Dynamics of F-actin during spermiogenesis in Gampsocleis gratiosa ( Orthoptera: Tettigoniidae)

为阐明F-肌动蛋白在优雅蝈螽Gampsocleis gratiosa Brunner von Wattenwyl精子形成过程中的动态变化,本研究利用微分干涉相衬技术和免疫荧光技术首次对优雅蝈螽精子形成过程中的F-肌动蛋白进行了细胞定位,利用透射电镜技术从超微水平观察了优雅蝈螽精子顶体复合体的结构.结果显示:精子形成早期,F-肌动蛋白富集于亚顶体区域,形态由“球状”转变为“棒锥状”;精子形成中期,F-肌动蛋白呈“倒Y型”分布于亚顶体区域和细胞核前端两侧;精子形成后期,亚顶体区域的F-肌动蛋白解聚消失,F-肌动蛋白呈“箭头状”,仅分布于顶体复合体扩张的两翼中.F-肌动蛋白动态变化伴随着细胞核和精子头部的形态改变,F-肌动蛋白的动态装配在精子顶体复合体形态构建和细胞核的形变中起着重要的作用.本研究还发现未成熟的精子尾部有一些富含F-肌动蛋白的细胞质微滴,与精子形成过程中多余细胞质和细胞器的外排有关.F-肌动蛋白的动态变化研究为进一步阐明细胞骨架蛋白在昆虫精子形成过程中的功能和作用机制奠定了基础.     

优雅蝈螽精子形成过程中核的形态结构变化观察（英文）

【目的】螽斯精子结构复杂,具有特征性的箭头状顶体,是研究昆虫精子形成的理想材料。为了研究螽斯精子形成过程中的动态变化机制,特别是细胞核的凝集机制和箭头状顶体的发生机制,本研究对优雅蝈螽Gampsocleis gratiosa精细胞和精子的细胞核进行了观察。【方法】选择发育良好的优雅蝈螽成虫精巢为研究材料,利用透射电镜技术、普通光学显微镜和荧光显微镜技术,制作光镜切片和电镜切片进行观察。【结果】根据其形态结构变化特征,将优雅蝈螽精子形成过程中的细胞核分为4个阶段：圆形核、叶形核、柱状核和成熟阶段。我们还通过常规HE染色,结合DNA特异性荧光探针DAPI,证明了圆形核时期,精细胞内具有两个明显的球状结构,一个为细胞核,另一个是原顶体;精子成熟阶段,精子尾部排出的细胞质微滴中含有DNA。【结论】优雅蝈螽精子形成过程中,精细胞的细胞核经历了显著的形态变化,精细胞核的形态变化与细胞骨架微管相关,细胞核塑形伴随着染色质的重组。本研究为进一步阐明直翅目昆虫精子形成的分子机制奠定了基础。     

优雅蝈螽精子形成期间微丝和微管蛋白的动态变化

【目的】分析微丝和微管蛋白在优雅蝈螽Gampsocleis gratiosa精子形成过程中的作用,为昆虫精子顶体复合体形成和细胞核重建机制研究奠定基础。【方法】应用免疫荧光、PAS-苏木精染色和透射电镜等方法,对优雅蝈螽成虫的精巢、雄性贮精囊和雌性受精囊内精子的发育以及微丝和微管蛋白在精子形成各个时期的分布进行了观察。【结果】精巢中,早期圆形精子细胞中微丝在精子细胞的某区域大量聚集,而微管蛋白随机分布在细胞质中。伸长的精子细胞中,顶体开始形成时,微丝首先在亚顶体区域出现,历经球形、短棒形,然后向细胞核的两侧扩展成倒"Y"形,接着形成箭头形;在顶体的外周即微丝的周围,细胞核周围以及鞭毛中发现微管蛋白。在雄虫贮精囊和雌虫受精囊中,精子和精子束中仅有微管存在,且仅存在于鞭毛中;精子头部的微丝和微管蛋白均消失。【结论】综合分析,我们认为微丝和微管作为"脚手架"结构在优雅蝈螽精子形成期间参与顶体复合体形成和细胞核重建,精子成熟形成精子束过程中"脚手架"结构拆除。     

