肉发育特异性基因的时序表达差异和肌肉发育的影响,以及对受精率、孵化率和畸形率的影响。结果显示, 5个特异性基因在肌间刺缺失突变型与野生型斑马鱼各发育时期的表达量无显著性差异(P0.05)。其中,myf5、myod基因在原肠胚期表达量升高,在体节期和咽囊期开始逐步下降;mef2、myog和sox6基因在囊胚期和原肠胚期表达量很低,在体节期和咽囊期表达量迅速升高,并在孵化期开始下降。此外,受精率、孵化率和畸形率的统计结果显示,肌间刺突变型与野生型间亦无显著性差异(P0.05)。综上,推测肌间刺的缺失对斑马鱼胚胎发育及胚胎发育过程中肌肉的发育没有影响。     

不同亚硝酸盐对斑马鱼胚胎发育的急性毒性效应

用质量浓度为0.4 mg/mL、0.8 mg/mL、1.2 mg/mL、1.6 mg/mL和2.0 mg/mL的NaNO_2以及质量浓度为0.5mg/mL、1.0 mg/mL、1.5 mg/mL、2.0 mg/mL和3.0 mg/mL的KNO_2溶液分别处理受精后6 hpf (hours post fertilization)的斑马鱼Danio rerio胚胎,培养至96 hpf,每隔24 h在体视显微镜下观察记录胚胎死亡、孵化和发育异常状况,并测定胚胎自主活动次数、心率和体长,研究不同亚硝酸盐对斑马鱼胚胎发育的影响。结果表明:当亚硝酸盐质量浓度为0～0.5 mg/mL时,斑马鱼胚胎生长发育无明显异常;当亚硝酸盐质量浓度达到或超过1.0mg/mL时,开始出现毒害效应:胚胎自主活动、心跳、孵化率和体长下降,未孵出的胚胎发育异常,心包水肿、脊柱弯曲、头部发黑和死亡。NaNO_2和KNO_2的96 hpf-LC_(50)分别为1.30 mg/mL和1.44 mg/mL,96 hpf-EC_(50)分别为1.12mg/mL和1.50 mg/mL,96 hpf-IC_(50)分别为0.84 mg/mL和1.04 mg/mL。亚硝酸盐对斑马鱼胚胎发育有一定的抑制作用和致畸效应,且NaNO_2的毒害作用大于KNO_2。     

人工诱导栉江珧雌核发育的初步研究

利用紫外线诱导进行栉江珧(Atrina pectinata)雌核发育单倍体的研究,用强度为2 561μW/(cm2.s)的紫外线(254nm)照射不同时间的精子与正常卵子受精展开实验。结果发现随照射时间的增加,卵裂率、早期胚胎存活率和D型幼虫发生率总体呈下降趋势,遗传失活的精子与正常卵子受精后其胚胎发育至D型幼虫前期停止。经紫外线照射的精子受精后所产生的单倍体胚胎发育速度慢于正常受精所产生的二倍体胚胎,各照射组均出现非整倍体。40 s照射组中卵裂率达到51.2%,D型幼虫发生率为0,单倍体率最高,达到36.8%,出现“Hertwig效应”。由于雌核发育单倍体的有效诱导既需要精子遗传物质失活、又需要保证较高的卵裂率,因而在强度2 561μW/(cm2.s)的紫外线下照射40 s是获得栉江珧雌核发育单倍体的最佳条件。     

HSP90抑制剂根赤壳菌素对斑马鱼胚胎发育的影响

为研究热休克蛋白90(heat shock protein 90,HSP90)在斑马鱼(Danio rerio)胚胎发育中的作用,本实验采用2μmol/L、5μmol/L、10μmol/L的HSP90抑制剂根赤壳菌素(radicicol)对斑马鱼发育期胚胎进行处理,监测斑马鱼胚胎不同发育时期两个HSP90功能抑制标志基因BAG3(BCL2-associated athanogene3)和HSPB1(heat shock protein beta-1)的mRNA的表达水平,并观察不同发育时期的胚胎发育状况。结果如下:(1)实时荧光定量PCR结果显示,BAG3和HSPB1 mRNA水平在根赤壳菌素处理胚胎12 hpf(hours post-fertilization,hpf)或24 hpf后显著增高,Western blot检测到5μmol/L根赤壳菌素处理胚胎24 hpf后HSP70表达上调,证明该实验条件下HSP90功能受到抑制;(2)根赤壳菌素处理后斑马鱼胚胎发育变缓,胚胎成活率统计显示:2μmol/L、5μmol/L、10μmol/L根赤壳菌素处理24 hpf胚胎成活率分别是95%、77%、35%,成活率随根赤壳菌素浓度的增高而降低;(3)5μmol/L根赤壳菌素处理72 hpf可见部分个体色素沉积减少、心包膜增大、肌肉萎缩等发育畸形。研究结果证明HSP90在斑马鱼胚胎发育过程中发挥重要的作用,根赤壳菌素在5μmol/L时已达到较佳抑制效果且成活率较高,而且胚胎发育出现了各种形态学变化,为后期研究HSP90调节斑马鱼胚胎发育奠定了基础。     

