LHRH-A缓释剂促进雄性赤点石斑鱼性类固醇激素分泌和精巢发育与排精的研究

研究了促黄体生成素释放激素类似物(LHRHA)不同处理方式对雄性赤点石斑鱼性类固醇激素分泌和精子生成及精子排放的影响。对照组在0、7d二次注射生理盐水(50μg·kg-1BW),注射组在0、7d二次注射LHRHA(50μg·kg-1BW),埋植组在0d埋植LHRHA缓释剂(100μg·kg-1BW)。结果表明,通过埋植LHRHA缓释剂,血清睾酮(T)和11-酮基睾酮(11-KT)水平快速增加,显示血清睾酮和11-酮基睾酮水平与赤点石斑鱼精子生成和精子排放有密切关系。组织学观察表明,在21d,对照组和注射组鱼精巢充满了精子细胞、精母细胞和精原细胞,而在同一时间,LHRHA埋植组鱼精巢含有大量精子,表明精子排放仍在进行。在40d期间采集精液总量为对照组<注射组<埋植组,差异显著。当注射组在处理14d后精液量逐渐回落到对照组水平时,埋植组仍然维持较高精液量水平。各组采集的精子的活力与采集的精液量呈正相关,而精子密度没有显著差异。数据显示,LHRHA缓释剂能显著增加赤点石斑鱼的精液量及延长精子排放时间而不改变精子的质量。     

Binding characteristics and regulation of the 17,20β,21-trihydroxy-4-pregnen-3-one (20β-S) receptor on testicular and sperm plasma membranes of spotted seatrout (Cynoscion nebulosus)

The binding characteristics of 17,20,21-trihydroxy-4-pregnen-3-one (20-S) to plasma membranes prepared from the testes and sperm of spotted seatrout (Cynoscion nebulosus) were investigated using a filtration method to retain the bound 20-S. A single class of high affinity (K_d = 17.9 nM), low capacity (B_max = 0.072 nM g^-1 testes) binding sites was identified by saturation and Scatchard analyses on testicular membranes of spermiating spotted seatrout. A corresponding receptor (K_d = 22.17 nM, B_max = 0.00261 nM ml^-1 milt) was also detected in spermatozoan membrane preparations. The rates of 20-S association and dissociation were rapid, both had T_halfs of less than 1 min. Competition studies indicated that the receptor was highly specific for 20-S. 17,20-dihydroxy-4-pregnen-3-one, which had the highest affinity of the other steroids tested, had a relative binding affinity (RBA) of 14.3%. Progesterone, 11-deoxycortisol and testosterone competed with an order of magnitude less affinity (RBA's of 7.4, 1.8 and 1.1%, respectively). Estradiol displayed low affinity for the receptor (RBA = 0.4%) and cortisol did not cause any displacement at 1000-fold excess concentration. Specific 20-S receptor binding was detected in plasma membranes from testes of both spermiating and non-spermiating seatrout and on spermatozoa. Prolonged incubation of testicular fragments from a spermiating fish with gonadotropin (15 IU ml^-1 human chorionic gonadotropin) or forskolin (10 µM) caused a 2–3 fold increase in membrane receptor binding. Previous studies have shown that gonadotropin-induced upregulation of the 20-S plasma membrane receptor in seatrout ovaries is required for the oocytes to become responsive to 20-S and undergo final maturation. The existence of a 20-S membrane receptor on sperm and its upregulation in the testes by gonadotropin raises the possibility that final maturation of spermatozoa in male seatrout may be regulated by a similar mechanism.     

不同温度下雌激素对施氏鲟早期性腺发育及血清性激素的影响

在不同养殖温度下注射外源雌激素后对孵化后90日龄的施氏鲟性未分化的仔鱼进行培育,研究早期性腺发育过程中组织结构和血清性激素含量的变化,以了解温度和外源雌激素对施氏鲟幼鱼性分化和性别比率的影响,探讨施氏鲟的早期性别控制方法和机制。在不同试验温度下,注射外源雌激素-17β-estradiol(E2)30 d后,使施氏鲟幼鱼的卵巢分化提前,但未能改变施氏鲟雌雄1∶1的性别比率。注射24 h后各温度组血浆T含量下降到0 ng.mL-1,显著低于对照组0.46 ng.mL-1的水平(P<0.05)。注射30 d后,血浆T含量有所回升,但以高温组21、24和27℃组的血浆T含量开始回升,27℃组回升幅度最大,与处理前水平无显著差异(P>0.05)。注射后24 h,各温度组血浆E2含量显著升高(P<0.05),7 d后开始缓慢降低,15 d后降到最低值后开始回升,30 d时血浆E2含量与对照组差异不显著(P>0.05)。与注射后血清T含量一样,以温度高组回升快,最快组为21℃组。结果表明,注射17β-E2不能改变施氏鲟的性别比率,但对施氏鲟血浆T和E2含量具有一定影响作用。     

