Search

The advancement of nanotechnology and the widespread use of nanoparticles (NPs) in various industries have highlighted the importance of studying the potential harmful effects of nanomaterials on organisms. This study aimed to evaluate the lethal toxicity thresholds of Copper Oxide Nanoparticles (CuO-NPs). The investigation focused on examining the sub-lethal toxicity effects of CuO-NPs on blood parameters, as well as their influence on the gill tissue and liver of goldfish (Carassius auratus). Goldfish were exposed to varying concentrations of CuO-NPs (10, 20, 30, 40, 60, 80, and 100 mg/L) for 96 h. The Probit software was employed to determine the LC 50 (lethal concentration causing 50% fish mortality) by monitoring and documenting fish deaths at 24, 48, 72, and 96-hour intervals. Subsequently, sub-lethal concentrations of 5% LC 50 (T1), 10% LC 50 (T2), and 15% LC 50 (T3) of CuO-NPs were administered based on the LC 50 level to investigate their effects on haematological parameters, encompassing the number of red blood cells and white blood cells, hematocrit and haemoglobin levels, mean corpuscular volume, mean corpuscular haemoglobin and mean corpuscular haemoglobin concentration. Additionally, histopathological examinations were conducted on the gill and liver tissues of the studied fish. Results indicated concentration-response of fish mortalities. In general, changes in the blood biochemical parameters of fish exposed to sub-lethal concentrations of CuO-NPs included a significant decrease in leukocyte count and glucose level and an increase in protein and triglyceride levels. Furthermore, an escalation in tissue damage such as gill apical and basal hyperplasia, lamellae attachment, squamous cell swelling, blood cell infiltration, and cellular oedema in gills tissue. and bleeding, increased sinusoidal space, necrosis, lateralization of the nucleus, cell swelling, and water retention in the liver. The findings showed dose-dependent increasing toxicity in goldfish specimens exposed to CuO-NPs., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

The present study investigated dietary Enteromorpha prolifera polysaccharide (EPP) on the growth performance, body composition and non-specific immunity of crucian carp Carassius auratus. Crucian carps with body weight of 51.24 ± 4.08 g were randomly divided into five groups: one group was fed with basic diet, while the other four groups were fed with diets containing 20, 40, 60 and 80 g/kg EPP. After 60 days of feeding, dietary administration of 40 g/kg EPP increased the body weight gain rate, feed conversion ratio, specific growth rate, body crude protein content, intestine digestive enzyme activities, serum lysozyme activity, acid phosphatase activity, alkaline phosphatase activity, complement 3 level, superoxide dismutase activity and catalase activity and decreased body fat content and serum total cholesterol, triacylglycerol, alanine aminotransferase activity and glutamic oxaloacetic transaminase activity compared with those of the control group. All levels of EPP improved the resistance to Aeromonas hydrophilia compared with the control group. The results indicated that EPP could promote the growth of crucian carp and improve their disease resistance and may be used as a dietary supplement for crucian carp., (Copyright © 2020. Published by Elsevier B.V.)

Water temperature directly affects the body temperature in fish, so increasing water temperatures in oceans and rivers will lead to increases in fish body temperatures. Whilst a range of responses of fish to increases in water temperature have been measured, amino acid metabolism in a fish under high water temperature (HT) conditions has not been investigated. The aim of this study was to determine the effects of an acute increase in water temperature on oxygen consumption, plasma cortisol concentrations, and free amino acid concentrations in plasma and several tissues in goldfish (Carassius auratus). Oxygen consumption and plasma cortisol concentrations were increased in goldfish exposed to HT (30 ± 1 °C) for 200 min compared with goldfish at a control water temperature (CT 17 ± 1 °C). Oxygen consumption and plasma cortisol concentrations in both groups of fish combined were positively correlated. When goldfish were exposed to HT for 300 min oxygen consumption and plasma concentrations of 15 free amino acids were increased compared with goldish at CT. Concentrations of several free amino acids were increased to varying extents in the brain, liver, and muscle tissues. In conclusion, an acute increase in water temperature affected amino acid metabolism differently in the brain, liver, and muscle tissues. Goldfish will be a useful species for further studies of the possible roles of various amino acids in the brain, muscle, and liver during acute increases in water temperature in fish.

