LOCALIZATION OF A PUTATIVE EPIBOLY-DETERMINING FACTOR IN OOCYTES OF THE GOLDFISH (CARASSIUS AURATUS)

It is unknown whether cytoplasmic determinants in goldfish eggs are asymmetrically localized beforematuration. In this study fully grown goldfish oocytes with infact germinal vesicles (GVs) were ligated withbaby hair loops along desired planes into two parts, mature in vitro, and inseminated. Embryos developedfrom the animal halves with GV of oocytes ligated along the equator un unable to undergoepiboly, while those developed from the GV-containing parts of oocytes ligated along the animal-vegetal axiswere able to. Also, embryos developed from the animal halves obtained from fertilized eggs could undergo epiboly. From there results, it was postulated that the goldfish's premature oocyte possessed a putativefactor in the vegetal hemisphere which was responsible for epiboly of the embryonic cells.     

Localization of a putative epiboly-determining factor in oocytes of the goldfish (Carassius auratus)

It is unknown whether cytoplasmic determinants in goldfish eggs are asymetrically localized before maturation. In this study fully grown goldfish oocytes with intact germinal vesiles (GVs) were ligated with baby hair loops along desired planes into two parts, matured in vitro, and inseminated. Embryos developed from the animal halves with GV of oocytes ligated along the equator were unable to undergo epiboly, while those developed from the GV-containing parts of oocytes ligated along the animal-vegetal axis were able to. Also, embryos developed from the animal halves obtained from fertilized eggs could undergo epiboly. From these results, it was postulated that the goldfish's premature oocyte possessed a putative factor in the vegetal hemisphere which was responsible for epiboly of the embryonic cells. This work was supported by the NSFC and the “Climb Plan B” (PD-B6-1-1) of the Chinese Commission of Science and Technology. Contribution No. 2915 from the Institute of Oceanology, Chinese Academy of Sciences.     

Effects of inhibitors on 1-methyladenine induced maturation of starfish oocytes

1-methladenine (1-MA) induces starfish oocytes maturation via surface reaction followed by the appearance of a cytoplasmic maturation factor which in turn induces germinal vesicle breakdown (GVBD) to resume meiosis. Cellular mechanisms involved in GVBD were investigated by microinjection of metabolic inhibitors. Colchicine (Co) inhibited maturation, cytochalasin-B (CB) delayed GVBD and actinomycin-D-(Act-D) and puromycin (Pu) had no effect. It appears that the microtubule and the microfilament systems are associated with the nuclear membrane dissolution during the process of oocyte maturation of starfish.     

The insemination of goldfish (Carassium auratus) oocyte maturedin vitro

Full maturation of goldfish oocyte was inducedin vitro by 17 α-hydroxy-20β-dihydroprogesterone. The oocyte maturation involves GV migration to the periphery of the oocyte and germinal vesicle breakdown (GVBD). In the experiment, incubation duration for GVBD varied in different broods of oocytes. Generally, if the duration for GVBD was shorter than 6 h, oocytes would have a better chance to survive after maturation and insemination. The maturation of nucleus (GV) and cytoplasm are not synchronous. Cytoplasm maturation occurs several hs after GVBD. Oocytes inseminated 8–9 h after GVBD have the highest fertilizing and hatching rate. Fertilized ova maturedin vitro can develop to sexually mature adults capable of reproduction. Contribution No 1731 from the Institute of Oceanology, Academia Sinica.     

Role of animal pole protuberance and microtubules during meiosis in sea cucumber Apostichopus japonicus oocytes

Fully grown oocytes of Apostichopus japonicus have a cytoplasmic protuberance where the oocyte attaches to the follicle. The protuberance and the oolamina located on the opposite side of the oocyte indicate the animal-vegetal axis. Two pre-meiotic centrosomes are anchored to the protuberance by microtubules between centrosomes and protuberance. After meiosis reinitiation induced by DTT solution, the germinal vesicle (GV) migrates towards the protuberance. The GV breaks down after it migrates to the oocyte membrane on the protuberance side. The protuberance then contracts back into the oocyte and the first polar body extrudes from the site of the former protuberance. The second polar body forms beneath the first. Thus the oocyte protuberance indicates the presumptive animal pole well before maturation of the oocyte.     

