可口革囊星虫的精子发生及精子结构

为了探究可口革囊星虫(Phascolosoma esculenta)精子发生过程及结构上的特殊性,用显微及亚显微技术研究了可口革囊星虫的精子发生和精子结构。可口革囊星虫的精巢位于收吻肌基部,为一曲折的带状组织。成熟精巢内可观察到精原细胞、精母细胞以及精细胞等各阶段的生精细胞。在精子形成早期,很多精细胞脱离精巢,以精细胞团的形式掉落到体腔中。精细胞团内的精细胞同步发育为精子后,脱离精子团进入肾管。成熟精子由头部和尾部组成。头部由钟形顶体与鼓形细胞核构成。顶体后段下包于精核的前端。顶体分内、中、外三层,外层有横隔;顶体下腔内有颗粒状物质不均匀分布,中央有一束丝状纤维组成的顶体棒。核物质电子密度高,核内含空泡。无核前窝,具浅的核后窝。尾部分中段和末段,中段由6个(偶见5个或7个)线粒体围绕近、远端中心粒构成;末段细长鞭状,由轴丝及包绕轴丝的质膜组成,轴丝为典型的"9 2"结构。分析认为:可口革囊星虫精子发生过程以及超微结构上存在特殊的结构与机制:①精细胞团保证了精子形成的同步性;②顶体后段下包于精核的前端使精子头部小而灵巧,利于快速运动;③顶体的横隔使精子顶体的牢固性增强,确保受精时顶体反应的正常进行;④中段较多的线粒体使精子具有更强的环境适应性,有利于有效的受精。     

可口革囊星虫不同组织同工酶的比较

采用聚丙烯酰胺凝胶电泳(PAGE)技术,研究了可口革囊星虫5种组织(收吻肌、吻、肠、肾管及体壁)10种同工酶(ADH、GCDH、SDH、EST、LDH、ME、POD、SOD、MDH、CAT)的表达。结果表明:除CAT外,其余9种同工酶均得到了较为清晰的酶谱,除SOD和MDH在5种组织中表型相同外,其它7种同工酶的表达均具有明显的组织特异性。SDH-2、EST-1、LDH-1和POD-1为收吻肌所特有;ADH-3、ADH-4、GCDH-2、GCDH-4、GCDH-5、EST-10和LDH-6均为吻所特有;EST-4为肾管所特有。在肠中,ADH和GCDH均没有检测到活性。初步探讨了5种组织中各种同工酶的分布与活性及其与各组织生理功能的关系。     

可口革囊星虫肾管肌组织的结构特征

采用显微及亚显微技术观察了可1：7革囊星虫肾管肌组织的结构特征。肾管肌组织位于柱状上皮层下,由纵肌及环肌组成。肌细胞（肌纤维）呈长梭形,核位于细胞边缘并明显突向细胞外基质中,核周围有较多线粒体及少量内质网。肌纤维表面有许多囊状或指状突起的肌质囊,内含肌浆、光面内质网、线粒体及糖原颗粒。肌质囊之间的肌膜内面具膜相关电子致密斑。肌纤维内含粗、细两种肌丝,细肌丝围绕在粗肌丝周围,在肌丝之间分布有糖原颗粒、线粒体及胞质致密体。线粒体及糖原为肌纤维的代谢提供能量,肌组织的收缩对促进肾管的过滤排泄及繁殖时配子进入肾管可能起重要作用。     

可口革囊星虫消化道的形态及组织学结构

对可口革囊星虫(Phascolosoma esculenta)消化道各部分结构进行了形态学和组织学观察,表明可口革囊星虫消化道由翻吻、咽、食道、肠、直肠和肛门组成。翻吻收缩性很强;肠道极长,沿纺锤肌螺旋缠绕成肠索。消化道管壁由内向外分为粘膜层、粘膜下层、肌层和外膜。粘膜上皮主要由柱状细胞组成。除肠外,消化道上皮细胞均有发达的纤毛,肠上皮细胞主要具发达的微绒毛;粘膜下层及肌层的发达程度因消化道的部位不同而异。     

