Untersuchungen zur Dotterbildung in den Oocyten von Xiphophorus helleri (Heckel, 1848) (Teleostei,Poeciliidae)

Zusammenfassung Die Verteilung von Nucleinsäuren, Polysacchariden, Proteinen und Fetten in den verschiedenen Entwicklungsstadien der Oocyten von Xiphophorus helleri wurde histochemisch untersucht. Junge Oocyten enthalten viel RNS und Proteine, aber wenig Fett. Während der Vitellogenese steigt der Gehalt an Fetten, Glycoproteide werden eingebaut. Die dazu nötigen Stoffe werden aus dem mütterlichen Körper in niedermolekularer, löslicher Form über das Follikelepithel zur Oocyte transportiert. Diese histochemischen Befunde werden durch Ergebnisse der Elektrophorese gestützt. In elektronenmikroskopischen Untersuchungen wurden den histochemischen Fakten weitere morphologische Grundlagen zugeordnet. Während der Prävitellogenese tritt ein Dotterkern auf, der sich vor Beginn der Dotterbildung auflöst. Coated vesicles nehmen Material aus dem perioocytären Interzellularraum auf. Zellstrukturen der Oocyte beteiligen sich am Dotteraufbau. Besonders auffallend sind die Veränderungen an den Mitochondrien. In der Zusammensetzung des Dotters und in der Beteiligung der Organellen gibt es Unterschiede zu Lebistes. Injektionen von Myofer runden die Ergebnisse ab. Vom maternalen zum embryonalen Gewebe erfolgt ein Transport von Nährstoffen, nicht nur Gasaustausch. Die Definition der Ovoviviparie muß daher erweitert werden.

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Studies on yolk formation in oocytes of Xiphophorus helleri

Summary The distribution of nucleic acids, polysaccharides, proteins, and fats in different developmental stages of oocytes of Xiphophorus helleri was studied using histochemical methods. Young oocytes had high RNA and protein contents. During vitellogenesis, the fat content increased and glycoproteins were deposited. These substances were transported from the maternal body into the oocyte in a soluble, small molecular form. The histochemical results were supported by our studies using electrophoretic and electron microscopic methods. During previtellogenesis, a yolk nucleus appeared. Coated vesicles took up material from the intercellular space around the oocyte. Differences between Xiphophorus and Lebistes were observed in the composition of yolk and in the prevalent types of ultrastructural organelles present. In Xiphophorus, labelling by Myofer (intraperitoneal or intramuscular injections) verified the transport of nutritive material from the maternal body into oocytes (Stage III and IV) and embryonic tissues. This suggests that in Xiphophorus the definition of ovoviviparity has to be modified.

     

Genome organization in Xiphophorus (Poeciliidae; Telostei)

Summary Xiphophorus represents a valuable model for studying genomic contributions to neoplasia. For analyzing these contributions at the molecular level, basic information about the genome organization is a prerequisite. This study presents data on the organization and complexity of the genomes of three species of Xiphophorus, maculatus, variatus and helleri, representative of the problem. Their diploid nuclei, as measured in the erythrocyte, contain 1.19 pg, 1.23 pg, and 1.27 pg DNA, these values representing approximately 50% of that of birds, 20% of that of mammals. The melting curves of native, high molecular weight DNA are homogeneous, the T_m was determined for maculatus as 85.0° C (corresponding to a mean GC-content of 38.3%) for variatus as 86.0° C (GC=40.7%), for helleri as 85.0° C (GC=39.3%). Reassociation of sheared denatured DNA indicated approximately 90% single copy sequences, the remaining 10% are predominantly multiple copy sequences. The complexity of single copy DNA was determined from reassociation kinetics for maculatus as 3.97×10⁸ base pairs, for variatus as 4.31×10⁸ base pairs, and for helleri as 4.49×10⁸ base pairs. The DNA of the three species upon isopycnic density gradient centrifugation in the presence of the fluorescence dye Hoechst 33258 shows in addition to the main band, two heavy (GC-rich) satellites, denoted in the order of increasing density, components I and II. Analytical centrifugation reveals for the main band DNA a buoyant density of 1.6980 gcm^-3 (GC=38.7%), for component I 1.7080 gcm^-3 (GC=48.9%), for component II 1.7150 gcm^-3 (GC=56.1%). Each of the components comprises approximately 0.38% of the total DNA. Complete digestion of components I and II with restriction enzymes EcoRI and BamHI yields a complex banding pattern upon agarose gel-electrophoresis. A 2.4 kb fragment of component I and a 5.3 kb fragment of component II of helleri, cloned and amplified in the pBR322/E. coli RR1 system, hybridize efficiently to purified nuclei of liver. Furthermore, restriction fragments of component II DNA, transferred to nitrocellulose by Southern-blotting, hybridize with 18S and 28S ribosomal DNA.     

