家蝇的卵黄发生及其激素调节

用5—15%SDS-PAGE分析表明,家蝇Musce domestica viaina卵黄蛋白由三个亚基组成,其亚基分子量分别为58KD、50KD、48KD.火箭免疫电泳的结果表明,脂肪体、血淋巴和卵巢内卵黄原蛋白的变化具有密切的相关性,卵黄原蛋白在体内最早出现在羽化后30小时左右,然后迅速增加,在羽化后48小时,脂肪体和血淋巴中卵黄原蛋白含量达到最大值,卵巢开始沉积卵黄蛋白在羽化后30小时,到产卵前达到最大值,脂肪体在离体培养条件下,通过测定³H-亮氨酸掺入卵黄原蛋白的量,对不同发育时期家蝇脂肪体合成卵黄原蛋白的能力及激素的调节作用进行了研究,结果表明,羽化12小时后,合成能力迅速上升,48小时时形成高峰,60小时后迅速下跌直至产卵,其合成能力一直维持在低水平,产卵后合成能力又迅速回升,激素处理结果表明,保幼激素可以促进卵黄发生前期和后期家蝇脂肪体的卵黄原蛋白合成,20-羟基蜕皮酮可以大幅度促进卵黄发生期家蝇脂肪体的卵黄原蛋白合成.当二种激素共同处理时,对卵黄发生前期和卵黄发生期的家蝇脂肪体有协同促进作用,而对卵黄发生后期的脂肪体没有这种作用.本文还对家蝇卵黄发生过程中脂肪体、血淋巴和卵巢三者之间的关系及家蝇卵黄发生的激素调节进行了讨论.     

早熟素II对家蝇卵黄发生的影响

本实验通过卵巢发育分级的解剖观察、可溶性蛋白质和核酸的定量测定、火箭免疫电泳定量测定卵黄原蛋白及激素处理等方法,研究了早熟素对家蝇(Muscadomestica vicina)卵黄发生的影响。试验结果表明用20ug早熟素处理每头刚羽化家蝇时,家蝇卵黄发生处于不完全抑制状态,其卵黄发生过程比对照组“延迟”约12小时。处理后48小时,血淋巴中卵黄原蛋白的滴度为lo.5ug/ul,接近对照组,而其卵巢鲜重和发育等级明显低于对照组,这种不完全抑制状态表明卵母细胞对卵黄原蛋白的吸收作用受到抑制。当用高剂量100ug早熟素11处理每头刚羽化家蝇时,血淋巴中卵黄原蛋白滴度、卵巢鲜重及其发育均受到明显的抑制,这种抑制效应能自然恢复。 当早熟素11和保幼激素(JH-III)、20-羟基蜕皮酮共同处理时,保幼激素具有明显的去抑制作用,可使血淋巴中卵黄蛋白浓度成倍增加,20-羟基蜕皮酮的去抑制效应不明显。本文还对早熟素作用于双翅目昆虫的方式作了讨论。     

家蝇卵巢摄取卵黄蛋白的机理

在家蝇Musca domestica viaina 的卵黄发生过程中,卵母细胞摄取卵黄原蛋白与滤泡开放是相关的。观察不同发育时期的家蝇滤泡结果表明,在摄取活动最旺盛的时期也就是卵黄发生的顶盛时期,其滤泡开放程度最大,而在卵黄发生前期和后期基本上没有摄取活动,此时的滤泡上皮细胞间不开放。卵巢体外培养的激素处理表明,JH可以促进滤泡开放。家蝇卵巢微粒体制备物的Na⁺-K⁺ATP酶活力在卵巢发育过程中存在着动态变化。羽化后24小时时有一定的酶活性,随着卵黄发生的进行,酶活力逐渐增加,到羽化48小时时酶活力最高,然后又开始下降,到羽化72小时时已经很小。羽化32小时的家蝇点滴或注躬 JH之后,测得的卵巢微粒体制备物的Na⁺-K⁺ATP酶活力比正常羽化36小时的高,羽化44小时的家蝇点滴和注射JH之后,测得酶活力比正常羽化48小时的低。羽化36小时和48小时的家蝇卵巢微粒体制备物与JH共同作用后,其Na⁺-K⁺ATP酶的活力分别增加2.95倍和3.50倍,羽化48小时的家蝇卵巢在含有JH的培养液中培养启,其匀浆液的酶活性为对照组的1.26倍。 由此我们可以推测在家蝇的卵黄发生过程中,JH通过促进滤泡开放和增加卵巢微粒体制备物Na⁺-K⁺ATP酶的活力,从而调控卵母细胞对卵黄蛋白的摄取。     

