Muscle carotenoid concentrations in sexually maturing and immature Arctic charr, Salvelinus alpinus (L.)

Muscle carotenoid concentrations were measured in immature and maturing 2+ Arctic charr, Salvelinus alpinus (L.), sampled from April until ovulation started in November. In immature fish, muscle carotenoid concentration increased steadily and had reached about 10 mg kg^?1 by the end of the experiment. In mature fish of both sexes, pigment concentrations decreased during the autumn and muscle carotenoid concentrations were significantly lower than in the immature fish during the latter stages of the trial. No significant difference in muscle pigmentation was observed between sexes. Carotenoid concentrations were lowest in the cranial region of the fillet, but decreased in all parts of the fillet when the charr became mature. In females, the decrease seemed to be most marked in the caudal region. At the termination of the experiment, fillets accounted for about one-half of the carotenoid content in mature fish of both sexes. High concentrations of pigment were recorded in the skin and fins of mature males. In mature female charr, pigment was almost equally distributed among skin, fins and ovaries     

Expression of dmrt1 and sox9 during gonadal development in the Siberian sturgeon (Acipenser baerii)

The sex differentiation period of the Siberian sturgeon was investigated through expression profiling of two testicular markers (dmrt1 and sox9). At the molecular level, a clear sexual dimorphism of dmrt1 and sox9 was observed in 3-year-old fish with immature gonads, in which males showed higher expression of these genes. Among 16-month-old sturgeons cultured in Uruguay, gonad morphology analyses showed one group of fish with undifferentiated gonads and a second group which had started their histological differentiation into ovaries or testes. dmrt1 showed a significantly higher expression in testes of recently differentiated fish, but this was not the case for sox9. In undifferentiated fish, we observed two clearly different groups in terms of expression: one group of fish over-expressing male markers (dmrt1, sox9) and another group of fish showing very low expression of these genes. This suggests that fish undergoing male differentiation can be identified by their profiles of gene expression before they undergo morphological differentiation.     

Transitions of urine flow rate and kidney-somatic index in mature masu salmon

ABSTRACT:   The present study aimed to investigate functional changes of the kidney in mature masu salmon. The urine flow rate (UFR), kidney-somatic index (KSI), and epithelial cell height of second proximal convoluted segment of renal tubules were examined in mature and immature masu salmon Oncorhynchus masou . Two methods of calculating UFR were made as either the units per body weight, or body weight without gonads. The former calculation using body weight may be used for comparison between data obtained from immature fish, and the latter method was suitable when comparing mature sexes and assessing the transition from immature to finally mature stages. In both calculations of UFR, KSI, and epithelial cell height of the second proximal segment, the values for mature fish were significantly higher than for immature fish. These results first suggest that functional transition of the kidney occurs in maturing fish.     

Sex organ extracts and artificial hormonal compounds as sex pheromones to attract broodfish and to induce spawning of Chinese black sleeper (Bostrichthys sinensis Lacépède)

Ovarian, testicular and seminal vesicle extracts of mature Chinese black sleeper (Bostrichthys sinensis Lacepede), synthetic steroids 17α‐progesterone (17α‐P), 17α,20β‐dihydroxy‐4‐pregnen‐3one (17α,20β‐P) and prostaglandins PGE₂ and PGF_2α were used in an attempt to attract broodfish and to induce spawning. Furthermore, 17α‐P, PGE₂ and PGF_2α levels were determined in extracts, and PGE₂ and PGF_2α levels were determined in holding and spawning waters. The results showed that ovarian extracts attracted more males than females to enter artificial nests, while testicular and seminal vesicle extracts attracted more females than males. The highest percentage of spawned nests were found when 17α,20β‐P and PGE₂ were administered to the nests, while the largest egg numbers and the mean highest fertilization rate were observed when PGE₂ was added. In ovarian extract, 17α‐P level was higher than levels in testicular or seminal vesicle extracts. The PGE₂ levels were significantly (P<0.05) higher than PGF_2α in the ovarian, testicular and seminal vesicle extracts. The PGE₂ and PGF_2α levels in spawning waters were higher than those in holding waters. The PGE₂ levels were greatly significantly (P<0.01) higher than PGF_2α in spawning and in holding waters. The results suggest that the sex organs of B. sinensis contain 17α‐P, 17α, 20β‐P, PGE₂ and PGF_2α and that 17α‐P, 17α,20β‐P and PGE₂ may act as sex pheromones in this species, attracting both male and female conspesifics to spawning sites and inducing spawning. Among the compounds tested, PGE₂ was the most effective sex pheromone for induction of spawning in B. sinensis.     