可口革囊星虫消化道的形态结构特征

采用解剖及组织学显微技术观察研究了可口革囊星虫消化道的形态及组织学结构特征。消化道始于口,经咽、食道、游离肠、螺旋肠(上行肠及下行肠)、直肠,止于肛门,各段从内向外均由粘膜层、粘膜下层、肌层及外膜组成。粘膜层表面具微绒毛及纤毛,微绒毛增加了消化道的表面积,纤毛的摆动有利于消化道内食物或消化后残渣的运送;螺旋肠及直肠的粘膜层中均有杯状细胞,在下行肠前段与中段及上行肠前段与直肠中尤其多,杯状细胞能够分泌粘液,保护粘膜上皮,润滑食物及残渣。粘膜下层为结缔组织,肌层为环肌,外膜为浆膜。肠沿纺锤肌盘绕的特点延长了食物的滞留、消化和吸收时间,弥补了缺乏消化腺的不足。肠各段除直肠肌层较厚外,其余各段肌层薄或不明显。     

可口革囊星虫体腔中卵细胞大小组成的周年变化

在浙江温岭沿海逐月采样,显微观察了可口革囊星虫体腔中卵细胞大小组成的周年变化。3—5月,卵径30~90μm的卵细胞占大部分。6月,卵径70~130μm的卵细胞占多数。7月,大部分卵细胞卵径达100μm以上,其中130~150μm的卵细胞约占1/3。8月,卵细胞卵径均在110μm以上,其中130~150μm的卵细胞约占2/3。9月,仍多数卵细胞卵径在110μm以上,130~150μm的卵细胞约占1/3。10月,以110~140μm的卵细胞占多,40μm以下的卵细胞较少,大卵细胞开始退化。11月至翌年2月,随着大型卵细胞的逐渐退化,小型及中型卵细胞逐渐增多,2月仍可见少量未退化的大型卵细胞。分析认为,可口革囊星虫在浙江温岭的繁殖期主要在7—9月。     

可口革囊星虫体内纤溶酶的初步研究

目的从可口革囊星虫中寻找到纤溶酶,并对其进行初步研究。方法采用匀浆、抽提离心、Sephacryl S-300凝胶过滤等方法对可口革囊星虫纤溶酶初步分离,用纤维蛋白平板法和合成发色底物法检测其纤溶活性,并用该酶进行体外溶血凝块实验。结果可口革囊星虫内脏中存在纤溶酶。此酶既直接降解纤维蛋白又间接激活纤溶酶原,它对体外血凝块有明显溶解作用。结论可口革囊星虫内脏中存在着一种既具直接降解纤维蛋白作用又具激活纤溶酶原作用的纤溶酶。     

可口革囊星虫体液细胞的显微和亚显微结构

本文利用光学显微镜和透射电镜研究了可口革囊星虫体液细胞的显微和亚显微结构。研究发现该星虫体液中含有红细胞、颗粒细胞、桑椹胚样细胞和卵母细胞。可口革囊星虫的红细胞数量较多,红细胞呈盘状,从一边或两边略凹陷,最典型的是从三边发育的二裂片内陷,直径为24μm-32μm。圆形红细胞,核位于中央,细胞内很少有细胞器,细胞质内充满嗜锇的血红蛋白。体腔液内含有两种类型的颗粒细胞。Ⅰ型细胞含有大量的小颗粒,是典型的变形细胞,细胞质里含有大量的线粒体、最典型的高尔基体、内质网和各种形状不一的小池。Ⅱ型细胞含有大量的嗜锇细胞质。在可口革囊星虫体液中含有不同发育时期的卵母细胞。     

可口革囊星虫胚胎与幼体的发育

解剖经降温8-10℃刺激30-40h后的可口革囊星虫(Phascolosomaesculenta)亲体取得精、卵,用人工授精法获得的受精卵进行胚胎和幼体发育的研究。结果表明:可口革囊星虫的卵为圆形或椭圆形,卵的大小为120μm×142μm-125μm×148μm。精子由头部、中段和尾部组成,全长约为32-40μm。在海水盐度23.8ppt、pH8.1、温度15℃的条件下,精、卵不能受精;在海水盐度23.8ppt、pH8.1、温度20℃的条件下,精卵能受精,但胚胎发育到原肠胚期停止;在海水盐度23.8ppt、pH8.1条件下,25℃、30℃和35℃的不同水温,精卵均能受精,受精卵发育到初期海球幼虫分别需要72h0min、47h08min和42h53min。在海水盐度23.8-25.5ppt,pH8.1-8.2,温度28-30℃条件下,从初期海球幼虫发育到稚虫需要15-20d。     