暗褐蝈螽与优雅蝈螽精子超微结构的比较研究(直翅目,螽斯科)

研究了暗褐蝈螽Gampsocleis sedakovii(Fischer von Waldheim)和优雅蝈螽G.gratiosa Brunner von Wattenwyl精子的超微结构。这两种蝈螽精子头部的顶体复合体由顶体外层、顶体本体和顶体组成,顶体复合体位于细胞核前端,并包裹部分细胞核;颈部具5纵层细胞器;尾部鞭毛轴丝为典型的9+9+2型,线粒体衍生体部分晶状化。暗褐蝈螽精子较短,顶体复合体夹角较大,精子鞭毛横切面直径稍大;优雅蝈螽精子稍长,顶体复合体夹角较小,精子鞭毛横切面直径较小,两种精子超微结构差异不显著,其生殖隔离机制有待进一步研究。     

中华蝈螽与中华螽斯精子超微结构研究(直翅目,螽斯总科)

研究了中华蝈螽Gampsocleis sinensis与中华螽斯Tettigonia chinensis精子的超微结构.两种精子的顶体复合体侧生于核上且包裹了核的一部分,顶体复合体横切面两侧角分离或两侧角与细胞核远离,或仅见其一侧角;中华蝈螽顶体外层在顶体本体两侧角外多呈晕圈状;轴丝为典型的9 9 2型;轴丝两侧的副纤维结构为2副微管;轴丝与线粒体衍生体之间的连接带3条;中华蝈螽有的横切面中两扁平膜池连接在一起;两种精子的中心粒侧体部位具五纵层细胞器.     

优雅蝈螽雄性附腺的超微结构及其腺管提取物对精子束的作用

为阐明优雅蝈螽Gampsocleis gratiosa Brunner von Wattenwyl雄性附腺的结构与功能的关系, 本文利用透射电镜(transmission electron microscope, TEM)技术研究了优雅蝈螽雄性附腺的超微结构, 利用微分干涉相差显微镜(differential interference contrast microscope, DIC)技术并结合雄性附腺匀浆提取物与精子束在体外的短暂培养, 研究了优雅蝈螽雄性附腺对精子束的作用。结果表明： 优雅蝈螽雄性附腺3类腺管组织结构相似, 腺管管壁为单层上皮细胞, 缺少内表皮, 说明其来源于中胚层。上皮细胞富含粗面内质网、 线粒体、 高尔基体, 具有分泌细胞的特点。腺管管腔中分泌物有4种形态, 即电子透明的物质、 电子致密的颗粒物质、 细纤维状物质以及绒球状物质。上皮细胞的分泌方式主要有2种, 即顶质分泌和局部分泌。乳白短腺管的匀浆提取物参与了帽状精子束解聚的过程, 乳白长腺管和透明腺管的匀浆提取物有维持精子束活性的作用。本研究结果为进一步阐明螽斯雄性附腺的生理功能奠定了基础。     

优雅蝈螽与暗褐蝈螽精子束的显微观察

本文应用微分干涉相衬法对优雅蝈螽Ｇampsocleis gratiosa Brunner von Wattenwyl和暗褐蝈螽G. sedakovii （Fischer von Waldheim） 雄性精巢管基部、输精管、贮精囊和精包,及雌性受精囊中精子束的形态变化进行了观察,对探讨螽斯近缘种的生殖隔离机制和生殖生物学具有重要意义.结果表明：这两种蝈螽的精子束通过精包转移到雌性受精囊后,精子束的形态发生了显著变化.精巢管基部的精子为游离的单个精子;输精管、贮精囊和精包中精子成束排列形成较分散的精子束,精子束头部包裹有粘液帽;雌性受精囊中的精子束的精子呈羽状排列,精子的头部汇集在中央轴上.两种蝈螽精子束形态差异不显著.     