Lead (Pb), a heavy metal and an environmental stressor, may affect many physiological processes, including the serum index and the immune response. The aim of this study was to explore the toxic effects of Pb on the serum index and the immune response of Carassius auratus gibelio (C. gibelio) fed 0, 120, or 240 mg/kg Pb, and 10 9  cfu/g Bacillus subtilis (B. subtilis). After 15 and 30 days of dietary exposure, the serum indices and the immune responses of the fish were assessed. Dietary Pb exposure significantly affected various components of the serum index, including calcium, magnesium, glucose, cholesterol, total protein, glutamic-pyruvic transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), and lactate dehydrogenase (LDH). However, sIgA activity in the gut increased significantly following B. subtilis supplementation. Notable changes were also observed in the expression levels of immune-related genes, including HSP70, IgM, HSP90, IL-1β, IL-6, and TNF-α. B. subtilis supplementation effectively attenuated the effects of dietary Pb exposure.

A 30-days feeding trail was conducted to determine the sensitivity of Carassius auratus to the toxicological effects of elevated dietary Selenomethionine (Se-Met). C. auratus averaging 23.56 ± 1.82 g were exposed to four Se-Met concentrations (mg Se/kg): 0 (Se-Met0), 5 (Se-Met5), 10 (Se-Met10) and 20 (Se-Met20) to estimate the effects on tissue selenium (Se) accumulation, blood biochemical profiles, transcript expression and intestinal microbiota. Se accumulated in the kidney, liver and muscle in a dose-dependent manner and followed this order: kidney > liver > muscle, the highest accumulation were obtained in kidney of Se-Met20 diet after 30 days of feeding. Serum contents of alanine transaminase (ALT), aspartate transaminase (AST) and alkaline phosphatase (ALP) in fish exposed to Se-Met20 group was significantly highest among Se-Met exposure groups. Hydrogen peroxide (H 2 O 2 ) concentrations in liver were affected by dietary Se-Met exposures. Liver contents of total antioxidant capacity (TAC), catalase (CAT), glutathione peroxidase (GPx) and malondialdehyde (MDA) in fish exposure to Se-Met5 group was significantly highest among Se-Met exposure groups. Growth hormone receptor (GHR), insulin-like growth factor 1 (IGF-1) and antioxidant enzyme related genes including glutathione peroxidase (GPx), catalase (CAT) and glutathione S-transferase (GST) expression in liver were down-regulated with the concentration of Se-Met exposure groups. The results of high-throughput sequencing showed that gut microbial communities and hierarchy cluster heatmap analysis were significantly affected by Se-Met exposure. The abundances of Cetobacterium and Vibrio increased while fish exposed to Se-Met20 group. The abundance of Ralstonia increased when the Se-Met exposure dose reached 10 mg Se kg -1 . The results suggested that the exposure to elevated dietary Se-Met may result toxic effects in C. auratus., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Fish are the most diverse species of all vertebrate groups, and their blood cells have shown variable characteristics in terms of morphology. Cytochemical staining for enzyme activity in blood leukocytes will help assess the immune function of fish. We characterize blood cells from crucian carp (Carassius auratus) and grass carp (Ctenopharyngodon idellus) by using a Diff-Quick stain as well as different cytochemical methods. Blood specimens obtained from crucian carp and grass carp were evaluated after cytochemical staining for acid phosphatase (ACP), alkaline phosphatase (ALP), naphthol AS chloroacetate esterase (AS-DNCE), naphthyl acetate esterase (NAE), α-naphthyl butyrate esterase (NBE), peroxidase (MPO) and periodic acid-Schiff's reaction (PAS) using commercial kits. Blood cell types were evaluated based on their morphological characteristics and the presence or absence of specific chromogen. The expression pattern of enzymes was similar between the two Cyprinidae and was also broadly consistent with other fish species. However, there were some interesting differences detected between crucian carp and grass carp, including naphthol AS chloroacetate esterase activity in monocytes, peroxidase activity and location in thrombocytes. The ACP, ALP and MPO expressions of different leukocytes of the two Cyprinidae were evaluated by Image Pro Plus and were analysed for statistical significant differences. This investigation provides basic haematology and enzyme activity analyses for crucian carp and grass carp and serves as an approach to evaluating the immune response of fish., (© 2018 Blackwell Verlag GmbH.)