家蝇卵巢摄取卵黄蛋白的机理

在家蝇Ｍｕｓｃａ　ｄｏｍｅｓｔｉｃａ　ｖｉａｉｎａ的印黄发生过程中,卵母细胞摄取卵黄原蛋白与滤沟开放是相关的。观察不同发育时期的家蝇滤泡结果表明,在摄取活动最旺盛的时期也就是卵黄发生的顶盛时期,其滤泡开放程度最大,而在卵黄发生前期和后期基本上没有摄取活动,此时的滤泡上皮细胞间不开放。卵巢体外培养的激素处理表明,ＪＨ可以促进滤泡开放。家蝇卵巢微粒体制备物的Ｎａ＋－Ｋ＋ＡＴＰ酶活力在卵巢发育过程中存在着动态变化。羽化后２４小时时有一定的酶活性,随着卵黄发生的进行,酶活力逐渐增加,到现化４８小时时酶活力最高,然后又开始下降,到弱化７２小时时已经很小。羽化３２小时的家蝇点滴或注射ＪＨ之后,测得的卵巢微粒体制备物的Ｎａ＋－Ｋ＋ＡＴＰ酶活力比正常羽化３６小时的高,羽化率４４小时的家蝇点滴和注射ＪＨ之后,测得酶活力比正常羽化４８小时的低．羽化３６小时和４８小时的家蝇卵巢微粒体制备物与ＪＨ共同作用后,其Ｎａ＋－Ｋ＋ＡＴＰ酶的活力分别增加２．９５倍和３．５０倍,羽化４８小时的家蝇卵巢在含有ＪＨ的培养液中培养后,其匀浆液的酶活性为对照组的１．２６倍。由此我们可以推测在家蝇的卵黄发生过程中,ＪＨ通过促进滤泡开放和增加卵巢微粒体制     

天蚕卵黄原蛋白的合成、运转与沉积

系统测定了天蚕Antheraea yamamai吐丝结茧至成虫期脂肪体、血淋巴和卵巢中卵黄蛋白和可溶性蛋白总含量的动态变化。结果表明,脂肪体是卵黄原蛋白(Vg)合成场所,Vg合成始于吐丝结茧后第4天;脂肪体、血淋巴中Vg滴度在吐丝结茧后第4天开始上升,化蛹后第6天或第8天达高峰,成虫羽化第1天则明显下降。卵巢对Vg摄取始于化蛹第1天,此后随蛹日龄逐渐上升,并渐趋平稳。同一卵巢管中卵黄蛋白(Vt)含量自顶端至基端随卵室增大而逐渐升高,不同日龄蛹中相应序号卵室的Vt含量以日龄大者为高;卵室中Vt含量与卵室体积大小呈正线性关系。电镜观察表明,Vg被卵母细胞摄入后以卵黄体形式存在,不同发育阶段卵巢中卵母细胞内卵黄体大小不同,以早期者为小;同一卵巢管中不同卵母细胞内卵黄体以顶端为小,基端明显增大,且卵黄体呈网状。     

七星瓢虫雌成虫咽侧体的活性

以短期体外放射化学测定法测定了七星瓢虫雌成虫的咽侧体(CA)活性。结果表明,七星瓢虫的CA在TC 199培养液中培养时活性最高。在最适培养条件下CA合成保幼激素(JH)的速度在1—4小时呈直线增加。 七星瓢虫雌虫生殖期CA的活性变化与卵黄发生有相关性。羽化初期CA活性很低,羽化后4—8天CA活性增加,卵母细胞内卵黄沉积开始增多;羽化后8—12天CA活性高峰出现,此时卵母细胞内有大量卵黄沉积;羽化后12—15天CA活性下降,卵完全成熟并陆续出现产卵个体。 食料对七星瓢虫成虫的卵黄发生的影响:取食人工饲料雌虫的CA活性增长缓慢,直至羽化后15天CA活性仍很低,因而抑制了卵黄原蛋白的合成,使卵巢发育缓慢。此种雌虫中CA活性高峰的出现比取食蚜虫的延迟约2倍,前者的产卵前期也较后者延长约2倍。     