The reaction time of organ-forming substance in goldfish EGG and its relationship with mesodermal formation

Fertilized goldfish eggs were dechorionated with a pair of forceps and were cut off along or a little above the equator into animal and vegetative parts at desired stages with a glass needle or ligated into two connected fragments before cleavage with baby hair loop. Some of the ligated eggs were detached by further fastening soon after ligation, and some released later at different stages (2-cell, 16-cell, 128-cell, 512-cell, mid-blastula) to let the organ-forming substance (OFS) enter the blastoderm. The cholinesterase (ChE) in the resulting embryos was assayed. The results are as follows. 1. All the 142 embryos developed from the animal hemispheres cut off or ligated off before cleavage gave rise to hyperblastula in which no ChE activity was observed. 2. All 50 embryos obtained from animal halves isolated at the 8-cell stage produced ChE. 3. Embryos developed from the eggs released before the 512-cell stage formed ChE, but the later the releasing of the hair knots, the smaller the number of ChE-producing embryos. 4. After the 512-cell stage (excluding this stage), neither ChE nor tissue differentiation occurred in the embryos developed from the unfastened eggs though their OFS flow was set free. Since ChE is thought to be a muscle-specific enzyme in the early developmental stage, it is concluded that the OFS in goldfish egg appears to be indispensable for the establishment of the mesoderm. Contribution No. 1507 from the Institute of Oceanology, Academia Sinica     

Comparison of lipids in organs of the starfish Asterias amurensis associated with different treatments

Lipids were extracted from organs of the starfish Asterias amurensis associated with different treatments(raw-control,boiling and heating),and then analyzed for lipid content,lipid oxidation index,lipid classes and fatty acid composition.Results showed that boiling softened the hard starfish shells,thus facilitating the collection of starfish organs.As compared with raw organs,the boiled organs had lower water content and higher lipid content,possibly due to the loss of water-holding capacity caused by protein denaturation.Both boiling and heating increased the peroxide value(PV),thiobarbituric acid(TBA) value and carbon value(CV) of lipids.Despite slight increases in the content of complex lipids,associated lipid composition had no substantial variations upon boiling and heating.For simple lipids,the content of 1,2-diglyceride decreased in boiled and heated organs,with free fatty acids observed on thin layer chromatography(TLC).However,neither boiling nor heating significantly changed the fatty acid compositions of simple or complex lipids in starfish organs,suggesting that these two treatments had no significant effects on complex lipids in starfish organs.Together,our results indicated that boiling of starfish soon after capture facilitated the handling and extraction of useful complex lipids consisting of abundant glucosylceramide and eicosapentaenoic acid(EPA)-bounded phospholipids.     

Growth,reproduction & population structure of the freshwater crabSinopotamon yangtsekiense bott, 1967, from Zhejiang,China

Monthly investigations were mae on the population of Chinese freshwater crab,Sinopotamon yangtsekiense Bott, 1967 from April, 1984 to March, 1985. The data on 4413 specimens show that the growth was affected mainly by temperature. During the April to November growth period, the crabs' major development occurred from June through October. One year was required for a fine white oocyte to develop into a mature egg. The reproduction period was June–October. Females bearing eggs were taken from June–August, and crabs with young were found from July–October. The females reproduced once a year but could for more than one year. The number of eggs carried by a female varied greatly according to the size of the crab, ranging from 30 to 100 eggs. New-born crabs become mature after 1–2 years. The sex ratio was approximately 1∶1 in the overall population. However, the larger crabs are predominantly male. The age distribution ofS. yangtsekinese was estimated from size frequency histograms. There were more adult crabs (over 70%) from June to October and more immature crabs (over 50%) from November to May.     