Differentiation and development of Leydig cell, and induction of spermatogenesis during testicular differentiation in the Japanese eel, Anguilla japonica

The initial appearance and the development of Leydig cells (LCs), the sites of steroid hormone production in the testis, were investigated ultrastructurally during testicular differentiation in the Japanese eel, Anguilla japonica. In addition, the effects of a single injection of human chorionic gonadotropin (HCG; 5 IU g body weight^-1) on histological changes of the testes and serum 11-ketotestosterone (11-KT) were examined at various stages (15–18, 20–23, 26–29, 32–35, 38–41 and 46–50 cm body length (BL)) of testicular differentiation. Testicular differentiation was morphologically characterized by the development of loose connective tissue on the medial side in animals 18–29 cm in BL. Ultrastructurally, LCs were first identified in the loose connective tissue of the testis of the 23 cm fish. In the testes of fish over 32 cm, clusters of LCs were distributed throughout the interstitial region accompanying the increase in number of spermatogonia. In fish larger than 32 cm, spermatogenesis was induced by administration of HCG; serum 11-KT levels were also raised. On the other hand, there was no effect on spermatogenesis or serum 11-KT levels in fish less than 29 cm, or in the controls. These result suggests that morphological differentiation of LCs occurs in testis of the 23 cm eel, and subsequently, the testes of eels of BL more than 32 cm acquire the capability to produce steroid hormones.     

一个茄次碱苷的结构表征

首次从太白米大鳞茎中分离得到茄次碱-3-O-α-L-吡喃鼠李糖-(1→2)-[β-D-吡喃葡萄糖-(1→4)]-β-D-吡喃葡萄糖苷。通过对该化合物的IR,FABMS,SIMS,1H-NMR,13C-NMR,DEPT,HMQC,HMBC,1H-1HCOSY和NOSEY谱的综合解析确定了其结构,并对有关文献中的错误进行了更正。     

Effect of cortisol on the metabolism of 17-hydroxyprogesterone by Arctic charr and rainbow trout embryos

The effect of cortisol on the in vitro metabolism of [³H]17-hydroxyprogesterone ([³H]17OHP) was studied during embryonic development of Arctic charr (Salvelinus alpinus) and rainbow trout (Oncorhynchus mykiss). In the absence of cortisol, rainbow trout embryos metabolized [³H]17OHP largely to androstenedione (A₄) and androstenetrione (11-KA) with a minor conversion to 17,20ß-dihydroxy-4-pregnen-3-one (17,20P). In the presence of cortisol, this biosynthesis was inhibited. On the other hand, cortisol had no apparent inhibitory effect on the nature of metabolism of [³H]17OHP by Arctic charr embryos. In these embryos [³H]17OHP was metabolized mainly to 17,20P with a minor conversion to A₄ and without the formation of 11-KA that was seen in rainbow trout.When incubated in the presence of [³H]cortisol both Arctic charr and rainbow trout embryos produced 11ß-hydroxyandrostenedione (11ß-OHA) as the major metabolite, with a minor conversion to an unknown steroid. The catabolism of the cortisol by salmonid embryos may reflect the ability of the embryo to inactivate or detoxify cortisol to protect itself from the adverse effects of this biologically potent steroid hormone The study indicates the existence of species-specific differences in the nature of metabolism of [³H]17OHP and the inhibitory effect of cortisol on this metabolism.     

Evidence that 17α,20β,21-trihydroxy-4-pregnen-3-one is a maturation-inducing steroid in spotted seatrout

Ovarian steroidogenesis during final oocyte maturation (FOM) in the spotted seatrout (Cynoscion nebulosus) was investigated by incubating ovarian fragments with tritiated pregnenolone, followed by chromatographic separation of the radioactive products. The major tritiated steroid produced during FOM comigrated with 17α,20β,21-trihydroxy-4-pregnen-3-one (20β-dihydro-11-deoxycortisol, 20β-S) on HPLC and TLC. Only minor amounts of radioactive material coeluted with 17α,20β-dihydroxy-4-pregnen-3-one (17α,20β-P), 11-deoxycorticosterone (DOC), estradiol-17β and testosterone standards in the HPLC system. Additional chromatography by TLC confirmed the presence of radioactive estradiol-17β and testosterone but not 17α,20β-P and DOC. All the ovarian steroids producedin vitro during FOM were assayed for their ability to induce germinal vesicle breakdown (GVBD) of spotted seatrout oocytes. Twenty grams of ovarian tissue were incubated with human chorionic gonadotropin and exogenous pregnenolone. The steroidal products were purified by HPLC and TLC. Most of the maturation-inducing activity was confined to steroidal material which comigrated in these systems with 20β-S. This material was active at a concentration of 1 ng steroid/ml medium in the GVBD assay. Smaller amounts of material which coeluted with 11-deoxycortisol, DOC, 17α,20β-P and several minor unidentified fractions induced GVBD at concentrations of 10 ng steroid(s)/ml. The structure-activity relationships of authentic steroids in inducing GVBD of spotted seatrout oocytes was investigated. Hydroxylation at the 17α, 20β or 21 positions increased potency to induce GVBD. Steroids with multiple hydroxyl groups at the 17α and 20β positions (17α, 20β-P) and at the 17α, 20β, and 21 positions (20β-S) had maximum biological activity in the GVBD bioassay. The results suggest that 20β-S is a major maturation-inducing steroid in spotted seatrout.     