Immunoreactive gonadotropin-releasing hormone (GnRH) in the brain and pituitary of adult and juvenile swordtails (Xiphophorus helleri,Teleostei, Poeciliidae)

Different molecular variants of gonadotropin-releasing hormone (GnRH) were localized in the brain and pituitary of Xiphophorus helleri, from neonates up to mature animals of both sexes. Nine GnRH antisera to salmon (s), mammalian (m), chicken I (c-I), and chicken II (c-II) GnRH were utilized. In the first week after birth GnRH immunoreactivity (IR) emerges with pale staining of the nucleus olfactoretinalis (NOR) in the ventral forebrain. The intensity of IR in the NOR increases during the next weeks and an IR tract of nerve fibers appears, protruding from the NOR in dorsocaudal direction. Adult animals exhibit additional GnRH-positive structures. Some perikarya of the nucleus preopticus periventricularis (NPP) are IR and positive fibers extend from the NPP toward the pituitary. In the pituitary IR fibers are also detectable. A distinctive structure in adult animals is an IR cord of neurons (CN) at the bottom of the forebrain which extends from the NPP to the olfactory nerve. A comparison of antisera against different GnRH species indicates that sGnRH is present in the NOR, whereas a different form of GnRH is present in the NPP, CN, and pituitary. The early onset of GnRH IR in the NOR and the widespread distribution of positive fibers from that nucleus into other brain regions suggest neuromodulatory functions of sGnRH from the NOR. The NPP possibly plays a major role in direct stimulation of pituitary gonadotropes via a different type of GnRH. © 1996 Wiley-Liss, Inc.     

Demonstration of Strength and Concealment of Weakness in Escalating Fights of Male Swordtails (Xiphophorus helleri)

In the green swordtail (Xiphophorus helleri) confrontations between strange males regularly escalate to high levels of mutual Bites and Fin Grips, even between males differing greatly in size. The original expectation of early game theory models that the behaviours of the ultimate winners and losers are indistinguishable until shortly before the end of the fight could not be confirmed. A significant characteristic of loser behaviour is that the Biting rates are higher before and after escalation. Conversely, Fin Grips are more frequent in the ultimate winners than the losers. However, such behavioural differences are very poor predictors of the outcome of the fight from the viewpoint of a single fighting individual during the contest. Correct forecasts did not exceed 67%. The escalating fights of swordtails are considered as trials of strength in which the stronger male tries to demonstrate strength by the most costly behaviour pattern available, namely Fin Grips, and the weaker male conceals weakness quite successfully by countering with the same tactic and suppressing signs of weakness such as Avoidance behaviour.     

17α-甲基睾酮对剑尾鱼的影响

目的研究17α-甲基睾酮暴露对产后雌性剑尾鱼(Xiphophorus helleri)性逆转组织形态学变化的影响,探讨剑尾鱼第二性征变化作为水环境风险评价(ERA)的有效生物学标记的可能性。方法使用浸浴法以10μg/L 17α-甲基睾酮为剑尾鱼染毒持续7周,对实验鱼的体形、腹鳍、臀鳍、尾鳍及性腺等组织器官的变化进行观察;同时对幼鱼在实验室养殖条件下的性别分化进行统计。结果 17α-甲基睾酮对产后雌鱼有明显的影响,受诱导后出现性逆转,逐渐发育形成雄性第二性征:体形变纤细;腹鳍第1鳍条变短、第2和第3鳍条延长,臀鳍第3、4、5鳍条末端钩状化且第3鳍条变粗壮,尾鳍上下缘出现增生;体内与臀鳍相连的骨骼合并生长;受诱导的雌鱼性腺呈现退化趋势并伴有卵细胞坏死现象,生殖能力受到负面影响。结论剑尾鱼臀鳍和尾鳍变化可作为水环境雄激素物质污染监测的有效生物学标记。     