香螺精子发生及精子超微结构

本文采用透射电镜技术对香螺(NpatunedecumingiCrosse)精子发生过程进行了观察。结果表明,精原细胞胞质中含有大量的线粒体;初、次级精母细胞的细胞核和大量的线粒体呈极性分布;精子细胞分化过程中,细胞核形态、核内物质以及线粒体的形态发生显著变化;细胞核的核质由不均匀颗粒状浓缩成纤丝状,再浓缩成细线形,最后呈致密均匀状态,细胞核由近圆形伸长为粗线形,具有核后窝;在细胞核后端有8个膨大的线粒体,由卵圆形变为螺旋形,弯曲盘绕在轴丝外部,形成精子的中段;根据细胞核和线粒体的变化特点,将精子形成分为早、中、后三个时期。香螺典型性精子属于进化型,头部呈线形,中段加长,糖原颗粒包围轴丝构成主段。在精子发生过程中,细胞质内没有发达的高尔基复合体和前顶体池,没有观察到香螺精子的顶体。在成熟个体的精巢内,同时存在不具有受精能力的畸变精子。     

无蹼壁虎精子头部形成的研究

本文用透射电镜观察了无蹼壁虎精子头形成的过程。早期精细胞具有显著的高尔基复合体、线粒体集合及细胞质桥、接着高尔基体成熟面分泌出前顶体囊泡,并逐渐向核移动。以后精子形成可分四个时间：时间Ⅰ,当前顶体囊泡移至核膜时,核膜凹陷形成封闭的顶体囊泡,囊泡底部靠近核膜有一电子致密的顶体颗粒;时间Ⅱ,细胞核延长,顶体囊泡变扁平;时期Ⅲ,细胞核进一步延长,核内染色质纤维变粗并沿核纵轴方向排列有序;时间Ⅳ,精子发育     

精子形成中的表观遗传调控

精子形成是指单倍体球形精子细胞分化发育成为精子的过程。这一连续进程包含一系列复杂的生化事件和剧烈的形态变化,涉及顶体和尾部形成、组蛋白-鱼精蛋白转换、细胞核压缩和细胞质丢弃等。近期研究发现,表观遗传调控在精子形成过程中发挥重要作用,对确保精子细胞正常发育和精子生成至关重要。现总结近期的相关研究进展,从DNA甲基化和组蛋白修饰两个方面简介精子形成过程中的表观遗传调控功能和机制。     

The role of sertoli cells in spermatid maturation in the testis of the crayfish, Procambarus paeninsulanus

With the onset of spermiogenesis, many changes become apparent in the crayfish spermatid during its transition to mature sperm. The nucleus passes through a series of stages, excess cytoplasm is removed, the acrosome develops, and nuclear arms form and become wrapped around the sperm prior to its enclosure in a capsule. Changes are also apparent in the Sertoli cells surrounding the germ cells in the crayfish testis. The amount of cytoplasm of individual Sertoli cells appears to increase in quantity and changes in the intracellular organelles become apparent. As spermiogenesis commences, the cytoplasm along one side of Sertoli cells adjacent to the spermatids is devoid of obvious organelles. Numerous finger/like projections of Sertoli cytoplasm penetrate into the spermatid and appear to isolate portions of the sperm cytoplasm. During later stages of spermiogenesis, several vesicles in the Sertoli cells which appear to contain droplets of this isolated sperm cytoplasm. appear to undergo lytic changes, As the amount of cytoplasm of the spermatid is reduced, contact is maintained between the spermatid and Sertoli cell in the area of the acrosome. The nuclear arms of the sperm extend into the Sertoli cell during their formation and later become wrapped around the acrosomal area of the sperm. At this time, very little space exists between the Sertoli cell and its many sperm. Large vesicles of electron dense material appear to be released by the Sertoli cells into the space between the sperm and Sertoli cell. This material completely surrounds the sperm and forms the sperm capsule. Spermiation involves the gradual dissolution of the points of contact between the sperm capsule and the Sertoli cell.     