可口革囊星虫受精过程及早期卵裂的细胞学变化

为探究可口革囊星虫(Phascolosoma esculenta)受精过程中精子入卵、极体排放、雌雄原核的形成与结合以及早期卵裂的特点,为胚胎发育机制研究奠定基础及指导人工育苗,显微观察了可口革囊星虫卵母细胞的形态;用荧光染色剂HOECHST33258对已固定的成熟卵及受精卵进行染色的方法,在荧光显微镜下观察了可口革囊星虫受精过程及第一与第二次卵裂的细胞学变化。成熟卵呈椭圆形,卵膜较厚,核区偏位,染色体排列整齐。在水温30℃-31℃、盐度23条件下,授精后5min-10min,精子完成入卵;授精后10min-20min,完成第一次减数分裂、释放第一极体;授精后20min-30min,完成第二次减数分裂、放出第二极体,部分受精卵的第一极体分裂为二;授精后30min-40min,雌、雄原核形成,并相互靠近向卵中央迁移;授精后40min-50min,雌、雄原核在卵中央结合成合子核;授精后50min-70min及70min-90min分别完成第一及第二次卵裂。可口革囊星虫的受精过程及早期卵裂的细胞学特点为:1)成熟卵是处在第一次减数分裂中期的初级卵母细胞,具有受精能力;2)精子入卵位点是随机的,存在多精入卵现象;3)雌、雄原核以融合方式结合成合子核;4)第一及第二次卵裂均为经裂、不等裂。     

可口革囊星虫肾管纤毛的分布及结构特征

为了解可口革囊星虫(Phascolosoma esculenta)肾管纤毛的结构特点及其功能,采用显微及亚显微技术观察研究了可口革囊星虫肾管纤毛的分布位置及形态结构特征。结果表明,肾管外膜多纤毛细胞表面簇生纤毛、内部柱状上皮细胞与立方上皮细胞游离面着生分散的纤毛,肾口内面也着生纤毛。纤毛结构由纤毛干、过渡区、基体及其纤毛小根组成;纤毛干由"9+2"结构的轴丝外被纤毛膜构成;纤毛干与基体之间为过渡区,中央微管终止于此,外周双联微管通过过渡区和基体的外周轴丝相连;基体呈圆筒状,为"9+0"结构;纤毛小根分长根和短根,均为基体发出的由微细原纤维组成的圆锥形结构,具间隔70 nm的明显横纹。肾管纤毛可能在促进体腔液流动、提高肾管对体腔液的过滤作用以及引导成熟精卵进入肾管等方面发挥作用。     

Different physiological functions of free D- and L-alanine in three body parts of the intertidal sipunculid Phascolosoma arcuatum

Free D- and L-alanine contents were comparable in the body wall and introvert cum retractor muscles of Phascolosoma arcuatum. In contrast, the content of free D-alanine in the internal organs was twice that of free L-alanine. Since alanine aminotrans-ferase from P. arcuatum was L-alanine specific, D-alanine appeared to be synthesized from L-alanine through the action of alanine racemase. Alanine racemase activity was higher in the D-alanine-forming direction in the three body parts of P. arcuatum. In addition, the ratio of DL/LD racemase activity in the internal organs was the lowest among the body parts studied. These results indicate that free D-alanine might be of lesser importance than the free D-isomer to the internal organs as compared to the body wall and introvert cum retractor muscles. Indeed, L-alanine inhibited pyruvate kinase from the body wall and introvert cum retractor muscles but had no effect on the pyruvate kinase from the internal organs. Furthermore, the activity of alanopine dehydrogenase present in the internal organs was significantly lower than those of the body wall and introvert cum retractor muscles. L-Alanine was an essential substrate for alanopine formation in the body wall and introvert cum retractor muscles during hypoxia since alanopine dehydrogenases from these body parts were L-alanine specific. When P. arcuatum was confronted with hypo-osmotic stress, the free D-alanine/total free alanine ratio in the internal organs increased approximately from 0.6 to 0.8 as the total free alanine content decreased. In comparison, those ratios in the body wall and introvert cum retractor muscles remained relatively constant. It was concluded that D- and D-alanine had different physiological functions in the three body parts of P. arcuatum.Abbreviations ADP adenosine-5-diphosphate - ADH alanopine dehydrogenase - ALT alanine aminotransferase - AOD amino acid oxidase - BW body wall - EDTA ethylenediaminetetra-acetic acid - EGT A ethylene glyco-bis (-aminoethyl ether) - N,N,N,N tetra-acetic acid - ICRM introvert cum retractor muscles - IO internal organs - I ₅₀ inhibitor concentration producing 50% inhibition of enzyme activity - -KG -ketoglutarate - LDH lactate dehydrogenase - NAD nicotinamide adenine dinucleotide - NADH nicotinamide adenine dinucleotide (reduced form) - PEP phosphoenolpy-ruvate - PEPCK phosphoenolpyruvate carboxykinase - PK pyruvate kinase - PMSF phenylmethylsulphonyl fluoride - SE standard error - SW sea water - TCA trichloroacetic acid     