小体鲟卵黄蛋白生化特性及合成途径的研究

使用葡聚糖凝胶（Sephadex G-200）从小体鲟鱼卵粗提液中,提纯卵黄脂磷蛋白(Lipovitellin, Lv )和卵黄高磷蛋白(Phosvitin, Pv)。卵黄脂磷蛋白（含糖、磷和脂,等电点7.50）具有雌性特异性, 分子量为144 kD,由97.4 kD和30 kD的大小2个亚基组成。卵黄高磷蛋白（含糖、磷,等电点8.30）其分子量为66 kD,其具有两个亚基,分子量分别为47.6 kD、16.8 kD。制备卵黄脂磷蛋白兔抗血清,采用免疫组化方法对不同年龄小体鲟的肝脏、肠、卵（Ⅱ-Ⅴ期卵巢）及血涂片,进行免疫组织化学定位研究。小体鲟卵巢发育到 Ⅳ期前,卵黄蛋白主要靠卵母细胞自身合成,这个时期内源性合成卵黄蛋白;当卵巢发育到 Ⅳ期,卵母细胞自身不合成卵黄蛋白,主要是通过肝脏合成卵黄蛋白原,通过血液循环运送到卵巢,被卵母细胞吸收后,裂解为卵黄蛋白,这个时期外源性合成卵黄蛋白。     

家蝇卵巢在体外培育中摄取卵黄蛋白

本文报道家蝇Musca domestica卵巢在体外培养条件下,摄取异硫氰萤光素标记的家蝇卵黄蛋白的特点。用Grace's培养液标记蛋白浓度为2mg/m1,在27℃条件下培养2小时,卵巢摄取量依赖于培养液中卵黄蛋白浓度和温度,摄取高峰在羽化后48小时,正值卵母细胞发育阶段进入6-8时期。培养液中加入JHIII,能促进摄取,JHIIl的浓度和摄取量无明显相关性。乌本苷、牛血清蛋白和叠氮钠显著抑制卵巢的摄取活动。     

不同发育时期哲罗鱼卵黄的超微结构

应用透射电镜观察了不同发育时期哲罗鱼(Hucho taimen)卵黄的超微结构.根据哲罗鱼卵黄物质在卵母细胞中的加工合成、积累以及卵母细胞中参与卵黄颗粒形成的细胞器的变化,可将该鱼卵黄发生分为4个特征时期,即卵黄发生前期、卵黄泡期、卵黄积累期和卵黄积累完成期.卵黄发生前期是指卵母细胞发育过程中的卵黄物质开始积累前的时期,此时期核仁不断分裂,出现线粒体云和早期的滤泡细胞层、基层和鞘细胞层;卵黄泡期特点主要是细胞器不断变化产生卵黄泡和皮层泡;卵黄积累期的滤泡膜由内向外依次为放射带、颗粒细胞层、基层和鞘细胞层,此时外源性卵黄前体物质不断经过血液汇集于鞘细胞层,后经微胞饮作用穿过胶原纤维组成的基层,经过多泡体作用转运至颗粒细胞内,在细胞内经过加工和修饰形成小的卵黄蛋白颗粒,卵黄蛋白颗粒经微胞饮穿过放射带进入卵母细胞边缘形成的空泡中,不断积累形成卵黄球;进入卵黄积累完成期,卵黄球体积变大,向细胞中心聚集,填满大部分卵母细胞,卵黄积累完毕.     

七星瓢虫的卵黄发生:植入雄虫的卵巢中卵黄原蛋白的合成

为了证实七星瓢虫的卵巢能合成卵黄原蛋白, 并查明雄虫体内是否具有卵黄发生所必需的激素环境, 我们将刚羽化雌虫的一侧卵巢或数个卵巢管植入雄虫体内.移植的卵巢或卵巢管在雄虫体内能够发育, 其卵母细胞能沉积卵黄, 一部分可达成熟.体外培养证明移植的卵巢可合成卵黄原蛋白, 但受体雄虫的脂肪体不合成卵黄原蛋白, 而且其血淋巴中也不存在这种蛋白.用保幼激素类似物ZR-512处理受体雄虫, 可促进移植卵巢的发育, 但不能诱导其脂肪体合成卵黄原蛋白.此结果表明, 象大多数昆虫一样, 七星瓢虫的卵黄发生的性二型现象表现在激素的靶组织——脂肪体, 而不是激素本身.     

