雄烯二酮和甲基睾酮诱导雌性日本鳗鲡性发育的反馈调节作用

采用间隔１５ｄ多次埋植雄激素雄烯二酮（４－ａｎｄｒｏｓｔｅｎｅ－３,１７－ｄｉｎｅ,ＡＤＳＤ）或甲基睾酮（１７α－ｍｅｔｈｙｌ－ｔｅｓｔｏｓ－ｔｅｒｏｎｅ,ＭＴ）诱导雌性日本鳗鲡（Ａｎｇｕｉｌｌａ ｊａｐｏｎｉｃａ）性腺发育成熟;并着重研究埋植雄激素后,雌鳗血清中促性腺激素（ＧｔＨ）及脑和垂体中哺乳类促性腺激素释放激素（ｍＧｎＲＨ）含量的动态变化。一次或多次埋植ＡＤＳＤ后,１～５ｄ的血清ＧｔＨ含量上升,然后下降;并且,血清ＧｔＨ含量上升的幅度随埋植次数的增加而增加;对于ＭＴ处理组,只在埋植７次后第１５天测得血清ＧｔＨ水平显著高于对照组,但仍显著低于ＡＤＳＤ处理组。这表明埋植ＡＤＳＤ和ＭＴ可促进ＧｔＨ的分泌,但两者促进ＧｔＨ分泌的作用存在显著差异。埋植ＡＤＳＤ１ｄ后,脑和垂体中ｍＧｎＲＨ含明明显增加,第２天后和对     

雄烯二酮和甲基睾酮对日本鳗鲡血清睾酮和17_雌二醇含量的影响

探讨了间隔15d多次埋植雄烯二酮（4－androstene-3,17-dine,ADSD)或甲基睾酮（17α-methyl-testosterone,MT)诱导日本鳗鲡性腺发育过程中，血清睾酮（testosterone,T)和17β－雌二醇（17β-estradiol,17β-E2）含量的变化。在雌鳗中，埋植ADSD后血清T和17β-E2含量显著升高，随后缓慢下降；埋植后第15d的血清T含量随埋植次数的增加呈下降趋势，而血清17β-E2含量随埋植次数的增加而逐渐增加。埋植MT则使雌鳗血清中T含量显著降低，而雌鳗血清17β-E2含量在埋植MT2次后显著升高。在雄鳗中，埋植ADSD后第15天的血清T和17β－E2水平均显著升高，并随埋植次数的增加而增加；而埋植MT后第15天的血清T含量也显著升高，并随埋植次数的增加而增加，但血清17β-E2水平仅在埋植第3次后显著高于对照组。研究结果表明：埋植ADSD和MT对日本鳗鲡血清T和17β-E2的影响存在明显的性别差异。     

激素缓释制剂诱导海水养殖鱼类性腺发育成熟与生殖行为的研究进展

综述促性腺素释放激素类似物（GnRH-a)缓释制剂（SRDS），人绒毛膜促性腺激素（HCG）SRDS，类固醇激素SRDS；雄烯二酮（ADSD）和17a-甲基睾丸酮（MT），以及鲤垂体匀浆（CPE）混合SRDS诱导海水养殖鱼类性腺发育成熟和生殖行为的效果，认为采用GnRH-a SRDS处理性腺发育良好的鲈形目鱼类和鲆鲽鱼类，可以显著提高血液促性腺激素（GtH)水平，且持续时间长，诱导排卵成功，HCG乳胶SRDS可诱导日本鳗鲡（Anguila japonicus)血液GtH持续 升高，性成熟系数明显升高并且性腺发育成熟；HCG和CPE水／油／水（W／O／W）复乳SRDS对日本鳗鲡的催熟效果显著，采用ADSD和MT SRDS SRDS可成功诱导日本鳗鲡性腺成熟与排卵。     