中华鳖SOX9基因在胚胎期性腺中的表达模式及在性逆转中的表达变化

SOX9基因在脊椎动物性别决定和性腺分化中扮演重要的调控作用。从mRNA和蛋白水平分析中华鳖SOX9基因在不同组织中的差异性表达、在胚胎性腺和成年睾丸中的细胞定位以及在性逆转中的表达变化,研究SOX9基因在中华鳖性别分化中的调控作用。Real-time PCR结果显示,SOX9基因在中华鳖雄性性腺中特异性表达。免疫荧光染色分析显示,SOX9蛋白在雄性18期胚胎性腺中开始表达,随着性腺的发育,SOX9蛋白定位于性腺Sertoli前体细胞细胞核中;而在雌性胚胎性腺并未见其表达。此外,在雌激素诱导的雄性向雌性性逆转胚胎中,SOX9基因显著下调,而在芳香化酶抑制剂诱导的雌性向雄性性逆转胚胎中,SOX9基因表达则显著上升。研究表明,SOX9基因为中华鳖雄性特异性基因,参与雄性性腺的发育过程,可能在中华鳖早期性别分化过程中起调控作用。     

Change in flesh quality associated with early maturation of Atlantic halibut (Hippoglossus hippoglossus)

To determine the effect of maturation on flesh quality, 20 Atlantic halibut (Hippoglossus hippoglossus) consisting of mature and immature fish of both sexes were killed on site, exsanguinated and stored on ice. After 6 days of storage, the fish was gutted and filleted before colour, texture hardness and shear force were evaluated to determine the effect of maturation. Results show that mature males excreted black mucus from the skin during ice storage, while a greyish mucus was observed from immature fish. Mature fish had approximately 2% lower slaughter yield and 6% lower fillet yield compared with immature fish. The fillets from mature males were significantly harder compared with fillets from immature fish and the muscle structure proved to be stronger as an increased fracturability was observed in mature fish. In colour, fillets from mature fish proved to have a whiter appearance (L^*) than immature fish, while no difference was seen in a^* and b^* values. We conclude that the physiological changes associated with maturation affects the end quality, and may be related to slower growth.     

中华乌塘鳢嗅觉器官的形态结构

中华乌塘鳢（Bostrichthys sinellsis Lacepede）样品体长17．0～19．8cm，性成熟鱼性腺为Ⅳ～Ⅴ期，性未成熟鱼性腺为Ⅱ期。取嗅囊切片、染色、固定。分别以扫描电镜和透射电镜拍照。结果显示，中华乌塘鳢具一对纺锤形嗅囊，由前、后鼻孔与外界相通，嗅上皮向嗅囊腔内突起形成10～16个初级嗅板，初级嗅板上有次级嗅板，可增大嗅上皮的表面积。嗅板由嗅上皮和中央髓两部分构成，中央髓主要由疏松结缔组织和毛细血管构成；嗅上皮排列于中央髓的两侧，由多层细胞组成。扫描和透射电镜观察表明：嗅上皮分为非感觉区和感觉区两部分，非感觉区位于嗅板边缘，较薄且平滑，外缘高倍放大呈指纹状或块状结构；感觉区位于中央部位，呈连续分布，细胞种类多样，表层为纤毛非感觉细胞，中上层为纤毛感受细胞和柱状细胞，中卜层为支持细胞，底层为基细胞。纤毛感受细胞为一种双极神经元，树突在上皮表面形成嗅结；轴突则穿过基膜，在固有层内集合成束，形成嗅神经纤维，终止于嗅叶。     

Prostanoids-induced dispersion in the melanophores of a carp Labeo rohita (Ham.)

Effects of three prostaglandins (i.e., prostanoids) and one of its precursors, arachidonic acid, were examined on the melanophores of the fish Labeo rohita (Ham.). PGE₁, PGE₂, PGF_2α and arachidonic acid elicit a concentration-related dispersion in the fish melanophores. In vitro analysis of melanophores was performed through incubation of the isolated fish scales in different agonists and antagonists solutions. Dispersal effect of prostanoids may be mediated directly through the typical receptors or indirectly through release of neurotransmitter substance(s) from the melanophore nerve endings. Denervation of fish melanophores rendered them insensitive to prostanoid (PGF_2α). Propranolol and verapamil completely inhibited the dispersal effects of PGF_2α; theophylline and indomethacine blocked the effects of higher concentrations of PGF_2α. During dispersing influence of PGF_2α, a free flux of Ca²⁺ ions was required and the indirectly released substance(s) from melanophore nerve endings would be the catecholamines of adrenergic and purinergic in nature.     