Isolation and characterization of highly purified mosquito oostatic hormone

Oostatic hormone, the hormone that inhibits vitellogenesis in mosquitoes, was purified 7,000-fold with a recovery of 70% from the ovaries of the mosquito Aedes aegypti. The purification procedure included heat treatment and chromatography on ion exchange and gel filtration columns. The hormone is a small peptidelike molecule of molecular weight 2,200 at pH 4.5, which aggregates into larger molecular species of trimer and octamer at pH 7.0 as determined by gel filtration. The hormone is positively charged at pH 7.8 and has a low Rf at pH 9.4 on disc gel electrophoresis. Injection of purified oostatic hormone (9 ng) into female mosquitoes inhibited yolk deposition and vitellogenin synthesis. Activity of the oostatic hormone in the mosquito ovary increased rapidly following blood feeding and reached a maximum after 48 h. Oostatic hormone of A. aegypti injected into autogenous Aedes taeniorhynchus inhibited egg development. Repeated injections of dilute oostatic hormone at 24 h intervals partially arrested egg development, resulting in 60% reduction in the number of eggs laid. This hormone does not block release of egg development neurosecretory hormone (EDNH) from the mosquito brain but rather appears to act on the ovary.     

Hepatopancreas is the likely organ of vitellogenin synthesis in the freshwater prawn, Macrobrachium rosenbergii

The objective of the present study was to investigate the source of vitellogenin in the freshwater prawn, Macrobrachium rosenbergii. Ovarian development of M. rosenbergii was classified into five stages (stage I-V). Vitellin/vitellogenin was detected in the ovary and the hepatopancreas in different stages by native-PAGE and Western blotting. Two and three subunits of vitellin were observed in the ovary at the early- (I-II), mid- and late- (III-V) stages, respectively. The subunit of vitellogenin was not detected in the hepatopancreas at different stages of prawns. Hepatopancreas had positive immunocytological staining (against vitellin antibody) in different ovarian stages of prawn. Only vitellogenic oocyte but not previtellogenic oocytes and follicle cells had a positive immunocytological staining. Hepatopancreas could synthesize radiolabeled immunoreactive proteins after incubation with radiolabeled glycine on the basis of immunoprecipitation (against vitellin antiserum). Therefore, it is concluded that hepatopancreas is the most likely organ to synthesize vitellogenin in the freshwater prawn, M. rosenbergii.     

Oostatic hormone inhibits biosynthesis of midgut proteolytic enzymes and egg development in mosquitoes

Injection of partially purified oostatic hormone (0.7 μg) into female Aedes aegypti inhibited egg development, proteolytic enzyme activity, and blood digestion in the midgut, whereas control injections of saline or insulin chain A (0.7 μg) did not affect these processes. Oostatic hormone given by enema, on the other hand, did not inhibit proteolytic enzyme activity, indicating that the hormone acts outside the midgut. A single injection of oostatic hormone (0.7 μg) caused a 1.7–1.5-fold reduction in activity of trypsinlike enzymes during blood digestion, with a 10-h delay in peak activity. Using [1,3-³H]diisopropylfluorophosphate (DFP) in the presence of 8 mM tosylamide-2-phenylethyl chloromethyl ketone, the synthesis of trypsinlike derivatives was followed in the midgut of female A. aegypti. A 4-fold reduction in [1,3-³H]diisopropylphosphoryl-trypsinlike derivatives was noted after oostatic hormone treatment. Several isozymes that are normally synthesized were absent in the presence of DFP, as assessed by polyacrylamide gel electrophoresis. Injection of oostatic hormone into decapitated and ovariectomized females that did not synthesize ecdysteroids inhibited trypsinlike enzyme synthesis and blood digestion in the midgut, indicating that oostatic hormone inhibits the midgut cells and not the ovary or the brain's endocrine system. Comparison between oostatic hormone and soybean trypsin inhibitor indicated that the former inhibited trypsin synthesis whereas the latter inhibited trypsin activity. A. aegypti oostatic hormone is not species specific and injections of the hormone into Culex quinquefasciatus, Culex nigripalpus, and Anopheles albimanus caused inhibition of egg development, blood digestion, and synthesis of trypsinlike enzymes. A direct relation between oostatic hormone synthesis and the regulation of trypsinlike activity in the midgut is proposed.     