不同激素对雌性花鳗鲡性体指数和发育过程中促性腺激素基因(GtHα,FSHβ,LHβ)表达的影响

为评估不同激素诱导花鳗鲡卵巢发育的效果并探究促性腺激素在花鳗鲡发育过程中的作用,实验比较了不同激素组合处理后花鳗鲡性体指数(gonadosomatic index,GSI)的变化,并采用实时荧光定量PCR(qRT-PCR)对处于不同发育阶段雌鱼脑垂体中促性腺激素基因GtHα(促性腺激素亚基),LHβ(促黄体生成素亚基)及FSHβ(卵泡刺激素亚基)的表达进行了定量研究.结果发现,鲤脑垂体萃取液(CPE)与人类绒毛膜促性腺激素(hCG)组合能够有效促使雌性花鳗鲡性腺发育成熟,GSI平均达22.9％,与对照组相比差异显著.单独注射雄烯二酮(ADSD)和注射ADSD、CPE、hCG混合制剂诱导效果不明显,GSI仅为1％ ～3％.荧光定量发现,经CPE与hCG处理的亲鱼LHβ和GtHα的表达量不断上调,其中发育早期GtHα的表达量极显著升高,而FSHβ的表达量呈现下调趋势.研究表明,CPE与hCG组合注射能够成功诱导花鳗鲡卵巢发育成熟,而ADSD对花鳗鲡卵巢发育无明显促进作用.LH在花鳗鲡卵巢发育整个过程中扮演重要角色;FSH在花鳗鲡性腺发育早期作用显著,可能是诱导鳗鲡性腺早期发育的关键因素.     

高效新型鱼类催产合剂介绍

<正> 近年来对鱼类生殖生理的研究已经证明:(1)鱼类脑垂体促性腺激素(GtH)的分泌活动既受到下丘脑分泌的促性腺激素释放激素(GnRH)的促进,又受到下丘脑产生的促性腺激素释放的抑制因素(GRIF)的抑制。(2)下丘脑神经分泌细胞产生的多巴胺具有GRIF的作用,它既能直接抑制脑垂体GtH细胞的分泌活动,亦能抑制GnRH促进GtH的分泌作用。(3)多巴胺的拮抗物或合成抑制剂能消除多巴胺对鱼脑垂     

介绍效果更好的鱼类催产合剂——高效鱼类催产合剂2号和3号

鱼类促性腺激素(GtH)的分泌和生殖活动已经证明为双重的神经内分泌系统所调节，即促性腺激素释放激素(GnRH)刺激GtH分泌和多巴胺作为促性腺激素释放的抑制因素(GRIF)抑制GnRH的作用以及GtH的自行释放。我们先前对养殖鱼类的研究已证明多巴胺受体的拮抗物(如pimozide，PIM)，排除剂(如reserpine，RES)和多巴胺合成抑制剂都能显著增强LRH-A诱导GtH分泌和排卵的作用。     

海藻酸钠-HCG制剂对雌性金鱼(Carassius auratus)生殖激素水平的影响

采用海藻酸钠(Na Alg)-HCG缓释制剂进行激素埋植实验。实验开始时,雌性金鱼(Carassius auratus)卵母细胞处于Ⅲ期。实验期间记录雌性金鱼性腺发育、放免(RIA)法测定血清性激素含量并且通过RT-PCR检测激素相关基因表达。结果表明,埋植Na Alg-HCG缓释激素后,雌性金鱼性成熟系数(GSI)在6-21 d显著高于对照组,血清睾酮(T)和雌二醇(E2)水平在6-30 d显著高于对照组;性腺芳香化酶基因(CYP19A基因)和雌激素受体基因(ERα基因)m RNA相对表达量分别在2-21 d、14-30 d内显著高于对照组,其性腺GH基因m RNA相对表达量变化不显著。研究结果显示,一次性埋植Na Alg-HCG缓释制剂,与雌性金鱼繁殖相关的生物学指标、激素水平、基因表达量较对照组产生明显变化,同时这种缓释制剂可以作为埋植激素的载体。     

海藻酸钠-HCG制剂对雌性金鱼(Carassius auratus)生殖激素水平的影响

采用海藻酸钠(Na Alg)-HCG缓释制剂进行激素埋植实验。实验开始时,雌性金鱼(Carassius auratus)卵母细胞处于Ⅲ期。实验期间记录雌性金鱼性腺发育、放免(RIA)法测定血清性激素含量并且通过RT-PCR检测激素相关基因表达。结果表明,埋植Na Alg-HCG缓释激素后,雌性金鱼性成熟系数(GSI)在6-21 d显著高于对照组,血清睾酮(T)和雌二醇(E2)水平在6-30 d显著高于对照组;性腺芳香化酶基因(CYP19A基因)和雌激素受体基因(ERα基因)m RNA相对表达量分别在2-21 d、14-30 d内显著高于对照组,其性腺GH基因m RNA相对表达量变化不显著。研究结果显示,一次性埋植Na Alg-HCG缓释制剂,与雌性金鱼繁殖相关的生物学指标、激素水平、基因表达量较对照组产生明显变化,同时这种缓释制剂可以作为埋植激素的载体。     