Sperm nuclear expansion and meiotic maturation in normal and gynogenetic eggs of the scallop, <Emphasis Type="Italic">Chlamys farreri</Emphasis>

Sperm nuclear expansion, meiosis and the association of the male and female pronuclei leading to the four-cell stage in normal Chlamysfarreri eggs were observed under a fluorescence microscope. The effects of ultraviolet （UV） irradiation on the fertilizing sperm were also examined. Both normal and UV-irradiated sperm nuclei enlarged at three distinct phases （phase A, metaphase I; phase B, polar body formation; and phase C, female pronuclear development and expansion） that were temporally correlated with meiotic process of the maternal chromosomes. Sperm nuclei underwent a rapid, initial enlargement during phase A, but condensed slightly during phase B, then re-enlarged during phase C. The effects of UV irradiation were not apparent during transformation of the sperm nucleus into a male pronucleus, and there was not any apparent effect on meiotic maturation and development of the female pronucleus. However, the rate of expansion of the UV-irradiated sperm nuclei and the size of male pronuclei were reduced apparently. Unlike the female pronucleus, the male pronucleus derived from sperm genome inactivated by UV irradiation did not form chromosomes, but became a dense chromatin body （DCB）. At mitotic anaphase, DCB did not participate in the karyokinesis of the first cleavage as evidenced by chromosomal nondisjunction, demonstrating the effectiveness of using UV irradiation to induce gynogenetic scallop embryos.     

GROWTH, REPRODUCTION & POPULATION STRUCTURE OF THE FRESHWATER CRAB SINOPOTAMON YANG TSEKIENSE BOTT, 1967, FROM ZHEJIANG, CHINA

Monthly investigatiom were made on the population of Chinese freshwater crab, Sinopotamon yangteskiense Bott, 1967 from April, 1984 to Match, 1985. The data on 4413 specimens show that the growth was affected mainly by temperature. During the April to November growth period, the crabs' major development occured from June through October. One year was required for a fine white oocyte to develop into a mature egg. The reproduction period was June-October. Females bearing eggs were taken from June-August, and crabs with young were found from July-October. The females reproduced once a year but could for more than one year. The number of eggs carried by a female varied greatly according to the size of the crab, ranging from 30 to 100 eggs. New-bom crabs become mature after 1-2 yeats. The sex ratio was approximately 1:1 in the overall population. However, the larger crabs are predominantly male. The age distribution of S. yangtsekinese was estimated from size frequency histograms. There were more adult crabs     

Development of inter-family nuclear transplant embryos by transplanting the nuclei from the loach blastulae into the non-enucleated zebrafish eggs

The developmental fate of the pronuclei in recombined embryos obtained by transplanting the donor nuclei into the non-enucleated eggs remains controversial in the case of fish.In the present study,the nuclei from the loach blastulae were transplanted into non-enucleated zebrafish eggs,the resulting 9 inter-family nuclear transplant embryos developed to larval stages,Although the development timing of the nuclear transplants resembled that of zebrafish,chromosome examination revealed that most of the recombined embryos were diploids with karyotype characteristic of loach,which was also proved by RAPD analysis.Moreover,3 out of the 9 larval fish formed barb rudiments specific to loach.It was therefore concluded that the nuclear transplant larval fish were inter-family nucleo-cytoplasmic hybrids;and that only the donor nuclei were involved in the development of the nuclear transplant embryos,while the pronuclei in the non-enucleated eggs were likely automatically eliminated during the development.     

The role of phosphatidylinositol signaling pathway in regulating serotonin-induced oocyte maturation in Mercenaria mercenaria

Serotonin (5-HT) has been found to stimulate meiotic maturation of oocytes in many molluscs. During maturation, several signaling pathways are involved, especially the phosphatidylinositol and cAMP pathways. In order to examine the possible role of the phosphatidylinositol signaling pathway in regulating oocyte maturation in Mercenaria mercenaria, the effects of the activator/inhibitor of phospholipase (PLC) and protein kinase C (PKC) on serotonin-induced maturation were studied. Results show that high-concentrations of neomycin (inhibitor of PLC) blocked oocyte maturation, while 9, 10-dimethyl-1, 2-benzanthracene (DMBA, activator of PLC) promoted oocyte maturation in the presence of serotonin. It was also found that in the presence of serotonin, phorbol 12-myristate 13-acetate (PMA, activator of PKC) inhibited oocyte maturation, while sphingosine (inhibitor of PKC) stimulated oocyte maturation. These results indicate that serotonin-induced oocyte maturation requires the activation of the phosphatidylinositol pathway. Decrease of PLC inhibited serotonin-induced oocyte maturation, whereas a decrease of PKC stimulated the maturation. Thus, our study indicates that serotonin promotes maturation of M. mercenaria oocytes through PLC stimulated increase in calcium ion concentration via inositol-1, 4, 5-trisphosphate (IP3) but not PKC.     

