Effects of rearing water quality on expression of brain corticotropin-releasing hormone and gonadal development in goldfish (Carassius auratus)

The effects of changes in water quality in a recirculating system on brain corticotropin-releasing hormone (CRH) gene expression in goldfish were examined. In the experimental group, fish were reared without water change, while in the control group, 50% of water was changed everyday. In the experimental group, total ammonia-nitrogen, nitrate and phosphate were increased over the experimental period. Brain CRH mRNA levels in the control group seemed higher than those in the experimental group, but differences were confounded by variation among individuals. There were no differences in gonadal development between both groups.     

Characterization of growth hormone binding sites in the goldfish,Carassius auratus: effects of hypophysectomy and hormone injection

A recombinant carp growth hormone (rcGH) was used to develop for a GH radioreceptor binding assay in the goldfish (Carassius auratus). Specific binding of¹²⁵I-rcGH to goldfish liver membranes was a pH, time, temperature, and membrane protein dependent process. Scatchard and LIGAND analysis indicated a single class of high affinity and low capacity binding site, with an association constant (Ka) of 1.9×10¹⁰ M^–1 and a maximum binding capacity (B_max) of 9 fmol mg^–1 protein. Liver tissue displayed the highest¹²⁵I-rcGH binding of all the tissues examined. Displacement of¹²⁵I-rcGH with various unlabeled teleost and mammalian GHs and prolactins revealed that the goldfish hepatic binding site was highly specific for teleost GH. Intraperitoneal administration of 0.1, 1.0, and 10 g rcGH g^–1 body weight to hypophysectomized goldfish resulted in a 27, 52, and 68% decrease in total binding sites, respectively. Injection of a high dose of rat prolactin (rPRL) (5 g rPRL g^–1 body weight) also resulted in a 32% decrease in total binding sites. These results suggest that endogenous GH may have a role in the regulation of its own receptors in the goldfish.     

银鲫crh基因的克隆、组织表达谱及其对摄食的影响

为研究crh (Corticotropin-releasing hormone)基因对鱼类摄食的调控作用,首次克隆了银鲫(Carassius auratus gibelio) crh基因c DNA全长序列。运用实时荧光定量PCR(RT-qPCR)技术检测crh基因m RNA在不同组织的表达及餐前、餐后和禁食对其表达的影响。结果显示,银鲫crh基因的c DNA序列全长为920 bp,其中,5’-UTR为53 bp,3’-UTR为378 bp,开放阅读框(Open reading frame,ORF)为489 bp。推导银鲫crh基因的ORF区编码蛋白由162个氨基酸组成,其中,含有11个氨基酸的保守区,24个氨基酸的信号肽,41个氨基酸的成熟肽。氨基酸序列的多重比较分析显示,银鲫crh基因与金鱼(Carassiusauratus)的同源性为99%,与鲤(Cyprinuscarpio)的同源性为96%。荧光定量PCR表达分析显示,银鲫crh基因在下丘脑中的表达量最高,其次是端脑和心脏。crh基因的表达量在餐前与餐后没有显著变化(P0.05),在禁食第5天、第7天出现极显著降低(P0.01),而在复投喂后第9天、第11天出现极显著升高(P0.01)。以上结果显示,crh基因在银鲫摄食调控方面可能扮演着重要角色。     

久效磷对金鱼精巢超显微结构和特征性酶活性的影响

用0.01、0.10、1.00 mg.L-1久效磷暴露金鱼21 d,测定了精巢乳酸脱氢酶(LDH)、酸性磷酸酶(ACP)、碱性磷酸酶(ALP)的活性,并观察了精巢显微和超微结构的变化。结果表明:久效磷暴露21 d,精巢小叶基膜断裂,Leydig氏细胞水肿,小叶间质扩大,支持细胞核膜溶解,胞质内脂滴和髓磷脂象数量增多。1.00 mg.L-1久效磷造成了生殖腺指数的显著下降,对精巢发育有一定程度的延迟作用。随着久效磷暴露浓度的升高,LDH,ACP,ALP的活性逐渐下降,0.10、1.00 mg.L-1久效磷暴露组精巢LDH,ACP的活性下降显著,而各暴露组的精巢ALP活性呈现不显著下降趋势。推测久效磷可能通过损伤精巢超显微结构和降低LDH,ACP和ALP活性,干扰精巢能量代谢,造成对雄性金鱼的生殖毒性。     

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.     