Dominance hierarchies of male green swordtails (Xiphophorus helleri) in nature

Swordtail males formed dominance hierarchies at shallow sites in creeks or streams and invested much time in maintaining a high rank-order position (mean 11·1 chases per 5 min).     

Reproductive biology of Gambusia affinis holbrooki Baird and Girard, Xiphophorus helleri (Gunther) and X. maculatus (Heckel) (Pisces; Poeciliidae) in Queensland, Australia

The reproductive biology of three introduced Poeciliidae, Gambusia affinis holbrooki Baird and Girard, Xiphophorus helleri (Günther) and X. maculatus (Heckel) was studied over a 14-month period in Brisbane, Queensland. Photoperiod and water temperature determined the length of the breeding season in G. affinis , which produced up to nine broods between August and March, with the peak of reproductive activity in October. Fecundity was linearly related to body weight in G. affinis (mean fecundity = 22.78 ± 1.02, range = 3–108 offspring per brood).
Xiphophorus helleri bred from August to May, producing eight to nine broods, with peak reproductive activity from October to December. Fecundity was curvilinearly related to body weight (mean fecundity = 60.15 ± 3.8, range = 7–183 offspring per brood. Xiphophorus maculatus bred between September and March, producing five to six broods during the study period. Fecundity was linearly related to standard length (s.l.) (mean fecundity = 27.27 ± 2.4, range = 3–92 offspring per brood). Although the reproductive strategy of the three species differs, all have the capacity to build up large populations in south-eastern Queensland. Their present distributions and relative abundance are discussed.     

Uptake of foreign ferritin in platy Xiphophorus maculatus (Poeciliidae: Teleostei)

The ability and capacity of various tissues in platy Xiphophorus maculatus L. to take up horse-spleen ferritin injected into the blood stream are described. Ferritin was injected intraperioneally, and the cellular uptake was demonstrated as Prussian blue precipitations in tissues treated with acid ferrocyanide solutions. Ferritin was detected within the heart endocardial cells and macrophages in the trunk kidney and spleen, 1/4 h after the injection, i.e. foreign ferritin was taken up very rapidly by these cells. When the time elapsed between the ferritin injection and sacrifice exceeded 6 h, these cells, and also macrophages in the gill and intestine, were almost completely filled with ferritin. At these stages, however, the amounts of Prussian blue precipitations per volume unit of the tissue were much larger in the heart than in the other organs studied in the present work, i.e. the endocardial tissue seems to play an important role in the clearance of the blood circulation in this species. We suggest that this tissue in platy is specialized to endocytose waste and foreign macromolecules, including pathogenic particles, from the blood stream. The eosinophilic and neutrophilic granulocytes do not appear to take up foreign ferritin, i.e. these cells may play no endocytotic role in the clearance of foreign macromolecules in platy.     

17β-雌二醇对剑尾鱼卵黄蛋白原的诱导研究

目的研究17β-雌二醇(E2)暴露对雄性剑尾鱼(Xiphophorus helleri)卵黄蛋白原(vitellogenin,Vtg)诱导作用作为环境风险评价(ERA)的有效生物学标记的可行性。方法以E2诱导的剑尾鱼雄性个体的整体匀浆液为材料,采用Sephacryl S-300凝胶过滤层析柱和QSepharose阴离子交换柱从剑尾鱼体内提纯Vtg。结果确定了被纯化的剑尾鱼Vtg在4%~7.5%Native PAGE电泳中相对分子质量为540×103。4%~7.5%Native PAGE电泳后的凝胶分别利用苏丹黑B进行脂蛋白染色、高碘酸-Schiff试剂进行糖蛋白染色和甲基绿进行磷蛋白染色,表明剑尾鱼Vtg是一种富含糖、脂、磷的蛋白。结论表明雄性剑尾鱼卵黄蛋白原的诱导变化可作为环境风险评价(ERA)的有效生物学标记。     

