Studies of oogenesis in the rotifer,Asplanchna

Summary Oocyte development in Asplanchna brightwelli was studied by observation through the transparent body wall of living females and by electron microscopy. During oogenesis, which requires four to six hours, the oocyte increases in volume approximately 1000-fold. Most of its cytoplasm appears to be derived from the vitellarium by direct flow through the cytoplasmic bridge. This flow is rapid enough to be easily observed in the living female at low magnifications. Ribosomes, mitochondria, cortical granules, and lipid droplets were observed in the bridge area in electron micrographs.RNA synthesis during oogenesis was studied by means of autoradiography. Very little synthesis could be demonstrated in oocyte nuclei at any period of oogenesis, whereas the vitellarium nuclei show active incorporation of ³H-uridine throughout the reproductive life of the adult female. Most of this RNA is subsequently transferred to developing oocytes.This research was supported by USPHS Grant GM 121183 to Dr. C. W. Birky, Jr.     

The spatial arrangement of germ line cells in ovarian follicles of the mutantdicephalic inDrosophila melanogaster

Summary The pattern of intercellular connections between germ line cells has been studied in follicles of the mutantdicephalic (dic), which possess nurse cell clusters at both poles. Staining of follicles with a fluorescent rhodamine conjugate of phalloidin reveals ring canals and cell membranes and thus allows us to reconstruct the spatial organization of the follicle. Each germ line cell can be identified by the pattern of cell-cell connections which reflect the mitotic history of individual cells in the 16-cell cluster. The results indicate that in both wild-type anddicephalic cystocyte clusters one of the two cells with four ring canals normally becomes the pro-oocyte. However, in some follicles (dicephalic and wild-type) oocytes were found with fewer or more than four ring canals. Indic follicles, one or several nurse cells may become disconnected from the other cells during oocyte growth at stage 9–10. Such disconnected cells cannot later on empty their cytoplasm into the oocyte. This, in turn, might be of consequence for the determination of axial polarity of the embryo.     

蕨配子体发育及卵发生的显微结构观察

运用显微观察技术对蕨(Pteridium aquilinum var. latiusculum)配子体发育和卵发生进行了研究。结果表明：(1)蕨孢子黄褐色,四面体形,具三裂缝,接种后3～7 d萌发,经丝状体和片状体阶段发育成原叶体,成熟原叶体雌雄异株或同株。(2)蕨颈卵器产生于生长点下方的表面细胞(颈卵器原始细胞),该细胞经2次分裂形成3层细胞,其上层和下层细胞发育为颈卵器壁细胞,中间细胞为初生细胞,它经2次不等分裂产生3个细胞,分别为卵细胞、腹沟细胞和颈沟细胞;刚产生时,此3个细胞紧贴颈卵器壁,细胞质内液泡较多,随着发育,卵细胞和腹沟细胞之间产生了分离腔,但二者通过孔区相连,在卵细胞上表面可观察到卵膜;此后,颈沟细胞和腹沟细胞逐渐退化,颈卵器壁细胞内具有黑色颗粒物质。连续切片观察发现,成熟卵细胞上表面中央具有受精孔。卵发生的细节尚需超微结构的研究。     

Identification of novel oocyte and granulosa cell markers

Here we present novel gene expression patterns in the ovary as part of an ongoing assessment of published micro-array data from mouse oocytes and embryos. We present the expression patterns of 13 genes that had been determined by micro-array to be expressed in the mature egg, but not during subsequent preimplantation development. In-situ hybridization of sectioned ovaries revealed that these genes were expressed in one of two distinct patterns: (1) oocyte-specific or (2) expressed in both the oocyte and surrounding granulosa cells. Despite the fact that micro-array data demonstrated expression in the egg, several of these genes are expressed at low levels in the oocyte, but strongly expressed in granulosa cells. Eleven of these genes have no reported function or expression during oogenesis, indicating that this approach is a necessary step towards functional annotation of the genome. Also of note is that while some of these gene products have been well characterized in other tissues and cell types, others are relatively unstudied in the literature. Our results provide novel gene expression information that may provide insights into the molecular mechanisms of follicular recruitment, oocyte maturation and ovulation and will direct further experimentation into the role these genes play during oogenesis.     