Cloning,molecular characterization and expression analysis of insulin-like growth factor binding protein-2 (IGFBP-2) cDNA in goldfish,Carassius auratus

In the present study, a full-length cDNA encoding the insulin-like growth factor binding protein-2 (IGFBP-2) was cloned from the liver of goldfish (Carassius auratus) by rapid amplification of cDNA ends technique. The goldfish IGFBP-2 cDNA sequence was 1,513 bp long and had an open reading frame of 825 bp encoding a predicted polypeptide of 274 amino acid residues. Semi-quantitative RT-PCR results revealed that goldfish IGFBP-2 mRNA was expressed in all detected tissues. In liver, central nervous system and pituitary gland, goldfish IGFBP-2 expressed at high levels, followed by anterior intestine, middle intestine and kidney. In posterior intestine, ovary, skin, fat, spleen, muscle and gill, the goldfish IGFBP-2 expression levels were very low. Fasting and refeeding experiment showed that the mRNA expression of goldfish IGFBP-2 was up-regulated significantly in liver compared to the fed group and restored rapidly to normal level after refed. However, the mRNA expressions of IGFBP-2 in hypothalamus and pituitary of goldfish were insensitive to fasting. Furthermore, the mRNA expressions of IGFBP-2 in hypothalamus, pituitary and liver were varied in periprandial changes and significantly down-regulated at 2 and 4 h after meal. These results imply that the IGFBP-2 mRNA expression may be associated with anabolic and catabolic metabolism and regulated by metabolic factors in goldfish.     

The life cycle of Chloromyxum auratum (Myxozoa) from goldfish, Carassius auratus (L.), involves an antonactinomyxon actinospore

The myxozoan parasite Chloromyxum auratum Hallett, Atkinson, Holt, Banner & Bartholomew, 2006 , was shown experimentally to have a two‐host life cycle which involved a previously undescribed antonactinomyxon actinospore stage. Myxospores obtained from gall bladders of naturally infected feral goldfish, Carassius auratus (L.), were used to infect samples of mixed species of oligochaete worms obtained from the same locality as the fish: Fern Ridge Dam, Oregon, USA. After some 110 days post‐exposure, actinospores were detected from the water above the oligochaetes. The 18S rDNA sequence of these actinospores was identical to the original myxospores. Spore release was sporadic, of low intensity and short duration, which confounded efforts to identify the host oligochaete species and infect naïve fish. This is the first life cycle that incorporates an actinospore of the collective group Antonactinomyxon, and the first life cycle demonstrated in the laboratory for a species of Chloromyxum.     

Effect of microalgal biomass concentration and temperature on ornamental goldfish (Carassius auratus) skin pigmentation

The aim of this work was to investigate the effect of different carotenoid sources/concentrations and temperature on goldfish (Carassius auratus) skin pigmentation. In the first trial (trial A), the effect of carotenoid source (natural – microalgae Chlorella vulgaris and synthetic – Carophyll Pink) and carotenoid concentration (45, 80 and 120 mg pigment kg^?1 diet) was tested. Six homogeneous duplicate groups of juvenile goldfish (7.4 g) were fed, for 5 weeks, one of the diets containing 45, 80 or 120 mg of total pigments of C. vulgaris biomass or synthetic astaxanthin per kg of diet (Cv45, Cv80, Cv120, Ax45, Ax80, Ax120), respectively. In trial B, the effect of water temperature on skin pigmentation was studied. Five homogeneous duplicate groups of 25 goldfish each (5.2 g) were fed diet Ax45 over 9 weeks, to test the following temperatures: 22, 24, 26, 28 and 30 °C. At the end of both trials, samples of skin along the dorsal fin were withdrawn for subsequent analysis of total carotenoid content, intensity of colour, red and yellow hue and visual observation. The best carotenoid concentrations were achieved with astaxanthin diets. There was a tendency to an overall improvement of colour parameters (L and b) in fish fed diets with high levels of C. vulgaris. The results indicated that the best temperature range to maximize skin pigmentation was 26–30 °C.     