During the present research, C. idella and C. auratus fish were exposed to 2 ppm concentration of imidacloprid for 28 and 24 days, respectively, and the effect on biochemical and haematological parameters was investigated. During the study of biochemical parameters, there occurred significant increase (P<0.05) in the serum levels of ALT and creatinine of imidacloprid exposed groups of both species of fish. The level of serum albumin of imidacloprid exposed groups of both fish species was significantly lower as compared to control group (P<0.05). Serum globulin level in imidacloprid exposed group of C. idella was insignificantly lower as compared to control group, however the serum globulin level of C. auratus was significantly lower than the control group (P < 0.05). The level of total proteins in serum of imidacloprid exposed groups of both fish species was insignificantly lower as compared to control groups (P>0.05). During the study of haematological parameters, TLC of C. idella was insignificantly (P>0.05) higher than control group but the TLC of C. auratus was significantly (P<0.05) higher than control. There was also observed increasing trend in the percentage of neutrophils and lymphocytes of imidacloprid exposed group of each fish species. The platelets count of imidacloprid exposed group of each fish species was significantly (P<0.05) lower than control group. The haemoglobin concentration of imidacloprid exposed group of C. idella was significantly lower than control group (P<0.05). In case of C. auratus, the haemoglobin level of imidacloprid exposed group was insignificantly lower than control group (P>0.05). From the finding of the present research it was concluded that 28 days exposure of C. idella and 24 days exposure of C. auratus to 2 ppm concentration of imidacloprid does not cause mortality however the exposure causes alteration in the normal level of biochemical and haematological parameters.

The pharmacokinetic profiles of florfenicol (FF) or florfenicol amine (FFA) in crucian carp were compared at different water temperatures after single intramuscular administration of FF at 10 mg/kg bodyweight. The concentrations of FF and FFA were determined by a high-performance liquid chromatography method, and then, the concentration versus time data were subjected to compartmental analysis using a one-compartment open model. At the water temperatures of 10, 20, and 25°C, the peak concentrations (C max s) of FF were 2.28, 2.29, and 2.34 μg/ml, respectively, while those of FFA were 0.42, 0.71, and 0.82 μg/ml, respectively. And the absorption half-life (t 1/2ka ) of FF was 0.21, 0.19, and 0.21 hr, while the elimination half-life (t 1/2kel ) was 31.66, 24.77, and 21.48 hr, respectively. For FFA, the formation half-life (t 1/2kf ) was 3.85, 8.97, and 12.43 hr, while the t 1/2kel was 58.34, 30.27, and 21.22 hr, respectively. The results presented here demonstrated that the water temperature had effects on the elimination of both FF and FFA and the formation of FFA. Based on the T > MIC values calculated here, to treat the infections of bacterial with MIC value ≤ 0.5 μg/ml, FF intramuscularly given at 10 mg/kg bodyweight with a 72-hr interval is sufficient at the water temperature of 10°C, while the intervals of 60 and 48 hr were needed at 20 and 25°C, respectively. But to treat bacterial with higher MIC values, more FF or FF at 10 mg/kg BW but with shorter intervals should be intramuscularly given to the infected fish., (© 2018 John Wiley & Sons Ltd.)

The pharmacokinetics (PK) of cefquinome (CEQ) was studied in crucian carp (Carassius auratus gibelio) after single oral, intramuscular (i.m.), and intraperitoneal (i.p.) administration at a dose of 10 mg/kg body weight and following incubation in a 5 mg/L bath for 5 hr at 25°C. The plasma concentration of CEQ was determined using high-performance liquid chromatography (HPLC). PK parameters were calculated based on mean CEQ concentration using WinNonlin 6.1 software. The disposition of CEQ following oral, i.m., or i.p. administration was best described by a two-compartment open model with first-order absorption. After oral, i.m., and i.p. administration, the maximum plasma concentration (C max ) values were 1.52, 40.53, and 67.87 μg/ml obtained at 0.25, 0.23, and 0.35 hr, respectively, while the elimination half-life (T 1/2β ) values were 4.68, 7.39, and 6.88 hr, respectively; the area under the concentration-time curve (AUC) values were 8.61, 339.11, and 495.06 μg hr/ml, respectively. No CEQ was detected in the plasma after bath incubation. Therapeutic blood concentrations of CEQ can be achieved in the crucian carp following i.m. and i.p. administration at a dosage of 10 mg/kg once every 2 days., (© 2018 John Wiley & Sons Ltd.)