Immunogold distribution of actin during sperimiogenesis in the normal rabbit and after experimental cryptorchidism

Immunogold procedures for actin detection were used in combination with experimental cryptorchidism in the rabbit as a modei to shed more light on the function of subacrosomal actin during spermiogenesis. In the normal testis, actin was localized in the perinuclcar substance (PNS) from round spermatid onward but it was not detected in late spermatids. Actin labeling in each type of spermatid was essentially unmodified after 24 hr of cryptorchidism. However, among relevant immediate and delayed effects, discontinuous acrosomes overlying a continuous PNS with normal actin labeling were noted. Nuclear invaginations were seen in combination with subacrosomal dilatations: at this site actin labeling was found only in the PNS closely apposed to the nuclear envelope. In subacrosomal areas lacking PNS, actin labeling also was lacking. These results suggest that the subacrosomal actin (F-actin) is a component of the PNS that is tightly bound to the nuclear envelope rather than the overlying inner acrosomal membrane. Therefore, a function for the subacrosomal actin either in anchoring the acrosome to the nucleus or in capping the inner acrosomal membrane appears unlikely. The data rather suggest a capping function for the nuclear membrane during spermiogenesis.     

Characterization of filaments within the subacrosomal space of rat spermatids during spermiogenesis

Two types of filaments were observed within the subacrosomal space of rat spermatids. The first of these types was characterized as actin by demonstration of actin filament affinity for myosin S-1 subfragments. Actin filaments were noted in the subacrosomal space shortly after the acrosomal sac made contact with the nucleus. As the acrosome increased its surface area contact with the spermatid nucleus, the number of layers of subacrosomal filaments increased. Pre-treatment with detergent, which in addition to permeablizing cells to allow entry of S-1, also caused the acrosome to vesiculate and the subacrosomal space to widen. In such preparations filaments were more easily visualized and appeared to extend between the nuclear and acrosomal membranes, indicating, but not proving, attachment to these membranes. During spermatid clongation, the number of actin filaments in the subacrosomal space increased greatly, especially over the dorsal convex region of the spermatid head. The polarity of the majority of filaments was not ascertainable since filaments were tightly packed within the narrow subacrosomal space. In late spermiogenesis (steps 18 and 19), actin filaments were no longer detected within the subacrosomal space. A second and much thicker type of filamentous structure was observed in the subacrosomal space of spermatids at steps 14-17 of spermiogenesis. About 14 nm in diameter (10-15 nm measurement range depending on fixation protocol utilized), these filaments did not decorate with myosin S-1 subfragments and were found in subacrosomal regions not containing actin. Fourteen nanometer filaments were seen in parallel array along the ventral folded portion of the nuclear membrane and extended partially around the nucleus. Like actin filaments. 14 nm filaments were not seen in the subacrosomal space during late spermiogenesis.     

Acroplaxome, an F-actin-keratin-containing plate, anchors the acrosome to the nucleus during shaping of the spermatid head

Nuclear shaping is a critical event during sperm development as demonstrated by the incidence of male infertility associated with abnormal sperm ad shaping. Herein, we demonstrate that mouse and rat spermatids assemble in the subacrosomal space a cytoskeletal scaffold containing F-actin and Sak57, a keratin ortholog. The cytoskeletal plate, designated acroplaxome, anchors the developing acrosome to the nuclear envelope. The acroplaxome consists of a marginal ring containing keratin 5 10-nm-thick filaments and F-actin. The ring is closely associated with the leading edge of the acrosome and to the nuclear envelope during the elongation of the spermatid head. Anchorage of the acroplaxome to the gradually shaping nucleus is not disrupted by hypotonic treatment and brief Triton X-100 extraction. By examining spermiogenesis in the azh mutant mouse, characterized by abnormal spermatid/sperm head shaping, we have determined that a deformity of the spermatid nucleus is restricted to the acroplaxome region. These findings lead to the suggestion that the acroplaxome nucleates an F-actin-keratin-containing assembly with the purpose of stabilizing and anchoring the developing acrosome during spermatid nuclear elongation. The acroplaxome may also provide a mechanical planar scaffold modulating external clutching forces generated by a stack of Sertoli cell F-actin-containing hoops encircling the elongating spermatid nucleus.     