大黄鱼脑垂体组织学与免疫组织化学

用组织学和免疫组织化学方法研究不同发育时期大黄鱼（Pseudosciaena crocea）脑垂体的形态结构和各种促激素分泌细胞的分布。结果表明：（1）大黄鱼脑垂体呈“鸡心“形,由神经垂体和腺垂体组成,腺垂体可分为前外侧部（RPD）、中外侧部（PPD）和垂体中间部（PI）。（2）RPD部有3种细胞,分别为催乳激素（PRL）细胞,促肾上腺皮质激素（ACTH）细胞,促性腺激素（GtH）细胞。（3）PPD部位于腺垂体腹面,也有3种细胞,其中1种为促生长激素（GH）分泌细胞,呈嗜酸性,另2种呈嗜碱性,分别为GtH细胞与促甲状腺激素（TSH）分泌细胞。（4）垂体中间部有2种细胞,1种为促黑激素（MSH）细胞,呈嗜酸性,另1种为GtH细胞,呈嗜碱性。随性腺发育成熟,大黄鱼GtH细胞的分布从中外侧部扩展至前外侧部和中间部背面;GtH细胞胞质空泡可作为其分泌活动的标志。     

17α-甲基睾酮对赤点石斑鱼性逆转的影响

采用腹部埋植方式,用17α-甲基睾酮(17-αMT)以10mg.kg-1剂量处理2龄雌性赤点石斑鱼(每4周埋植1次,共埋植2次),检查埋植前后性腺结构、血清中性类固醇激素水平以及脑和性腺芳香化酶活性的变化。结果显示:一次埋植17-αMT即可诱导赤点石斑鱼发生不同程度的性逆转,性腺成熟系数(GSI)明显下降,性腺中卵细胞退化,精原细胞增殖,出现大量精母细胞和精子细胞,性逆转雄鱼精巢的组织结构与正常雄鱼精巢相似,部分鱼转变为功能性雄鱼,经轻微的腹部挤压可排精,精子活力与正常雄鱼相似。赤点石斑鱼埋植17-αMT后前脑、中脑的芳香化酶活性明显提高而后脑的芳香化酶活性降低,但性腺芳香化酶活性无显著变化。埋植17α-MT后第4周血清中11-酮基睾酮(11-ketotestosterone,11-KT)浓度显著升高,而睾酮(testosterone,T)和雌二醇(estradiol-17,βE2)浓度变化不显著。这些结果表明,脑部芳香化酶活性的显著升高与性逆转密切相关,17-αMT主要通过提高血清中11-KT水平诱导赤点石斑鱼发生性逆转。     

鲤脑垂体匀浆液和人绒毛膜促性腺激_省略_性腺激素及性类固醇激素含量的影响

雄鳗注射５－６次、雌鳗注射９－１０次鲤脑体匀浆液（ＣＰＥ）＋人绒毛膜促性腺激素（ＨＣＧ）能分别诱导精巢和卵巢发育成熟。在雌雄鳗鲡,注射ＣＰＥ＋ＨＣＧ可显著增加端脑、间脑、中脑和下丘脑ｍＧｎＲＨ的含量,而对后脑和延髓ｍＧｎＲＨ的影响较小;注射ＣＰＥ＋ＨＣＧ增加雄鳗后脑和延髓ｃＧｎＲＨ－Ⅱ含量,对雌鳗脑区ｃＧｎＲＨ－Ⅱ则无显著影响。雌雄鳗鲡每次注射ＣＰＥ＋ＨＣＧ后１天,血清促性腺激素（ＧｔＨ）急剧上升     

Differential distribution and response to experimental sexual maturation of two forms of brain gonadotropin-releasing hormone (GnRH) in the European eel,Anguilla anguilla

Using specific radioimmunoassays for the two GnRH molecular forms present in the European eel, Anguilla anguilla, (mGnRH and cGnRH II), we compared their distributions in the pituitary and different parts of the brain of female silver eels, as well as the modifications of their levels in experimentally matured female eels (treated with carp pituitary extract). In control eels, mGnRH levels were higher than cGnRH II levels in the pituitary, olfactory lobes and telencephalon, di- and mesencephalon, while the opposite was found in the posterior part of the brain (met- and myelencephalon). Experimental sexual maturation of the gonads significantly increased mGnRH levels in the pituitary and anterior parts of the brain; such a positive effect was not observed on the low cGnRH II levels, which were, in contrast, reduced. These data indicate that the positive feedback of gonadal hormones on GnRH, that we previously demonstrated, would specifically affect the mGnRH form. The differential distribution and control of mGnRH and cGnRH II suggest that these two forms have different physiological roles in the eel. The large increase in mGnRH during sexual maturation suggests the prime implication of this form in the neuroendocrine control of reproduction.     