细鳞鱼卵子发生过程中细胞器的形态变化与作用

通过组织学方法和透射电镜技术对细鳞鱼(Brachymystax lenok)卵子发生过程中细胞器的变化与作用进行了研究.结果表明,从刚分化的卵原细胞至成熟卵母细胞时期(Ⅰ～Ⅴ时相)胞质内均能观察到线粒体,其形态最初为圆形,随着其大量增殖,形态变为棒状、弯曲状或长形(Ⅱ时相),并导致线粒体簇形成,其嵴也由单个变为多个,电子密度呈由低到高的规律变化;但到Ⅲ时相末期线粒体又退化为圆形,个别线粒体还通过对分或牙分进行裂变,线粒体嵴被不断释放,形成空泡,其基质电子密度呈降低的规律变化;在此过程中线粒体主要参与各种囊泡的形成,为后期卵黄前体物质进入、积累创造条件.在Ⅱ时相卵母细胞早期的细胞核附近开始出现内质网和高尔基体,但数量少,结构简单,随着它们的大量增殖(Ⅲ～Ⅳ时相),这两种细胞器将对卵黄物质的合成与加工起到关键作用.内质网主要呈弓形,少数呈圆形或杯形,早期与高尔基体相伴出现,但随着内质网大量增殖,其合成功能也随之增强.早期高尔基体也呈弓形,但随着其高度发育,几个分散的高尔基体聚集形成高度发育的高尔基体复合体,其加工与修饰功能也不断增强,同时其周围伴有大量潴泡或电子密度不同的囊泡体(多层结构)出现,且这些多泡体常常与环形片层(annulate lamellae,AL)一同出现.AL与核膜结构相似,早期呈弧形排列,本研究推测环形片层起源于核膜,其主要作用可能是膜的储藏地.     

Developmental profiles of PERIOD and DOUBLETIME in Drosophila melanogaster ovary

The clock protein PERIOD (PER) displays circadian cycles of accumulation, phosphorylation, nuclear translocation and degradation in Drosophila melanogaster clock cells. One exception to this pattern is in follicular cells enclosing previtellogenic ovarian egg chambers. In these cells, PER remains high and cytoplasmic at all times of day. Genetic evidence suggest that PER and its clock partner TIMELESS (TIM) interact in these cells, yet, they do not translocate to the nucleus. Here, we investigated the levels and subcellular localization of PER in older vitellogenic follicles. Cytoplasmic PER levels decreased in the follicular cells at the onset of vitellogenesis (stage 9). Interestingly, PER was observed in the nuclei of some follicular cells at this stage. PER signal disappeared in more advanced (stage 10) vitellogenic follicles. Since the phosphorylation state of PER is critical for the progression of circadian cycle, we investigated the status of PER phosphorylation in the ovary and the expression patterns of DOUBLETIME (DBT), a kinase known to affect PER in the clock cells. DBT was absent in previtellogenic follicular cells, but present in the cytoplasm of some stage 9 follicular cells. DBT was not distributed uniformly but was present in patches of adjacent cells, in a pattern resembling PER distribution at the same stage. Our data suggest that the absence of dbt expression in the follicular cells of previtellogenic egg chambers may be related to stable and cytoplasmic expression of PER in these cells. Onset of dbt expression in vitellogenic follicles coincides with nuclear localization of PER protein.     

Reproduction in a variable environment: How does Eupelmus vuilleti, a parasitoid wasp, adjust oogenesis to host availability?

Oogenesis of the parasitoid wasp Eupelmus vuilleti is known to be dependent on host availability. However, examination of ovarian dynamics by microscopy showed that oogenesis and vitellogenesis are initiated before female eclosion and proceed 1-2 days after, independent of host presence. Oogenesis continued beyond the 2nd day only in the presence of hosts, otherwise it was replaced by egg resorption. It is thus possible to distinguish between host-independent and host-dependent periods of oogenesis. In the presence of host, each ovariole (three per ovary) contained generally three oocytes: a fully mature oocyte, a nearly mature one and an immature one. However, host deprived-females resorbed their most mature and their smallest oocytes, but kept one almost mature oocyte per ovariole. Comparison of zero, short and long host deprivation periods showed that females always had the ability to quickly lay eggs to exploit any new host. However, increased deprivation led to a reduction in the number and the viability of eggs. Enzymo-immunological measurements of ecdysteroids were made in whole females, in dissected ovaries and in newly laid eggs. Our results indicated that ecdysteroids play a major role as circulating hormones involved in the regulation of oogenesis.     

Arrest of oogenesis in the bug Rhodnius prolixus challenged with the fungus Aspergillus niger is mediated by immune response-derived PGE2

In this work we characterized the immune response of the insect Rhodnius prolixus to a direct injection into the hemocoel of the non-entomopathogenic fungus Aspergillus niger, and evaluated its consequences on host oogenesis. These animals were able to respond by mounting effective cellular and humoral responses to this fungus; these responses were shown, however, to have reproductive fitness costs, as the number of eggs laid per female was significantly reduced. The disturbance of egg formation during infectious process correlated with an elevation in the titer of hemolymph prostaglandin E₂ 48 h post-challenge. Administration of Zymosan A as an immunogenic non-infectious challenge produced similar effects on phenoloxidase and prophenoloxidase activities, oocyte development and prostaglandin E₂ titer, precluding the hypothesis of an effect mediated by fungal metabolites in animals challenged with fungus. Ovaries at 48 h post-challenge showed absence of vitellogenic ovarian follicles, and the in vivo administration of prostaglandin E₂ or its receptor agonist misoprostol, partially reproduced this phenotype. Together these data led us to hypothesize that immune-derived prostaglandin E₂ raised from the insect response to the fungal challenge is involved in disturbing follicle development, contributing to a reduction in host reproductive output and acting as a host-derived adaptive effector to infection.