Ultrastructural differentiation of spermiogenesis in Scincus scincus (Scincidae,Reptilia)

BackgroundKnowledge of spermiogenesis in reptiles, especially in lizards, is very limited. Lizards found in Arabian deserts have not been considered for detailed studies due to many reasons and the paucity of these animals. Therefore, we designed a study on the differentiation and morphogenesis of spermiogenesis, at an ultrastructural level, in a rare lizard species, Scincus scincus.ResultsThe spermiogenesis process includes the development of an acrosomal vesicle, aggregation of acrosomal granules, formation of subacrosomal nuclear space, and nuclear elongation. A role for manchette microtubules was described in nuclear shaping and organelle movement. During head differentiation, the fine granular chromatin of the early spermatid is gradually replaced by highly condensed contents in a process called chromatin condensation. Furthermore, ultrastructural features of sperm tail differentiation in S. scincus were described in detail. The commencement was with caudal migration toward centrioles, insertion of the proximal centriole in the nuclear fossa, and extension of the distal centrioles to form the microtubular axoneme. Subsequently, tail differentiation consists of the enlargement of neck portion, middle piece, the main and end pieces.ConclusionsThis study aids in the understanding of different aspects of spermiogenesis in the lizard family and unfurls evolutionary links within and outside reptiles.     

The subacrosomal granule and its evolution during spermiogenesis in a lizard

Summary Some aspects of spermiogenesis have been studied in the testis of the teiid lizard Cnemidophorus lemniscatus lemniscatus by electron microscopy. Shortly after the acrosomal vesicle is lodged in a nuclear concavity of the spermatid, a dense granule differentiates in the center of the subacrosomal space. It is cone-shaped and shows a longitudinal striation. Its base applies to the acrosomal membrane and, through this, to the acrosomal granule. Its rounded vertex causes a depression of the nuclear membranes which, initially juxtaposed, separates at this point to form a vesicle. The granule develops and becomes a rod when spermiogenesis is advanced and the subacrosomal space has taken the form of a secondary cap. The rod is cylindrical, retains its original striation and has a convex acrosomal end. It encloses the vesicle formed by the nuclear envelope in its base and follows the apex of the nucleus. Meanwhile, the acrosomal granule loses its identity and the acrosomal cap is filled with a dense substance, in which a fringe of translucent material differentiates. This fringe lies in the dorsal and apical margins of the acrosome and is incompletely divided by longitudinal crests of the dense acrosomal substance. A projection of the Sertoli cell forms an accessory cap which envelops the acrosome and is in turn covered by the cytoplasm of the spermatid, constituting an intricate association. Two reflex membranes underlie the plasmalemma in the outer surface of the projection of the Sertoli cell. They are continuous with one another at their ends and with the cell membrane in the edge of pores. In the peripheral cytoplasm of the spermatid facing the accessory cap, numerous microtubules run longitudinally. By means of thin membranes some are interconnected or connected with the plasmalemma, from which they seem to originate.This research forms part of project N. 31.26.S1-0244 supported by the Consejo Nacional de Investigaciones Científicas y Tecnológicas     

Spermiogenesis in the bullfrog (Rana catesbeiana): a study of cytoplasmic events including cell volume changes and cytoplasmic elimination