Effects of embryonic exposure to α‐lipoic acid or ascorbic acid on hatching rate and development of zebrafish (Danio rerio)

This study investigates the effects of embryonic exposure to two different antioxidants on growth and development in fish. Zebrafish (Danio rerio) embryos (100 per group) were exposed to lipoic acid (LA, 6–12 μM) or ascorbic acid (AA, 100–200 μM) and the hatching rate, standard lengths (SL) at hatching, development and growth post‐hatching monitored. The SLs at hatching were increased (P<0.05) in both antioxidant‐exposed groups relative to the controls, with no effect on yolk reserves. This enhanced development persisted up to 15 days post hatching. At hatching, cell proliferation rates (P<0.0005) and basic fibroblast growth factor (P<0.001), were greater in the antioxidant‐exposed fish than in the controls (0 μM antioxidant); no oxidative DNA damage was detected (P>0.05). Activity of the endogenous antioxidant enzyme superoxide dismutase was greater (P<0.001) in LA‐treated fish than in the controls. The results suggest that embryonic treatment of zebrafish with LA or AA during embryogenesis enhanced cell proliferation, leading to increased somatic growth in the larval stages, persisting into the juvenile stage. The findings support the treatment of embryonic fish with antioxidants for enhanced results in aquaculture.     

Induction of centrum defects in amberjack Seriola dumerili by exposure of embryos to hypoxia

ABSTRACT:   Artificially hatched Seriola species have the problem of malformation, mainly in their vertebrae, head, and mouth parts. To clarify the cause of vertebral malformation, the effects of hypoxia during embryogenesis on the induction of centrum defects was investigated in artificially hatched amberjack Seriola dumerili . Firstly, 7-somite stage embryos were exposed to waters of 0, 12.5, 25, 50, and 100% dissolved oxygen (DO) for 0.5, 1, 2, 3, 4, and 5 h to confirm the effective dose (DO concentration and duration of exposure) of hypoxia that induces somitic disturbances in newly hatched larvae. Exposure of embryos to 12.5% DO concentration for longer than 0.5 h induced somitic disturbances. Following this result, centrum defects in juveniles were investigated by an induction experiment with embryos exposed to 12.5% DO for 2 h at the gastrula, 1- or 2-somite, 10-somite, 15-somite, or heart beating stage. This experiment revealed that centrum defects were induced only during somitogenesis, and somitic disturbances were the premonitory symptom of centrum defects. These results indicate hypoxia during somitogenesis as a possible cause of centrum defects in amberjack.     

遮阴和紫外辐射对凤眼莲株型与光合作用的影响

采用室外培养,单因子设计,设置紫外辐射（UV-B 100 μW/cm2）、遮阴（光强为自然光的70%）和全自然光3个处理,测定凤眼莲（Eichhornia crassipes）30d处理前后生长势和光合参数的变化。结果表明：（1）凤眼莲是对光因子高度敏感的植物,不同光环境下株型不同;与全自然光处理材料相比,遮阴后增加了...     