Effects of short‐term starvation on the rhythmic expression of microRNAs in skeletal muscle of goldfish (Carassius auratus)

Molecular oscillators exist in peripheral tissues such as in skeletal muscles and diet is a dominant factor to affect Zeitgeber for peripheral clocks. MicroRNAs (miRNAs) play an important role in muscle cell proliferation and differentiation. However, the knowledge of starvation effects on miRNA rhythmic expression remains limited in teleost. In this study, the circadian expression pattern of miRNAs was investigated in goldfish muscle upon restricting feeding treatment. The data showed that 15 miRNAs exhibited a daily rhythmicity among the 70 miRNAs assayed in muscles of normally fed goldfish. While after 7‐day and 15‐day fasting treatment, 23 and 18 miRNAs showed circadian rhythmicity in goldfish muscles respectively. Only 4 miRNAs (miR‐23a, miR‐29, miR‐199a‐3p and miR‐455) exhibited a daily rhythmicity in all three groups of different nutrient treatments. Correlation analysis of the circadian‐miRNAs indicates different feeding stimulation could alter the circadian‐miRNA components and their expression profile in skeletal muscle upon short‐term feed deprivation. These miRNAs may play important roles in regulating circadian expression of genes involved in muscle cell differentiation and growth by nutritional alteration, thus having a potential application in fish aquaculture.     

Effect of azadirachtin on haematological and biochemical parameters of Argulus-infested goldfish Carassius auratus (Linn. 1758)

Argulosis hampers aquaculture production and alters the host physiology and growth. Azadirachtin is recognized as a potential antiparasitic agent against Argulus sp. The present study aimed to investigate the effect of different concentration of azadirachtin solution on haematological and serum biochemical parameters of Argulus-infested goldfish Carassius auratus. Ninety Argulus-infested goldfish were randomly divided into six equal groups. Fish of group 1–5 were treated with azadirachtin solution through bath of 1, 5, 10, 15 and 20 mg L^?1 as T1, T2, T3, T4 and T5, respectively, and group 6 was exposed to 2 % DMSO solution without azadirachtin and considered as negative control T0^?. Along with six treatment groups, a positive control T0⁺ of healthy goldfish free from Argulus infestation was also maintained. Parasitic mortality was evaluated after 3 days of consecutive bath treatment. After 7 days of post-treatment, the blood and serum were drawn from each of the treatment groups and haematological and serum biochemical parameters were evaluated. Total leucocyte count (TLC), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), blood glucose, total protein (TP), globulin, serum glutamate oxaloacetate transaminase (SGOT) and serum glutamate pyruvate transaminase (SGPT) were significantly (p < 0.05) high in negative control group when compared with positive control group. It could be concluded that Argulus infestation altered marked haematological and serum biochemical parameters. However, in treated groups complete elimination of Argulus was found in T4 and T5 groups. Also significant (p < 0.05) reduction in haematological and serum biochemical parameters of all the treatment groups were recorded in comparison with negative control group. In addition, T4 and T5 groups showed significantly (p < 0.05) high superoxide dismutase (SOD), catalase, total erythrocyte count (TEC) and haemoglobin (Hb). However, higher mean corpuscular haemoglobin concentration (MCHC), blood glucose and lactate dehydrogenase (LDH) levels in T5 group revealed that higher concentration of azadirachtin have notable effects on activity of vital tissues function and physiology of the host. Argulus spp. from infested goldfish could be eliminated using bath treatment with solution of azadirachtin having concentration of 15 mg L^?1 and that also shifted haematological and serum biochemical parameters towards homeostasis.     

Use of canola oil in the feed of larval and juvenile goldfish, Carassius auratus (L.)

Goldfish were used as a model for the evaluation of canola oil as a lipid source in the feeds of larval and juvenile cyprinids. Goldfish larvae were raised from hatching until 24 weeks of age on diets containing cod liver oil. canola oil or a mixture of the two oils as the lipid source. Survival, weight gain and weight-length relationship did not differ among groups of fish fed the three diets. Carcass fatty acid profiles largely reflected those of the diets except that carcasses of fish fed canola oil contained long chain (n-3) and (n-6) polyunsaturated fatty acids (PUFA) that are not found in canola oil. This indicates that goldfish are capable of producing these fatty acids from 18-carbon precursors. The flesh of fish fed canola oil would be inferior for human nutrition to that offish fed marine oils, due to lower (n-3) PUFA levels. However, the results do indicate that canola oil has good potential as a lipid source in larval cyprinid diets.     