The process by which spermatid cytoplasmic volume is reduced and cytoplasm eliminated during spermiogenesis was investigated in the bullfrog Rana catesbeiana. At early phases of spermiogenesis, newly formed, rounded spermatids were found within spermatocysts. As acrosomal development, nuclear elongation, and chromatin condensation occurred, spermatid nuclei became eccentric within the cell. A cytoplasmic lobe formed from the caudal spermatid head and flagellum and extended toward the seminiferous tubule lumen. The cytoplasmic lobe underwent progressive condensation whereby most of its cytoplasm became extremely electron dense and contrasted sharply with numerous electron-translucent vesicles contained therein. At the completion of spermiogenesis, many spermatids with their highly condensed cytoplasm still attached were released from their Sertoli cell into the lumen of the seminiferous tubule. There was no evidence of the phagocytosis of residual bodies by Sertoli cells. Because spermatozoa are normally retained in the testis in winter and are not released until the following breeding season, sperm were induced to traverse the duct system with a single injection of hCG. Some spermatids remained attached to their cytoplasm during the sojourn through the testicular and kidney ducts; however, by the time the sperm reached the Wolffian duct, separation had occurred. The discarded cytoplasmic lobe (residual body) appeared to be degraded with the epithelium of the Wolffian duct. It was determined that the volume of the spermatid was reduced by 87% during spermiogenesis through a nuclear volume decrease of 76% and cytoplasmic volume decrease of 95.3%.     

Studies on the polymorphic spermatozoa of a marine snail. 2. Genesis of the eupyrene sperm

Spermiogenesis of the eupyrene sperm in the snail, Fusitriton oregonensis, was studied with light and electron microscopes. Endoplasmic reticulum, which encircles the nucleus in each spermatid, appears to connect with the Golgi body and to interconnect between adjacent spermatids via cytoplasmic bridges. It is suggested that as the Golgi body migrates around the nucleus the endoplasmic reticulum may circulate with it. The alignment of the proacrosome with the nucleus is effected by a 180° rotation of the Golgi body, after which it separates and migrates posteriorly with the residual cytoplasm. Each sperm possesses a well-developed intracellular digestive system as indicated by multivesicular bodies, residual bodies, and myeloid figures. Autophagy begins in the residual cytoplasm before it is released from the middle piece. Microtubules are found outside the nucleus and mitochondria during the final stages of spermiogenesis, when elongation is almost complete. These microtubules appear to be involved in the final shaping and twisting process, in which torsion is locked in the nucleus and the mitochondria spiral around the axoneme. The annulus attaches the distal centriole to the plasma membrane in the early spermatid and as flagellar production begins they move towards the implantation fossa at the base of the nucleus. There are two centrioles in the early spermatid, the distal centriole and procentriole. The small procentriole fuses with the distal centriole in the intranuclear canal to form the centriolar cap of the basal body. This cap is pushed through the end of the nuclear tube and is separated from the subacrosomal space by only the nuclear membranes.     

Spermiogenesis in the imbricate alligator lizard,Barisia imbricata (Reptilia,Squamata, Anguidae)

Although the events of spermiogenesis are commonly studied in amniotes, the amount of research available for Squamata is lacking. Many studies have described the morphological characteristics of mature spermatozoa in squamates, but few detail the ultrastructural changes that occur during spermiogenesis. This study's purpose is to gain a better understanding of the subcellular events of spermatid development within the Imbricate Alligator Lizard, Barisia imbricata. The morphological data presented here represent the first complete ultrastructural study of spermiogenesis within the family Anguidae. Samples of testes from four specimens collected on the northwest side of the Nevado de Toluca, México, were prepared using standard techniques for transmission electron microscopy. Many of the ultrastructural changes occurring during spermiogenesis within B. imbricata are similar to that of other squamates (i.e., early acrosome formation, chromatin condensation, flagella formation, annulus present, and a prominent manchette). However, there are a few unique characteristics within B. imbricata spermatids that to date have not been described during spermiogenesis in other squamates. For example, penetration of the acrosomal granule into the subacrosomal space to form the basal plate of the perforatorium during round spermatid development, the clover‐shaped morphology of the developing nuclear fossa of the flagellum, and the bulbous shape to the perforatorium are all unique to the Imbricate Alligator Lizard. These anatomical character differences may be valuable nontraditional data that along with more traditional matrices (such as DNA sequences and gross morphological data) may help elucidate phylogenetic relationships, which are historically considered controversial within Squamata. J. Morphol., 2013. © 2013 Wiley Periodicals, Inc.