Changes in plasma somatolactin levels during spawning migration of chum salmon (Oncorhynchus keta)

Plasma somatolactin (SL) concentrations were examined in chum salmon in relation to gonadal maturation; immature salmon in the Bering Sea at various stages of maturation, and mature salmon during upstream migration caught at the ocean, bay and river. Plasma SL concentrations as well as plasma prolactin (PRL) and growth hormone (GH) levels in the immature fish caught in the Bering Sea were maintained essentially at similar levels. Plasma SL in mature salmon increased significantly from the fish in the ocean to the fish in the river in both sexes. Although all the fish had fully developed gonads, females completed ovulation while still in the bay, whereas final spermeation in males was achieved after entry into the river. Thus, no clear correlation was seen between plasma SL levels and final gonadal maturation. On the other hand, plasma PRL concentrations in both male and female fish were higher in the fish in the river than those in the ocean and bay, and plasma GH levels were higher in both sexes in the fish in the bay and river than those in the ocean. Plasma levels of triglycerides, glucose, free fatty acids and ionized sodium and calcium were also examined. Significant-negative correlations were seen between plasma SL and plasma ionized calcium in mature male salmon, and between plasma SL and plasma triglycerides in mature female salmon. Although our findings do not rule out the possibility of the involvement of SL in final maturation, the results indicate that SL seems to be involved at least in energy and/or calcium metabolism during the spawning migration.     

Structural and functional effects of early exposure to estradiol-17β and 17α-ethynylestradiol on the gonads of the gonochoristic teleost Dicentrarchus labrax

This study investigated the effects of estrogens on sexual differentiation in sea bass (Dicentrarchus labrax L.), a gonochoristic marine teleost that under culture conditions has a histologically sexual undifferentiated period that covers most of the first year of life, after which most individuals develop as males. Sea bass that had no noticeable histological sign of sex differentiation were fed estrogens at two doses (5 or 10 mg kg^-1 food) and for different periods ranging from 48 to 426 days post fertilization (DPF). Exposure to the synthetic estrogen 17-ethynylestradiol (EE₂) at 10 mg kg^-1 food from 60 to 260 DPF, including the sensitive period to equivalent doses of synthetic androgens previously determined for this species (126-226 DPF), significantly (p < 0.05) more than doubled the number of juvenile females to 80%, compared to the control value of 33%, and completely suppressed gonadal development in the remaining 20% of the population. This suggests that the period during which sea bass gonads exhibit high sensitivity to androgens is also very sensitive to estrogens. A comparable exposure to the natural estrogen estradiol-17 (E₂) resulted in 13% of the fish having suppressed gonadal development, but induced 57% of the fish to develop gonads with germinal tissue of both sexes, suggesting a pivotal role for E₂ during this sensitive period. Earlier exposure to EE₂ at 10 mg kg^-1 food from 48-88 DPF, significantly (p < 0.05) increased the number of females to 62% from 36% in the control group, allowing for the normal testicular development in the remaining fish. In contrast, a later chronic exposure (226-426 DPF) to E₂, at either 5 or 10 mg kg^-1 food, starting when the gonads showed no sign of sexual differentiation but past the critical sensitive period, had no effect on the resulting overall sex ratios, indicating that after this period responsiveness of the gonads to estrogens decreases as gonadal sexual differentiation progresses. However, the consequences of this apparently innocuous exposure were later manifested in adults, exemplified by a significant dose-dependent reduction in the number of mature males at 626 DPF, coinciding with the second reproductive season, the time when males normally reach sexual maturation in cultured sea bass. This suggests that chronic exposure to E₂ past the critical sensitive period may not affect the sex ratio, but could result in alterations in the male reproductive organs. This was later verified by histological analysis which revealed a significant (p < 0.05) dose-dependent reduction of the surface of the testicular lobules in the remaining males that did not mature. Together, these experiments illustrate both readily observable and subtle effects of estrogens on sex proportions, gonadal morphology and maturation rates, providing evidence that estrogen exposure can have delayed action in a teleost in a manner similar to the effects described for mammalian species. The possible existence of effects of this latter type in adult fish could be considered when evaluating the consequences of deliberate or accidental exposure to estrogens or putative estrogenic chemicals, particularly if such exposure had taken place during sex differentiation.     