UV-B紫外辐射对条斑紫菜生理生化的影响

条斑紫菜(Pyropia yezoensis)作为一种生长在潮间带的经济海藻，在适应干露失水时的高紫外辐射上有其独特的特性。本研究以无UV-B辐射为对照组，0.5 W/m2(低)和2 W/m2(高)UV-B辐射为实验组，检测条斑紫菜对UV-B紫外辐射响应的叶绿素荧光光合参数、多光谱荧光参数以及酪氨酸酶活性的变化。结果显示，受到UV-B辐射的条斑紫菜的叶绿素荧光光合参数(QY_max、NPQ_Lss、qP_Lss和Rfd_Lss)随辐照时间呈不同程度的下降趋势，并且高辐射强度下的各光合参数值低于低辐射强度，表明条斑紫菜光合作用受到辐射的负面影响；极早期胁迫参数F440/F690和F440/F740的差异表明，2 W/m2 UV-B辐射组条斑紫菜比0.5 W/m2辐照组更早(1 h)进入胁迫状态，且UV-B辐射下条斑紫菜酚醛类物质减少；与叶绿素浓度呈负相关的参数F690/F740变化表明，0.5 W/m2辐照组叶绿素浓度高于对照组，而在辐照3 h内低于2 W/m2辐照组，2 W/m2辐照组叶绿素浓度则先上升，并在辐照4 h下降，至6 h呈显著降低，说明藻体可耐受较低UV-B辐射，而高UV-B长时间辐照对藻体造成明显的伤害；对酪氨酸酶的活性检测结果表明，受到UV-B辐照的条斑紫菜酪氨酸酶活性显著高于对照组，且高辐照组的酶活性高于低辐照组的酶活性。     

Stress-induced apoptosis by heat shock, UV and γ-ray irradiation in zebrafish embryos detected by increased caspase activity and whole-mount TUNEL staining

SUMMARY: Environmental stress-induced apoptosis in zebrafish Danio rerio embryos was characterized by assaying caspase-3-like activity and whole-mount terminal deoxynucleotidyl nick-end labeling (TUNEL) staining. Severe stress conditions, such as heat shock at 39°C for 1 h, ultraviolet light at 10–100 mJ/cm² and γ-ray irradiation at 5–20 Gy induced extensive apoptosis in embryos. Apoptotic cells were observed after the bud and 1-somite stages in normal embryos by TUNEL staining, and after stress treatment many TUNEL-positive cells were found in the enveloping and deep cell layers and the larval fin. The caspase-3-like activity increased severalfold during stress-induced apoptosis in a dose-dependent manner. These findings indicate that apoptotic pathways, mediated by caspase-3-like activity, play a major role in zebrafish embryogenesis under stress conditions.     

不同光强处理对长茎葡萄蕨藻叶绿素荧光特性的影响

通过调制叶绿素荧光仪(MINI-PAM),对长茎葡萄蕨藻(Caulerpa lentillifera)在10μmol/(m~2·s)、120μmol/(m~2·s)、360μmol/(m~2·s)光强下的叶绿素荧光特性进行了研究。结果表明:随着光照强度从10μmol/(m~2·s)上升到360μmol/(m~2·s),长茎葡萄蕨藻最大光量子产量(F_v/F_m)、实际光量子产量、相对电子传递速率(rETR)、快速光曲线初始斜率均呈下降趋势;用120μmol/(m~2·s)和360μmol/(m~2·s)光强处理的样品光化学猝灭(qP)较高,而非光化学猝灭(NPQ)较低;不同光强处理4 h和8 h时,与10μmol/(m~2·s)处理组相比,120μmol/(m~2·s)处理组最大相对电子传递速率(rETR_(max))、半饱和光强(I_k)均显著上升,360μmol/(m~2·s)处理组I_k显著上升,rETR_(max)明显降低;在360μmol/(m~2·s)光强下处理12 h与处理8 h相比,样品rETR_(max)、I_k均显著降低。上述结果说明长茎葡萄蕨藻偏向适应10μmol/(m~2·s)的低光条件,在120μmol/(m~2·s)和360μmol/(m~2·s)的高光条件下能通过不同的方式提高对光强的耐受能力,但是在360μmol/(m~2·s)光强下处理12 h会引起光氧化对光合结构造成破坏。     

Lipid-based transfection as a method for gene delivery in zebrafish (Danio rerio) embryos

A major challenge to the widespread production of transgenic, knockout and knockdown zebrafish has been the absence of a simple and effective procedure for introducing macromolecules into the fertilized egg. None of the existing techniques for gene transfer in fish embryos has proven to be a major advance over cytoplasm microinjection, which is a technically demanding and time‐consuming procedure. This report addresses this need, considering that the development of protocols for lipid‐based transfection with fish embryos would considerably simplify gene transfer in this complex biological model. In this study, lipid‐based transfection with two different reporter vectors was carried out in zebrafish embryos at different developmental stages. The parameters tested included different plasmid/transfection reagent ratios as well as the influence of an added transfection enhancer reagent. When embryos were transfected in the blastula stage with a pEGFP‐N1 vector, more than 35% successfully incorporated the plasmid and expressed the fluorescent protein 24 h after transfection. The transfection enhancer did not show any significant effect in our experiments. This work presents an approach to implement this technique as a faster, cheaper and more practical alternative than microinjection.