Chromosome behavior of heat shock induced triploid in Fenneropenaeus chinensis

Triploidy was induced in Chinese shrimpFenneropenaeus chinensis by 30±0.5°C heat shock treatment (initiated at 20 min after fertilization) for 10 min to inhibit the release of PB2 at 18.0°C. The highest triploid rate obtained was 84.5% in nauplius stage. The effect of heat shock treatment on meiosis and cleavage of eggs was investigated in this work aimed to establish efficient procedures for triploid induction and to gain understanding of the mechanism of triploid production. Three pronuclei that could be observed in the treated eggs under fluorescence microscope developed into triploid embryos. Some abnormal chromosome behavior was observed in heat shocked eggs. Contribution No. 4407 from the Institute of Oceanology, Chinese Academy of Sciences. This work was supported by Project under Major State Basic Development Program (G1999012009) and Knowledge Innovation Program of the Chinese Academy of Sciences (KZCX 2-211)     

CYTOLOGICAL COMPARISON OF GAMETOGENESIS OF SCALLOPS, ARGOPECTEN IRRADIANS AND CHLAMYS FARRERI

Histological characteristics of gametogenesis of two kinds of scallops, gonochoric Chinese scallop, Chlamys farreri and hermaphroditic bay scallop, Argopecten irradians were investigated in this study. Spermatogenesis in C. farreri has different developmental stages: spermatogonia, primary spermatocyte, second spermatocyte, spermatid and spermatozoon. A large number of same developmental stage spermatic cells converge at a definite area of the testis. Premeiotic, previtellogenic and vitellogenic oocytes can be found during oogenesis in C. farreri, where oocyte distribution is obviously irregular. The A. irradians gonad consists of two different parts in one individual: one part functions as testis, the other as ovary. Between these two parts is a special appearance area, where a large number of spermatic cells are bound with two layers of acellular substance with many oocytes in it.     

Chromosome Behavior of Heat Shock Induced Triploid in Fenneropenaeus Chinensis

INTRODUCTIONTriploidanimalsareusefulbecausetheyaresterile ,characterizedbypositivegrowthinthere productiveseason ,usefulfortesting ,forintroductionofnon nativespecies ,andforprotectionofde velopedstrains.Theinductionoftriploidyhadbeenreportedinmanyaquaculturespecies .Triploidshavebeensuccessfullyinducedinshrimpusingtemperatureshockorchemicalshock (CB ,6 DMAP)in 4species :Sicyoniaingentis (Xiangetal.,1 991 ) ,Fenneropenaeuschinensis (Xiangetal.,1 992 ;Daietal.,1 993;Baoetal.,1 993;Li…     

Reproductive biology of bream A bramis brama(L.) in the lower reaches of the Irtysh River,China

The reproductive biology of bream A bramis brama(L.) was studied from 546 fish collected from the lower reaches of the Irtysh River in Xinjiang, north-west China, from March to November 2013. The overall sex ratio(M/F) was 1.06:1, and was not significantly diff erent from the theoretical 1:1 ratio( P 0.5). However, there was a dominance of males during the pre-spawning season(1.33 in March and 1.56 in April, P 0.5), while females were dominant during the peak spawning season(0.88 in June, P 0.5). The monthly variation in gonadosomatic index(GSI) and proportion of gonads at each macroscopic maturity stage, indicated that A. brama spawned once a year with peak spawning occurring from late May to June. The unimodal distribution of oocyte diameter each month indicated that A. brama is a single spawner, with a high degree of spawning synchronicity. The standard lengths(SL 50) and ages( A 50) at first maturity for males and females, were 178 and 204 mm, and 5.6 and 6.8 years, respectively. The mean absolute fecundity(AF) was 77 311 eggs per fish, and mean relative fecundity(RF) was 162 eggs per gram of body weight(BW). The AF of A. brama increased linearly with increasing of gonad weight(GW), eviscerated weight(EW) and standard length(SL), but was not significantly correlated with age. As, A. brama in the lower reaches of the Irtysh River reaches reproductive maturity relatively late in their life span, is mature for a short period and spawns in aggregations, this fish is vulnerable to overexploitation.     