The cytochrome P450 system of atlantic salmon (Salmo salar): I. Basal properties and induction of P450 1A1 in liver of immature and mature fish

The major components of the cytochrome P450 (P450) system in liver microsomes of Atlantic salmon were studied using spectrophotometric, catalytic and immunochemical techniques. In juvenile fish sampled during the winter season, high basal activities of 7-ethoxyresorufin O-deethylase (EROD) were found. The K_m for 7-ethoxyresorufin was 0.4 µM, and V_max 1.23 nmol/min/mg protein in juvenile fish. In mature fish sampled from the same group of fish in December, EROD activity was barely detectable (20–30 pmol/min/mg protein). Treatment with the P450 1A1 inducer -naphthoflavone (BNF) resulted in almost 2-fold induction of total P450, and 30–40-fold induction of EROD activity in immature fish. A similar fold increase was seen in mature fish. The differences in EROD activity between untreated and BNF-treated fish, was accompanied by similar differences in a P450 1A1 cross-reacting protein (M_r=58,000 D) in immunochemical studies using rabbit anti-cod P450 1A1 IgG. However, judging from these studies, the levels of P450 1A1-protein in mature salmon far exceeded those accounted for by the measured EROD activity in comparison to immature fish (both before and after BNF-treatment), indicating inhibiting effects of sex steroids on the measured activity. This effect was not seen on 7-ethoxycoumarin O-deethylase activity. A long-term storage experiment indicated that Atlantic salmon liver microsomes can be stored for 2 years at –80°C in 20% glycerol without losing more than 20–40% of its catalytic activity.Parts of this work were presented at the 5th International Symposium on Responses of Marine Organisms to Pollutants, April 1989 in Plymouth, United Kingdom (Larsen and Goksøyr 1989).     

Immunochemical sexing of living yellowfin tuna, Thunnus albacares (Bonnaterre), using a vitellogeninlike protein

Sexing of living yellowfin tuna, Thunnus albacares (Bonnaterre), was carried out based on immunochemical detection of vitellogenin (VTG), a female-specific protein. Tissue samples were taken from swimming fish in a sea pen by a hand-made apparatus. Recently fertilized eggs were also collected before tissue sample trials. A double immunodiffusion pattern showed that the egg homogenate of T. albacares reacted with rabbit antibody against the purified VTG of greater amberjack, Seriola dumerili (Risso). This suggests that the VTG of T albacares has an antigenicity similar to that of S, dumerili and that this antibody can be used to detect T albacares VTG. Immunodot-blotting analysis revealed that, out of the two fish tested, the tissue sample from one, but not the other, reacted with this antibody, suggesting that the former was a mature female, and the latter a male or immature female. Examination of the fish gonads revealed an ovary and a testis, respectively. Immunological and morphological results corresponded completely. Therefore, the method used in the present study is available for discriminating the sexes of T albacares.     

Mercuric chloride induced toxicity responses in the olfactory epithelium of Labeo rohita (HAMILTON): a light and electron microscopy study

Bioaccumulation of mercury and histomorphological changes in the olfactory epithelium of Labeo rohita were investigated after exposing the fish to two sublethal concentrations of HgCl₂ (66 and 132 μg/L) for 15 and 30 days. Mercury deposition increased in the tissue significantly (p < 0.05) with dose- and duration-dependent manner. Severe damage to the olfactory epithelium was evident. When fish exposed to 66 μg/L for 15 days, the histology of olfactory epithelium exhibited that mucous cell proliferation was upregulated and cell size was significantly increased from the control. Similar trends were found in 30 days exposure in both treated groups. Histology showed that mercury induced degeneration of columnar sensory cells, supporting cells and ciliated non-sensory cells and induced basal cell proliferation. Basal cell hyperplasia led to form intraepithelial proliferative lesion, thickening of epithelium, basal lamina disruption and cyst formation. Scanning electron microscopy revealed that mercury exposure at 66 μg/L caused clumping and loss of cilia, erosion in microridges on the supporting cells and proliferation of mucous cell opening. Complete degeneration of ciliated cells and cyst formation was observed in the fish when exposed to 132 μg/L HgCl₂. This result suggests that prolonged exposure to mercury might cause irreversible damage to the olfactory epithelium and impair the olfactory function of fish.     

青蛤染色体制备及核型分析

以青蛤成体鳃组织和性成熟的雌、雄青蛤性腺为材料,采用常规制片方法,分别制备青蛤二倍体染色体和单倍体染色体,统计染色体数目。结果显示,青蛤二倍体与单倍体染色体数目分别为2n=38和n=19;核型分析发现,青蛤核型组成为11m+6sm+2st,NF=76,未发现性染色体和随体染色体。本研究以青蛤成熟性腺为制备材料,成功制备单倍体染色体,与青蛤二倍体染色体互相印证,弥补了以往研究的不足,进一步丰富了对青蛤染色体数目及核型组成的认知。同时,也为青蛤染色体水平全基因组测序组装提供更加科学的基础资料。     

The progeny of sex-inverted female Atlantic salmon (Salmo salar L.)