Intermediary metabolism of normal and tumorous tissue of Xiphophorus (Teleostei: Poeciliidae)

• 1.1. The Xiphophorus melanoma system is one of the few well established and genetically well understood in vivo models in experimental carcinogenesis. However, data describing features of intermediary metabolism of the genetically caused melanoma or the different inducible neoplasia, as well as that of the different transformed cell lines of Xiphophorus, are still lacking. For this reason we initiated a comparative study of enolase-, pyruvate kinase-, lactate dehydrogenase- and malate dehydrogenase-activities and pyruvate and lactate levels in transformed as well as normal tissues of Xiphophorus.
• 2.2. We observed tissue specific and age dependent activities of the different enzymes and substrate levels.
• 3.3. Enzyme activities and substrate levels from all tumors analyzed differ from that of any normal tissue. They are dependent on the tumor sections analyzed, the histiotype and the etiology of the tumors.
• 4.4. Analysis of enzyme activities from different in vitro cultured fish cell lines and the human Hela cell line revealed dependency of the intermediary metabolism on oxygen supply, on the proliferative state of the cells and on the cell types.
• 5.5. We could not find a correlation between our data and the expression of the c-src gene of Xiphophorus and no genotype-dependent changes in enzyme activities were detected.

     

多氯联苯对剑尾鱼超氧化物岐化酶活性的影响

目的　研究多氯联苯 (PCBs)暴露对剑尾鱼 (Xiphophorushelleri)超氧化物歧化酶 (SOD)活性的影响 ,探讨剑尾鱼器官组织内SOD活性变化作为环境风险评价 (ERA)的有效生物学标记的可行性。方法　测定了PCBs对剑尾鱼 30d的半致死浓度 (LC50 ) ;使用浸浴法以 0、2、5 0 μg L三个PCBs浓度为剑尾鱼染毒 ,定量测定了 72h内肝、鳃及卵巢组织中的SOD的活性变化。结果　PCBs对剑尾鱼的 30dLC50 为 1 0 5 80 μg L ;PCBs对剑尾鱼肝脏和卵巢的SOD活性有明显 (P <0 0 1 )的影响。在最初染毒的 1 2h内 ,SOD活性略有上升 ,但随暴露时间延长 ,浓度增加 ,肝和卵巢的SOD活性均呈下降趋势。此外 ,结果还显示肝组织的SOD活性较高于卵巢的SOD活性 ,表明不同器官组织的SOD活性对PCBs胁迫的敏感性存在一定的差异。实验中鳃组织SOD活性在PCBs暴露后其变化不明显 (P >0 0 5 )。结论　表明剑尾鱼肝细胞SOD活性可作为环境风险评价 (ERA)的有效生物学标记     

Eavesdropping on visual cues in green swordtail (Xiphophorus helleri) fights: a case for networking

Aggressive contests probably occur in networking environments where information about fighting ability is conveyed both to an opponent and to individuals peripheral to the fight itself, the bystanders. Our primary aim was to investigate the relative influences of eavesdropping and prior social experience on the dynamics of aggressive contests in Xiphophorus helleri. A bystander's ability to witness an encounter was manipulated using clear, one-way mirror, and opaque partitions. After watching (or not watching) the initial contest, the bystander encountered either the winner or loser of the bout. Treatment comparisons of bystander-winner or bystander-loser contest dynamics indicated the presence or absence of winner, loser, or eavesdropping effects. Winner and loser effects had negligible influences on bystander contest dynamics. Eavesdropping significantly reduced the bystander's propensity to initiate aggression, escalate, and win against seen winners regardless of whether the watched bout had escalated or not. Though eavesdropping had relatively little effect on bystander-loser contest dynamics, bystanders were less prone to initiate aggression and win against losers that had escalated in the witnessed bout. Thus, bystanders appear to preferentially retain and utilize information gained about potentially dangerous opponents (winners or persistent losers). Our data lend clear support for the importance of eavesdropping in visually based aggressive signalling systems.     