以江蓠为栖息环境养殖的双齿围沙蚕性腺发育

研究以江蓠为养殖基质的双齿围沙蚕的配子发生及性腺发育。结果表明:1)根据双齿围沙蚕卵细胞形态特征、内部结构、卵径大小,卵母细胞的发生可划分为第Ⅰ时相(卵原细胞)、第Ⅱ时相(小生长期初级卵母细胞)、第Ⅲ时相(大生长期初级卵母细胞,卵黄形成前期、卵黄形成中期、卵黄形成后期)和第Ⅳ时相(成熟卵母细胞,胶膜形成前期、胶膜形成后期)等4时相;根据各时相卵子的数量变化及体腔中其他结构的形态变化,将雌性性腺分为Ⅰ期卵巢、Ⅱ卵巢、Ⅲ卵巢、Ⅳ卵巢和Ⅴ期卵巢等5个阶段。2)根据双齿围沙蚕体腔内精母细胞的大小、形态和所处的位置,将双齿围沙蚕的精子发生划分为精原细胞、初级精母细胞、次级精母细胞、精细胞和精子;根据各时相精子的形态和数量变化以及体腔中其他组织器官的形态变化,将双齿围沙蚕的雄性性腺划分为Ⅰ期精巢、Ⅱ期精巢、Ⅲ期精巢、Ⅳ期精巢和V期精巢等5个阶段。3)Ⅰ-Ⅳ期的性腺主要包含对应时相的生殖细胞,同时,也存在有少量其他时相的生殖细胞,生殖细胞的发育速度并不完全一致。     

The in vitro effects of steroids,human chorionic gonadotropin and cyanoketone on germinal vesicle breakdown of striped mullet (Mugil cephalus L.) oocytes

The in vitro effects of steroids, human chorionic gonadotropin (HCG) and cyanoketone on germinal vesicle breakdown (GVBD) of striped mullet (Mugil cephalus L.) oocytes were investigated. All concentrations of HCG (5,10,50 I.U./ml), progesterone and pregnenolone at the highest concentrations(lug/ml) were moderately effective in inducing GVBD, whereas 17β-estrodiol, cortisol, testosterone, 11β-hydroxyandrostenedione and 11-ketotestosterone did not stimulate GVBD. 17α, 20βdihydroxy-4-pregnen-3-one (17α, 20βdiOHprog) and deoxycorticosterone (DOC) were the most potent steroids in stimulating final oocyte maturation. The results indicate that C21 hydroxylated steroids are potent inducers of final maturation in mullet. Further, co-incubations with 17β-estradiol, cortisol and testosterone did not alter the maturation-inducing effects of HCG or 17α,20βdiOHprog. Cyanoketone, a blocker of 3βHSD activity, was only partially effective in blocking GVBD induced by HCG. This suggests that Δ5 (pregnenolone derived) and Δ4 steroids may be involved in final oocyte maturation in this species.     

不同条件延缓未受精卵激活对鱼类细胞电融合的影响

泥鳅未受精卵先在合成卵巢液、郝氏液中分别浸泡15、20、30min,然后再受精孵化,统计发育情况,结果表明,合成卵巢液可有效延缓未受精卵遇水后被激活的时间,郝氏液对延缓未受精卵被激活也有一定作用,但效果比合成卵巢液差。用合成卵巢液配制不同浓度胰酶处理受体卵进行细胞电融合实验,胰酶质量浓度为5 mg/mL时,融合卵发育至囊胚期的比例比对照组(郝氏液)高2%,统计分析表明该差异是显著的(P<0.05);但发育至原肠期及孵出期的比例则与对照组无显著差异。胰酶质量浓度为2和1 mg/mL时,融合卵发育到各阶段的比例均低于对照组,统计分析表明两者的差异是显著的(P<0.05)。研究表明,溶液种类、胰酶浓度等对电融合卵的发育有明显的影响,但延缓受体卵的激活时间并不能提高电融合卵的出苗率。用郝氏液配制的2 mg/mL的胰酶溶液处理受体卵,可获得较高比例的融合鱼。