Positive feedback control by the gonads on gonadotropin (GTH) and gonadoliberin (GnRH) levels in experimentally matured female silver eels,Anguilla anguilla

Treatment of sham-operated female silver eels with carp pituitary extract stimulated ovarian development and induced increases in pituitary gonadotropin (GTH) and gonadoliberin (GnRH) contents. Both effects of carp pituitary extract were abolished in ovariectomized eels, indicating the involvement of the gonads. Endogenous sexual steroids, the secretion of which was increased during sexual maturation, should be responsible for the stimulation of GTH and GnRH levels. Ovariectomy itself had no significant effect on pituitary GTH and GnRH contents, reflecting the fact that, at the silver stage, sexual steroid levels are too low to exert any significant effect on pituitary GTH and GnRH. The positive feedback control exerted by the gonads on GTH and GnRH levels during sexual maturation, in the eel as well as in some other teleosts, would produce an amplification of the pubertal stimulation of the hypothalamo-pituitary-gonadal axis.     

Studies on the mode of action of neuropeptide Y (NPY) on maturational gonadotropin (GtH) secretion from perifused rainbow trout pituitary glands

The action of neuropeptide Y (NPY) and gonadotropin releasing hormone (s-GnRH) have been compared on the release of gonadotropin (GtH) by perifused rainbow trout pituitary glands sampled from freshly ovulated female rainbow trout. We have already demonstrated that NPY stimulated the secretion of GtH in vitro. The pituitary responses to s-GnRH and NPY were similar either after repeated 10 min infusions or a one hour prolonged application. In both cases, after the first application, the pituitary did not responded to subsequent secretagogues stimulations, and appeared to be desensitized. The stimulatory action of s-GnRH was partly inhibited (60%) by LH-RH antagonist (DpGlu¹, DPhe², DTrp^3–6) LH-RH, which completely inhibited the response to NPY in perifused pituitary glands sampled from freshly ovulated females, but did not modify the response of pituitaries taken from vitellogenic animals in which NPY induced a transient inhibition of the GtH secretion. These results may indicate that the mode of action of NPY would differ between vitellogenic and matured animals. NPY also stimulated the GtH secretion from perifused pituitary dispersed cells prepared from pituitaries taken from freshly ovulated rainbow trout, indicating that NPY may act directly on the pituitary gonadotropic cells to stimulate GtH secretion.     

Effect of aromatable androgen (17‐methyltestosterone) on induced maturation of silver European eels (Anguilla Anguilla): Oocyte performance and synchronization

The reproductive performances of silver European eel in term of gonad development and egg production, employing slow‐release implants with the androgen 17‐MT (1 mg) in combination with traditional weekly injection of carp pituitary extract (CPE) was evaluated. Wild female European eels (Anguilla anguilla) underwent a standard induction protocol with CPE and were randomly divided into three groups (N‐group, no implant; Y‐group, with implant; and control, C‐group, no treatment). The results showed that 17‐MT‐treated females (Y‐group) reproduced spontaneously about 6 weeks earlier than the N‐group females with a saving of almost 40% in CPE and time of induction. Concerning artificial induction of maturation in female silver eels, our study demonstrated that they positively respond to androgen exposure also in terms of eggs productivity. Indeed, Y‐group was more productive than N‐group: in Y‐group, 11 eels ensured an eggs production that exceeded 50% of initial body weight (BW), whereas in N‐group only three eels have exceeded this value. The results suggest that 17‐MT should be considered in future protocols for the improvement of the artificial reproduction of female silver European eels.     

Gonadal regulation of gonadotropin subunit expression and pituitary Lh protein content in female hybrid striped bass

To assess the response of the GtH system to the removal of gonadal feedback, effects of bilateral gonadectomy and 17β-estradiol (E₂) replacement on gonadotropin (GtH) subunit expression, as well as LH protein levels, were monitored in the pituitary. Adult female hybrid striped bass undergoing mid-vitellogenesis were divided into one sham and two gonadectomized (gdx) groups. One gdx group received E₂ (2 mg kg^?1) via microspheric delivery systems, the other was given a vehicle injection. Groups were sampled on days 3 and 14 post-surgery (PS). Expression of all GtH subunits was significantly increased in the gdx group on both days compared to the shams, whereas E₂ replacement in gdx fish restored alpha and LHβ mRNA levels to those of the sham fish and dramatically reduced FSHβ mRNA levels. Gdx had no effect on the pituitary concentration of LH. E₂ replacement, however, reduced protein levels significantly on day 3, but not day 14. In conclusion, during mid-vitellogenesis expression of both GtHβ subunits is negatively controlled by gonadal feedback, which is also evident by restoration of gdx subunit mRNA levels to those of the shams after E₂ replacement.     