Lead (Pb) poisoning in humans and fish represents a significant global problem. Bacillus subtilis (B. subtilis) is a widely used probiotic in aquaculture. Carassius auratus gibelio (C. gibelio) is one of the most important aquaculture species with great commercial value. The objective of this study was to evaluate the potential of B. subtilis in ameliorating lead-induced toxicity in C. gibelio. The fish were exposed for 60 days to waterborne Pb at 0, 0.05, 0.5 and 1 mg/L and/or dietary B. subtilis at 10 9 cfu/g. After 30 and 60 days, the fish were sampled and bioaccumulation, antioxidant activity and immune responses were assessed. The results revealed that B. subtilis confers significant protective effects against lead toxicity by preventing alterations in the levels of bioaccumulation, superoxide dismutase, catalase and glutathione. B. subtilis also assists in the recovery of blood δ-aminolevulinic acid dehydratase, lysozyme, and IgM levels while regulating the expression of immune-related genes including IL-10, lysozyme, TNF-α, IgM and Hsp70 after 60 days of lead exposure. Our results suggest that administration of B. subtilis (10 9 cfu/g) has the potential to combat lead toxicity in C. gibelio., (Copyright © 2018. Published by Elsevier Inc.)

This study tested the hypothesis that blood-borne prostaglandin F 2α (PGF 2α ) produced at the time of ovulation by female goldfish, a typical scramble-spawning, egg-laying cyprinid fish, functions as a hormone which stimulates female sexual receptivity, behavior, and pheromone release, thereby synchronizing female mating behavior with egg availability. We conducted 5 experiments. First, we tested whether PGF 2α is found in the blood of female fish and if it increases at the time of ovulation. Using gas chromatography-mass spectrometry, we found that circulating PGF 2α was approximately 1 ng/ml prior to ovulation, increased over 50-fold within 3 h of ovulation and returned to preovulatory values after spawning and egg release. Ovulated fish also released over 2 ng/h of PGF 2α and 800 ng/h of 15-keto-PGF 2α , a metabolite of PGF 2α - both compounds with known pheromonal function. Second, we tested how closely levels of circulating PGF 2α tracked the timing of ovulation by sampling fish at the time of ovulation and discovered that PGF 2α increased within 15 min of ovulation, peaked after 9 h, and fell to basal levels as fish spawned and released their eggs. Third, we tested whether an interaction between eggs and the reproductive tract serves as a source of circulating PGF 2α and its relationship with female sexual receptivity by injecting ovulated eggs (or an egg-substitute) into the reproductive tract of females stripped of ovulated eggs. We found both of these treatments elicited measurable increases in plasma PGF 2α as well as female sexual behavior. A fourth experiment showed that indothemacin, a PG synthase inhibitor, blocked both PGF 2α increase and female sexual behavior in egg-substitute-injected fish. Finally, we tested the relationship between the expression of female behavior and PGF 2α in PGF 2α -injected fish and found that circulating PGF 2α levels closely paralleled behavior, rising within 15 min and peaking at 45 min. Together, these experiments establish that PGF 2α functions as a behavioral blood-borne hormone in the goldfish, suggesting it likely has similar activity in other related, externally-fertilizing fishes., (Crown Copyright © 2018. Published by Elsevier Inc. All rights reserved.)