A group of Atlantic salmon juveniles received dietary 17α-methyl-testosterone (3 mg/kg of food) for 60 days after first-feeding. Many of these fish became sexually mature as males at 1.5 years of age, while they remained in freshwater. Supposed sex-inverted females among these were identified by their failure to strip manually. Milt from the excised gonads of five of these fish was used separately to fertilise mixed ova from normal adult females. The progeny of each male, when examined 8 weeks after first-feeding, all had typically female gonads (n = 50).     

Long-term effects of intraperitoneal injection of estradiol-17β on the growth and physiology of juvenile stellate sturgeon Acipenser stellatus

Juvenile stellate sturgeon Acipenser stellatus were intraperitoneally injected with estradiol-17β (E₂; 0 and 5 mg/kg fish) to investigate the possibility of sex reversal and also determine the changes in biochemical parameters. Five-month-old fish (40.9 ± 1.1 g) were injected every 3-week interval during a 190-day trial. At the termination of the experiment, final weight and other growth parameters including weight gain and specific growth rate, hepatosomatic and viscerosomatic indices were not affected by repetitive injection of E₂. Hematological features of E₂-treated fish showed significant reductions in number of red blood cells, hemoglobin concentration, hematocrit value and mean corpuscular hemoglobin (P < 0.05), but no significant changes were observed in number of white blood cells, mean corpuscular volume and mean corpuscular hemoglobin concentration (P > 0.05). Calcium, phosphorus, glucose, triacylglycerol, cholesterol, total protein and estradiol concentrations were significantly increased in fish injected with E₂ (P < 0.001). Plasma progesterone and testosterone levels were noticeably lower in fish injected with 5 mg/kg E₂ rather than the control fish (P < 0.001). Histological observations of gonads showed that all fish injected with 5 mg/kg E₂ apparently feminized, while 66.6 % of the control group was female. These results revealed that the injection of E₂ is an effective method for feminization of stellate sturgeon without having significant inhibitory effects on growth and survival.     

性早熟中华绒螯蟹肝胰腺与性腺的酸性和碱性磷酸酶活性

采用磷酸苯二钠法在520nm波长测定了未成熟、早熟和正常成熟中华绒螯蟹肝胰腺与性腺中的酸性磷酸酶（ACP）和碱性磷酸酶（ALP）。结果表明：（1）各组蟹肝胰腺ACP和ALP活性都显著高于性腺;（2）早熟蟹肝胰腺中ACP活性都显著高于未成熟蟹而低于正常成熟蟹,在各发育阶段没有性别差异,但是,早熟雌蟹和正常成熟雌蟹卵巢中ACP活性都较早熟雄蟹和正常雄蟹精巢低,早熟与正常成熟个体没有差异;（3）早熟雌蟹肝胰腺ALP活性显著高于未成熟和正常成熟雌蟹,而与三组雄蟹处于同一水平,但是,早熟和正常成熟雌雄蟹的性腺中ALP活性没有显著差异。由此得出结论：（1）性腺发育所需营养物质的动员或合成主要与肝胰腺ACP有关;（2）性早熟的发生与肝胰腺中ACP和ALP活性的显著升高有关。     

Effects of individual and binary mixtures of estrogens on male goldfish (Carassius auratus)

Adverse effects of five typical environmental estrogens, namely estrone (E₁), 17β-estradiol (E₂), 4-n-octylphenol (4-n-OP), 4-n-nonylphenol (4-n-NP) and bisphenol A (BPA) on adult male goldfish (Carassius auratus) were investigated both individually and in binary mixtures, using serum vitellogenin (VTG) induction and gonadosomatic index (GSI) as the endpoints. Doses of individual and binary mixtures of estrogens were chosen at broad ranges. Five individual estrogens induced common dose-dependent increases of serum VTG in the experimental fish when injection doses of the estrogen series were comparatively low. The levels of VTG induction in fish descended after peaked at a certain dose of the individual estrogen. Significant GSI decreases were observed in fish treated by all dose series of E₁ and E₂, and comparatively high doses of 4-n-OP, 4-n-NP and BPA when compared with that of solvent control (SC). Effects of binary mixtures of the five typical estrogens on VTG induction in male goldfish were in additive manner at low-effect doses, but divergences occurred at high dose levels, with the predicted effects by additive manner exceeding those were observed. All of GSI of fish treated by the binary mixtures were about or lower than 10^?3 %. Serious atrophy of gonads was observed in all the mixture treatment groups when compared with that of SC. These findings highlight the potential reproductive risk of fish resulted from existing mixtures of hormones in the aquatic environment, and they have important implications for environmental estrogen hazard assessment.