日本鳗鲡产卵后生存及其繁育

为证明日本鳗鲡(Anguilla japonica)生活史最后一步一产后鳗的命运,本研究模拟产后的日本鳗鲡继续在海水中养殖,观察其存活率及繁育情况.结果表明,产后鳗在海水中停食约18 d后,体能得到恢复,部分亲鱼开始出现摄食,1个月左右全部恢复摄食,经244 d养殖,雌、雄鳗体质量增加,存活率达94.6%.随后,给产后鳗注射外源性促性腺激素(鲤鱼脑垂体匀浆CPE和人绒毛膜促性腺激素HCG)后激发其退化的性腺(卵巢和精巢)重新发育(与当年银鳗作对照).通过性腺组织切片观察产后鳗和对照鳗性腺发育成熟的全过程及其差异,发现产后鳗起初性腺发育比当年银鳗差,但经多次注射激素后,产后鳗性腺成熟与当年银鳗同步,证明产后鳗生殖细胞对激素的敏感件高.应用17α,20β-双羟孕酮和促性腺激素释放激素类似物(GnRH-A3)使催熟的产后鳗和对照鳗均产卵和排精,并孵化出仔鱼,从而有力地证明,鳗鲡产后虽体质弱,但待体能恢复后能够继续生存和繁殖.本研究旨在探讨利用产后鳗作为今后鳗鲡人工繁殖亲鳗的可行性.     

Growth hormone secretion during longterm incubation of the pituitary of the Japanese eel,Anguilla japonica

Growth hormone (GH) secretion from organ-cultured pituitaries of the eel (Anguilla japonica) was studied during incubation in a defined medium for 2 weeks, using a homologous radioimmunoassay which does not distinguish between the two molecular forms of eel GH. The total amount of GH secreted increased gradually during the incubation period; so that the amount of GH released on day 14 was about 30 times greater than that on day 1. On day 14, the proportion of GH released relative to the total amount of GH present (the sum of GH released into the medium and residual content in the pituitary) was 96% and the amount produced on day 14 was 4 times greater than the content in the unincubated pituitary. Somatostatin (SRIF, 1.8 × 10^-7 M) inhibited the increase in GH release. On day 7, the proportion of GH released by pituitaries treated with SRIF (28%) was less than that released by the control pituitary (91%). There was no significant difference in GH release between the pituitaries incubated in isotonic medium (300 mOsm) and those in hypotonic medium (240 mOsm) for 2 weeks except for the first 3 days, when the pituitaries in hypotonic medium secreted significantly greater amounts of GH than those incubated under isotonic condition. Hypertonic medium (350 mOsm) had no effect on GH release except for significant inhibition on days 6 and 14. When secretion of the two forms of GH (GH I and II) was examined after separation by polyacrylamide gel electrophoresis followed by densitometry, slightly more GH I tended to be secreted than GH II during the culture period, although the effects of SRIF and osmolality of the media on GH I release were similar to those on GH II. It is concluded that GH secretion and production in the eel is mainly under the inhibitory control of hypothalamus, and that osmolality has a minimum influence on the GH release.     

LHRH-A、DA对施氏鲟脑垂体碎片释放GtH的影响

为了探讨在古老的软骨硬鳞鱼中促性腺激素(GtH)的双重内分泌调节作用,本实验设计用离体灌流的方法研究促黄体素释放激素类似物(LHRH-A)和多巴胺(DA)对施氏鲟脑垂体碎片分泌GtH的影响。引入10、100和1 000 nmol/L 3个浓度的LHRH-A对施氏鲟脑垂体碎片3次脉冲式刺激实验;每次间隔1 h,持续5 min,研究不同剂量LHRH-A对鲟鱼脑垂体释放GtH的作用;用200 nmol/L DA对施氏鲟脑垂体碎片持续2 h灌流后引入5 min的1 000nmol/L LHRH-A刺激实验,研究DA如何抑制鲟鱼脑垂体释放GtH。每5 min收集一管灌流液,用放射免疫测定法(RIA)检测灌流液中GtH的含量。结果显示,低剂量LHRH-A随着刺激引入脑垂体释放GtH出现波浪式的增加,中、高剂量出现释放延后现象。LHRH-A在10nmol/L到1 000 nmol/L范围内对刺激脑垂体释放GtH没有剂量依存关系。DA对施氏鲟脑垂体碎片GtH的分泌没有显著影响,但是可以抑制LHRH-A引起的GtH分泌,即DA不能抑制施氏鲟GtH的基础分泌,而只能抑制LHRH-A诱导的GtH分泌。研究结果证明,在高等硬骨鱼类中存在的双重神经内分泌调节在古老的鲟鱼中也存在。