This study investigated the effect of dietary supplementation of probiotic Pedicoccus acidilactici and nucleotide (separately or combined) on growth performance, intestinal microbiota, hemato-immunological parameters, and immunity response in goldfish (Carassius auratus). Fish (average weight 5-6 g) were acclimatized and divided into eight experimental diets supplemented with P. acidilactici of different concentrations (0.1, 0.2, and 0.3% diet) and nucleotides (0 and 0.5% diet) for 6 months. Fish fed with experimental diets showed significant differences in terms of final weight, weight gain, feed conversion ratio, daily growth rate, and condition factor when compared to control diet (P < 0.05). Fish fed with probiotic (0.3%) separately and combined with nucleotide (0.5%) had highest RBC and WBC when compared to other diets (P < 0.05), while the highest values for Hb and Hct as well as total protein, glucose, albumin, and globulin were observed in probiotic (0.2%) and nucleotide (0.5%) combined diet. Serum lysozyme and anti-protease activities were significantly higher in probiotic (0.1 and 0.2%) and nucleotide (0.5%) combined diets. Similarly, these two diets combined showed the highest colonization of P. acidilactici when compared to other diets. In conclusion, combined dietary probiotic and nucleotide improve the growth performance, hemato-biochemical parameters, and intestine growth in C. auratus.

Cross-regulation occurs at many points between the hypothalamic-pituitary-gonad (HPG) and hypothalamic-pituitary-thyroid (HPT) axes. Monocrotophos (MCP) pesticide could disrupt HPG and HPT axes, but its direct target within the endocrine system is still unclear. In the present study, hormone concentrations and transcriptional profiles of HPG and HPT genes were examined in male goldfish (Carassius auratus) exposed to 0, 4, 40, and 400 μg/L MCP for 2, 4, 8, and 12 d. In vivo data were analyzed by multiple linear regression and correlation analysis, quantitatively indicating that MCP-induced plasma 17β-estradiol (E 2 ) levels were most associated with alteration of cyp19a transcription, which was also a potential point indirectly modulated by the MCP-altered thyroid hormones (THs) status; disturbance of THs pathways was most related with effect of MCP on regulation of the hypothalamic-pituitary hormones involved in the thyroid system, and the increased E 2 levels might enhance the impact of MCP on HPT axis by modulating hepatic deiodinase expression. Our finding, based on these correlational data, gave a whole view of the regulations, especially on the cross-talk between sex hormone and thyroid hormone pathways upon exposure to chemicals with unknown direct target in vivo, and cautions should be exercised when developing adverse outcome pathway networks for reproductive and thyroidal endocrine disruption., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The pharmacokinetics of orbifloxacin was studied after a single dose (7.5 mg/kg) of intravenous or intramuscular administration to crucian carp (Carassius auratus) reared in freshwater at 25°C. Plasma samples were collected from six fish per sampling point. Orbifloxacin concentrations were determined by high-performance liquid chromatography with a 0.02 μg/ml limit of detection, then were subjected to noncompartmental analysis. After intravenous injection, initial concentration of 5.83 μg/ml, apparent elimination rate constant (λ z ) of 0.039 hr -1 , apparent elimination half-life (T 1/2λz ) of 17.90 hr, systemic total body clearance (Cl) of 75.47 ml hr -1  kg -1 , volume of distribution (Vz) of 1,948.76 ml/kg, and volume of distribution at steady-state (Vss) of 1,863.97 ml/kg were determined, respectively. While after intramuscular administration, the λ z , T 1/2λz , mean absorption time (MAT), absorption half-life (T 1/2ka ), and bioavailability were determined as 0.027 hr -1 , 25.69, 10.26, 7.11 hr, and 96.46%, respectively, while the peak concentration was observed as 3.11 ± 0.06 μg/ml at 2.0 hr. It was shown that orbifloxacin was completely but relatively slowly absorbed, extensively distributed, and slowly eliminated in crucian carp, and an orbifloxacin dosage of 10 mg/kg administered intravenously or intramuscularly would be expected to successfully treat crucian carp infected by strains with MIC values ≤0.5 μg/ml., (© 2018 John Wiley & Sons Ltd.)