剑尾鱼线粒体细胞色素b基因的序列分析

目的　克隆和测定剑尾鱼 (Xiphophorushelleri)线粒体细胞色素b基因 (cytb)的全序列。方法　提取剑尾鱼肝脏的总DNA。设计合成特异引物进行PCR扩增。扩增产物经琼脂糖电泳检测、纯化后克隆到pGEM Teasyvectorsystem中的T载体上 ,筛选转化子 ,提取质粒 ,酶切鉴定。挑取重组质粒pGEM T xhcytb 11进行序列测定。结果　获得了剑尾鱼线粒体cytb基因的全序列 ,共 114 0bp。结论　用BLAST与GenBank中的线粒体DNA序列进行比较 ,显示剑尾鱼与其他鱼类的cytb基因具有较高的同源性 ;根据剑尾鱼与其他 13种鱼的cytb基因序列同源性所建立的进化树 ,与传统的分类地位基本吻合     

Genetic Linkage Map of Fishes of the Genus Xiphophorus (Teleostei: Poeciliidae)

Analysis of genotypes of 76 polymorphic loci in more than 2600 backcross hybrid individuals derived from intra- and interspecific genetic crosses of fishes of the genus Xiphophorus (Poeciliidae) resulted in the identification of 17 multipoint linkage groups containing 55 protein-coding loci and one sex chromosome-linked pigment pattern gene. Multipoint linkage analyses identified highly probable gene orders for 10 linkage groups. The total genome length was estimated to be approximately 18 Morgans. Comparisons of the Xiphophorus linkage map with those of other fishes, amphibians and mammals suggested that fish gene maps are remarkably similar and probably retain many syntenic groups present in the ancestor of all vertebrates.     

Genetic variation and phylogeography of the swordtail fish Xiphophorus cortezi (Cyprinodontiformes, Poeciliidae)

Swordtail fish have been studied extensively in relation to diverse aspects of biology; however, little attention has been paid to the patterns of genetic variation within and among populations of swordtails. In this study, we sequenced the mtDNA control region from 65 individuals and 10 populations of Xiphophorus cortezi to investigate the genetic variation within and among populations, including tests for correlations between genetic and geographic distances and tests for species monophyly. We found low gene and nucleotide diversity within populations and high degrees of genetic differentiation among populations. Significant and positive correlations between genetic distance and both river and straight-line geographic distance indicate that genetic differentiation among X. cortezi populations can be explained, to some extent, by an isolation-by-distance model and provide evidence of stream capture. Phylogenetic analyses suggest that X. cortezi is paraphyletic relative to X. malinche, raising questions concerning the status of these taxa as separate species.     

Plasma membrane structure in spermatogenic cells of the swordtail (teleostei,Xiphophorus helleri)

The plasma membrane of spermatogenic cells of the teleost Xiphophorus helleri was examined ultrastructurally and cytochemically in order to characterize the temporal development of the membrane specializations characteristic of the mature spermatozoon. Mature sperm display a mosaic distribution of Concanavalin A and Ricinus comrnunis I binding sites; the anterior region of the head displays an intense binding that is not seen in other surface regions. This asymmetric binding is evident in early spermatids and the area of lectin binding appears associated with the plasma membrane overlying the nucleus. Transmission electron microscopy reveals that the plasma membrane over the anterior region of the head is characterized by an ordered glycocalyx and a tight adherence to the underlying nucleus. Additional membrane differentiations were revealed both in the midpiece region where a “submitochondrial net” is attached to the plasma membrane and at the base of the axoneme where the plasma membrane possesses a “collar-like” arrangement of circumferential rings. The possible functions of these differentiations, as well as their correlation to differentiations seen in sperm of other animal groups, are discussed.