Detoxification of environmental sulfide to sulfane sulfur in the intertidal sipunculid Phascolosoma arcuatum

The capability of Phascolosoma arcuatum to detoxify sulfide in anaerobic conditions was examined. Sulfane sulfur, which underwent cold cyanolysis, was the major excretory end product of sulfide detoxification during anoxia. Thiosulfate was not excreted into the external medium. Instead, it was absorbed by P. arcuatum and its absorption was stimulated by the presence of sodium sulfide (Na₂S) in the incubation medium. The effective formation and excretion of sulfane sulfur by P.␣arcuatum required the presence of both Na₂S and sodium thiosulfate (Na₂S₂O₃). Results obtained indicate that rhodanese might be involved in sulfide detoxification in this sipunculid. Rhodanese could act as a catalyst in the transfer of sulfur atoms from thiosulfate to HS⁻. The body wall and the introvert were the main sites of sulfide detoxification. However, it is unlikely that epibiotic bacteria associated with the outside surface of the worm were involved in the detoxification process. A time-course study on the contents of thiosulfate and sulfane sulfur in the body wall of P. arcuatum incubated anaerobically in the presence of Na₂S + Na₂S₂O₃ verified that thiosulfate absorbed was utilized to detoxify sulfide to sulfane sulfur. Accepted: 24 October 1996     

中缅树鼩精子形态特征及冷冻损伤

树鼩作为一种新型的、接近灵长类的实验动物,在医学生物学上的应用受到越来越多的重视。精子的结构特性研究及冷冻后结构的完整性分析是精子生物学的主要内容,也有助于树鼩的实验室快速繁殖。该研究采用人工饲养的中缅树鼩(Tupaia belangeri chinensis),结果显示其睾丸占总体重的(1.05±0.07)%,总体积为(1.12±0.10)mL。附睾尾及输精管精子总量估计在2.2×107～8.8×107,其运动度和顶体完整率分别为(68.8±3.9)%和(90.0±2.1)%。利用扫描电子显微镜和透射电子显微镜对树鼩附睾精子的超微结构进行的观察和分析显示精子头部呈圆形或卵圆形;头部长度、宽度平均分别为6.65和5.82μm;精子尾部中段、主段、尾段和精子总长度平均分别为13.39、52.35、65.74和73.05μm;尾部中段的线粒体螺旋数量为48个,其轴丝结构为典型的"9+9+2"结构。冷冻解冻后的精子主要表现在顶体与质膜不完整、精子断裂、尾部扭曲和膨大。上述结果提示树鼩精子与其他哺乳动物精子的结构特征相似,但是精子大小和线粒体螺旋数目有明显的差别,且超微结构改变仍是冷冻精子运动和受精能力下降的主要原因。     

A light and electron microscopic study of the ciliated urn of Phascolosoma agassizii (sipunculida)

Summary The free swimming ciliated urn found in the coelomic fluid of Phascolosoma agassizii has been studied by electron microscopy. The urn is a multicellular structure composed of three cell types: (a) ciliated cells which possibly function in capturing cell debris and foreign particles; (b) cupola cells which are capable of phagocytozing latex particles; and (c) lobe cells which are capable of phagocytozing carbon particles. The lobes are separated from the ciliated cells by a semilunar area, with mucoprotein staining characteristics, containing fibrils which appear to be the structural support for the urn. Ciliated cells and lobes are attached to the semilunar area by hemidesmosomes.This work is based on a thesis submitted in partial fulfillment of the requirements for the degree Master of Arts in the Department of Biology, San Francisco State University, San Francisco, California, U.S.A.I wish to thank Dr. John C. Lee, formerly of the Department of Pathology, University of California Medical Center, San Francisco, for encouragement and use of his electron microscope facilities     

多穗金粟兰营养器官的形态结构及组织化学研究

采用植物解剖学和组织化学定位方法研究多穗金粟兰（Chloranthus multistachysPei）营养器官的显微结构及萜类和黄酮类化合物在营养器官中的分布规律。结果表明：多穗金粟兰的根皮层细胞排列不整齐,液泡化明显,细胞间隙较大,有散在石细胞分布;茎皮层细胞排列不整齐,在韧皮部外方包围有少数几层束状纤维束;叶为异面叶,栅栏组织和海绵组织分化明显,叶片上、下表皮均无毛被,气孔器多为不规则型。组织化学定位结果显示萜类和黄酮类化合物分布在根的皮层、茎的皮层及韧皮部细胞中。