Cellular responses of goldfish, Carassius auratus (L.), to metacercariae of Ribeiroia marini (Faust & Hoffman, 1934)

Goldfish, Carassius auratus (L.), mobilized an acute, non-specific cellular inflammatory response against experimental infection with metacercariae of the digenean Ribeiroia marini in lateral line scale canals. The host cellular response to primary infection consisted of a rapid infiltration of neutrophilic granulocytes and macrophages that adhered to the metacercial cyst wall. Granulocytes released cytoplasmic granules, forming a necrotic zone around the cyst. Outside the necrotic zone, a fibrocytic zone and epithelioid cells formed a matrix upon which reactive leucocytes were attached in a progressive granulomatous encapsulation. Interdigitating epitheliod cells formed a syncytial border separating the necrotic zone from the outer fibrocytic region of the granuloma. Acute inflammatory response preceded the expulsion of most metacercariae from the scale canals. Few cysts remained encapsulated in epidermal tissue. Cellular response to challenge infections was more intense, but no major differences in leucocyte composition between primary and challenge infections suggested a continuum of inflammation and cell-mediated responses. In late primary and challenge infections, the predominant eosinophilic granular cell released cytoplasmic granules. We propose that the eosinophilic granular cell of the goldfish has an antiparasitic function, releasing bioactive granules that alter the environment of the scale canal and cause the expulsion of parasites.     

The stimulatory effect of LED light spectra on genes related to photoreceptors and skin pigmentation in goldfish (Carassius auratus)

This study aimed to assess differences in genes related to skin color of goldfish (Carassius auratus) exposed to light-emitting diodes (LEDs): red, green, and purple. We investigated differences in the expression of mammalian-like melanopsin (Opn4m), rhodopsin (RH), melanin-concentrating hormone (MCH), melanin-concentrating hormone receptor (MCH-R), and proopiomelanocortin (POMC) in goldfish exposed to different LED light spectra. Opn4m, RH, MCH, and MCH-R mRNA levels were significantly higher in the green and purple LED groups than in the white fluorescent bulb (control) and red LED groups. Furthermore, skin cells were isolated to measure the MCH-R mRNA expression levels. The results show that the mRNA expression levels were significantly higher in the green and purple LED groups than in the control and red LED groups. In addition, body weights in the green and purple LED groups were significantly higher than those in the control and red LED groups. However, POMC mRNA expression levels in the green and purple LED groups were significantly lower than those in the control and red LED groups. These results suggest that specific wavelengths regulate fish skin color through neuropeptide hormones and photoreceptors, and POMC, which is related to stress hormones and melatonin, is associated with stress levels as well as skin color.     

Lutein as a natural carotenoid source: Effect on growth,survival and skin pigmentation of goldfish juveniles (Carassius auratus)

This study aimed to evaluate the effect of lutein supplementation on growth, survival and skin pigmentation for goldfish juveniles. Four diets enriched with different carotenoid sources (lutein, astaxanthin, canthaxanthin and a combination of lutein and canthaxanthin) were compared to a control diet without carotenoid supplementation. The carotenoid inclusion level was standardized at 50 mg kg‐1 in all treatments. 240 goldfish juveniles (1.07?0.57 g) were cultivated in 30 aquariums (30L) during 84 days. The experimental design was completely randomized with five treatments and six replicates. The dietary inclusion of carotenoid pigments did not affect the growth and feeding efficiency of goldfish juveniles. Supplementation with lutein presented higher survival values when compared to the other treatments. Astaxanthin and canthaxanthin supplementation increased the concentration of carotenoids on the skin of goldfish juveniles in relation to the control treatment. For the fish fed with the diet containing lutein, the skin pigmentation was as efficient as astaxanthin and canthaxanthin, but did not differ from the control and combined treatment (canthaxanthin + lutein). The lutein supplementation (50 mg kg‐1) improved survival and promoted efficient carotenoid pigmentation on the skin of goldfish juveniles.