Immunolocalization of sex steroid receptors in the epididymis and ductus deferens of immature and mature Japanese Quail, Coturnix Japonica

The goal of this study was to determine whether in the Japanese quail the male genital tract contains receptors for progesterone, androgen and estrogen (PR, AR and ER, respectively), which have significant roles in reproductive functions, and whether their localization changes during sexual maturation. The epididymis and ductus deferens (middle and ampulla regions) of immature (approximately 30-day-old) and mature male Japanese quail were collected and frozen sections of them were immunostained for PR, AR and ER. The immunoreaction products for AR and PR were found in the nuclei of epithelial cells in the efferent ductules, epididymal duct, and the middle and ampulla regions of the ductus deferens of mature and immature birds. In the mature birds, the epithelial cells of the efferent ductules, epididymal duct, and the middle and ampulla regions of the ductus deferens were positive for ER, although some of the cells in the ductus deferens were negative. The epithelial cells of the ductules in the epididymis stained positive for ER, but the immunoreactions were negligible in the ductus deferens of immature birds. These results suggest that the epididymis and ductus deferens in quail possesses PR, AR and ER receptors. Each receptor is expressed before sexual maturation, although enhancement of ER expression may occur during maturation.     

Vergleichende Untersuchungen zur Steuerung der Ovarfunktion beim Rind mit PMSG und FSH

Inhalt: Nach Steuerung der Ovarfunktion beim Rind mit PMSG/PGF_2α und FSH/PGF_2α war 6.5 h nach der PGF_2α-Applikation eine deutliche Abhängigkeit zwischen den Estradiol-17β (E₂)- bzw. Testosteron (T)-Konzentrationen in der Follikelflüssigkeit und der Gröβe der Ovarialfollikel zu erkennen. In präovulatorischen Follikeln wurden unabhängig von der gewählten Hormonbehandlung teilweise signifikant höhere (P < 0,05) mittlere E₂- und LH-Konzentrationen als in nonovulatorischen Follikeln festgestellt. Follikel mit intakten oder degenerierten Oozyten wiesen keine signifikanten Unterschiede im Steroidgehalt auf. Der Reifungsprozeß der Cumulus-Oozyten-Komplexe bzw. Oozyten war mit charakteristischen Verschiebungen in den intrafollikulären Hormonspiegeln verbunden. Auch die veränderten Verhältnisse der Steroidhormonkonzentrationen in der Follikelflüssigkeit bestätigen die Rolle der Steroide bei der Oozytenreifung.     

Oocyte development and serum profiles of vitellogenin and steroid hormone levels in captive female Pacific herring Clupea pallasii during their first maturational cycle

ABSTRACT:   The present study investigates the relationship between oocyte development and serum steroid hormone levels in captive Pacific herring, Clupea pallasii , during the first reproductive cycle. The process of oocyte development in Pacific herring belongs to the group-synchronous type. Maturity of the ovary was divided into six periods based on histological observation (i.e. immature (April to September), onset of vitellogenesis (August to October), progress of vitellogenesis (October to December), completion of vitellogenesis (December to March), maturation and spawning (March to April) and spent (late April)). The pattern of seasonal change in the gonadosomatic index (GSI) well reflected the ovarian maturity. Serum vitellogenin levels showed good correlation with change in GSI, which increased from September to a peak (4.2 ± 0.3 mg/mL) in March. Serum estradiol-17β (E2) levels elevated from September and reached a peak (15.8 ± 4.2 ng/mL) in December, and remained comparatively high until March, suggesting that the active vitellogenin synthesis during vitellogenesis is controlled by the high E2 level. 17,20β-Dihydroxy-4-pregnen-3-one showed a single sharp peak (2.4 ± 0.28 ng/mL) in early April of the second year, suggesting it was a maturation-inducing steroid in this species.