Role of juvenile hormone in the synthesis and sequestration of vitellogenins in the red cotton stainer, Dysdercus koenigii (Heteroptera: Pyrrhocoridae)

Investigations were carried out to determine the role of juvenile hormone (JH) and 20-hydroxy ecdysone in the synthesis and uptake of vitellogenins, which were earlier identified, purified and characterised, in Dysdercus koenigii. The concentration(s) of vitellogenin(s) in fat body, haemolymph and that of vitellin(s) in ovary were significantly lower after chemical allatectomy at eclosion. In addition, at 70 h after emergence, chemical allatectomy reduced ovarian vitellin concentration, but vitellogenin levels remained normal in the fat body and haemolymph. The haemolymph vitellogenins were not incorporated into oocytes in such insects. Administration of JH-III at 20 h after allatectomy restored vitellogenin levels in the fat body and haemolymph, but the ovary failed to incorporate the available vitellogenins from haemolymph in such insects. However, when JH-III was administered twice, one at 20 h and then at 70 h after allatectomy, vitellogenin concentrations in fat body and haemolymph and also vitellin concentrations in ovary approached control levels. It is suggested that JH has two separate roles, one in vitellogenin synthesis and the other in uptake. 20-hydroxy ecdysone had no apparent role in either vitellogenin synthesis or uptake in D. koenigii.     

Gonad maturation,morphological and physiological changes during the first reproductive cycle of the crayfish Cherax quadricarinatus female

Summary

Concurrent morphological, anatomical and physiological changes took place during the first reproductive cycle in the Australian red-claw crayfish Cherax quadricarinatus, which prepared the female for spawning and holding of the newly deposited eggs. The endopod became longer and wider than the exopod and developed a mixture of plumose and long thin simple (ovigerous) setae. Small oocytes (0.24±0.05 mm) were present in the immature ovary. The growing ovary contained two distinct oocyte populations: one consisted of small (0.55±0.07 mm), barely growing oocytes, while the other consisted of large oocytes, which increased in size continuously (0.73 to 2.55 mm) until egg laying took place. A gradual change in the relative abundance of ovarian polypeptides occurred until the late vitellogenic stage (large oocytes < 1.8 mm). Three predominant female-specific, SDS-PAGE separated, polypeptides were observed (103, 78 and 73 kDa) that may represent vitellin subunits. The most abundant carotenoid in the ovary was astaxanthin, while β-carotene was present at a lower concentration. The strong correlation between the increasing diameter of the oocyte and the concentration of astaxanthin in the ovary and in the hemolymph suggested an association of astaxanthin with vitellin and vitellogenin.     

A correlation between juvenile hormone deficiency and vitellogenic oocyte degeneration inDrosophila melanogaster

Summary It is known from previous work that juvenile hormone (JH) is required to initiate vitellogenin uptake into maturing oocytes ofDrosophila melanogaster, but additional requirements for this hormone during oocyte maturation have not been fully understood. To determine if early vitellogenic oocytes (stages 8 and 9) require JH for continued development, these oocytes were transplanted toDrosophila female and male hosts which were rendered deficient in JH by three methods. Implanted stage 9 and usually stage 8 oocytes were found to degenerate in JH-deficient hosts unless ZR-515, a JH analogue, was applied to the host shortly after implantation.These results were confirmed during in situ ovary development. JH deficiency was produced in gravid females, and ovaries examined at subsequent time intervals were found to be deficient in stage 8–10 oocytes as early as 6 h after treatment. Degenerating oocytes corresponding to these stages were commonly found. ZR-515 prevented oocyte degeneration during at least the first 8 h and continued to support stage 8–10 oocyte development 24 h after application to these females. The results suggest that JH is required not only for initiation but also for continuation of vitellogenin uptake and oocyte development.     

Vitellogenin synthesis in the ovary of scallop,Patinopecten yessoensis: Control by estradiol-17 beta and the central nervous system

Elucidation of a profile of scallop vitellin formation associated with oogenesis and its endocrine control, and identification of a vitellogenin synthesizing site were immunologically undertaken by using anti-scallop Vn serum. Vn content increased during ovarian growth and accounted for more than 80% of the water soluble protein of the ovary at the mature stage. In vivo injection of estradiol-17 beta (E(2)) resulted in an increase in Vn content in the ovary. In vitro accumulation of Vn in the ovarian tissue was promoted with E2 and a vitellogenesis promoting factor (VPF) from cerebral plus pedal ganglion which was heat stable, less than MW 10,000 and trypsin/chymotrypsin resistant. Estrogen receptor (ER)-like immunoreactivity was found in the growing oocyte and the auxiliary cell in close contact with growing oocytes, in which Vn immunoreactivity was also found. It is suggested that the vitellogenin synthesis occurred inside the ovary, especially in the auxiliary cell, and is controlled by E2 and VPF via ER.