We examined the effects of α-melanocyte-stimulating hormone (α-MSH) on bone metabolism using regenerating goldfish scales. Normally developed scales on the bodies of goldfish were removed to allow the regeneration of scales under anesthesia. Thereafter, the influence of α-MSH on the regeneration of goldfish scales was investigated in vivo. In brief, α-MSH was injected at a low dose (0.1 μg/g body weight) or a high dose (1 μg/g body weight) into goldfish every other day. Ten days after removing the scales, we collected regenerating scales and analyzed osteoblastic and osteoclastic activities as respective marker enzyme (alkaline phosphatase for osteoblasts, tartrate-resistant acid phosphatase for osteoclasts) activity in the regenerating scales as well as plasma calcium levels. At both doses, osteoblastic and osteoclastic activities in the regenerating scales increased significantly. Plasma calcium concentrations in the α-MSH-treated group (high doses) were significantly higher than those in the control group. Next, in vitro experiments were performed to confirm the results of in vivo experiments. In the cultured regenerating scales, osteoblastic and osteoclastic activities significantly increased with α-MSH (10 -7 and 10 -6  M) treatment. In addition, real-time PCR analysis indicated that osteoclastogenesis in α-MSH-treated scales was induced by the receptor activator of the NF-κB/receptor activator of the NF-κB ligand/osteoprotegerin pathway. Furthermore, we found that α-MSH receptors (melanocortin receptors 4 and 5) were detected in the regenerating scales. Thus, in teleosts, we are the first to demonstrate that α-MSH functions in bone metabolism and promotes bone resorption via melatonin receptors 4 and/or 5., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Mebendazole is approved for use in aquatic animals and is widely used in Chinese aquaculture. We developed a pharmacokinetic and residue analysis for mebendazole levels in the goldfish (Carassius auratus). Plasma and muscle samples of C. auratus were taken after oral administration of 10 mg/kg mebendazole. The maximal drug plasma concentration of 0.55 mg/L was achieved at 48 hr and then declined with the elimination half-life (T 1/2β ) of 7.99 hr. Administration of 10 mg/kg by oral gavage for 5 successive days resulted in a peak mebendazole concentration of 0.70 mg/kg in muscle at 96 hr after the last dose. The drug was then eliminated at a relatively slow rate from muscle with T 1/2β of 68.41 hr. There was no detectable mebendazole in any muscle samples at 24 days postadministration. The AUC last in plasma and muscle was 19.42 and 105.33 mg hr/L, respectively. These data provide information for dosage recommendations and withdrawal time determinations for mebendazole use in aquariums., (© 2017 John Wiley & Sons Ltd.)

Diel cyclic hypoxia occurs with varying frequency and duration in freshwater habitats, yet little is known about its effects on reproduction of freshwater fishes. The present study shows that long-term exposure of goldfish (Carassius auratus) to cyclic hypoxia (0.8 ± 0.2 mg/l dissolved oxygen) for 9 h or more, per day, altered plasma lipid and sex steroid profiles, which in turn directly or indirectly suppressed ovarian growth and viable spermatozoa production. Hypoxia decreased total cholesterol and high density lipoprotein (HDL p < 0.05) and elevated triglycerides (TG; p < 0.05) in both sexes. Plasma steroid concentrations particularly of 17α-hydroxyprogesterone (17-HP), estradiol (E2), testosterone (T) in females, and T and 11-ketotestosterone (11-KT) in males were attenuated under diel hypoxic conditions. Intriguingly, both diel and continuous hypoxia elevated plasma E2 and vitellogenin levels in males. However, neither lipid nor steroid profiles recorded any variation in a dose-dependent manner in response to diel hypoxia. The reduced GSI, decreased number of tertiary oocytes, and motile spermatozoa in hypoxic fish clearly indicate suppression of gametogenesis. Thereby, prolonged diel cyclic hypoxia may affect valuable fishery resources and fish population structure by impairing reproductive performances and inducing estrogenic effects in males.

Reference intervals for diagnostic tests are vitally important for clinical decision making. Despite the popularity of pet goldfish (Carassius auratus), reference intervals have not been generated for routine biochemistry panel analytes in this species. This study establishes de novo reference intervals for packed cell volume and total solids, using 47 apparently healthy immature goldfish, and for 11 common chemistry panel analytes (albumin, aspartate aminotransferase, calcium, creatine kinase, globulin, blood glucose, sodium, potassium, phosphorous, total protein, and uric acid) using 39 immature goldfish. Robust reference intervals were generated following recommendations of the American Society for Veterinary Clinical Pathology. Linear regression was used to demonstrate a statistically significant relationship between body weight and calcium, albumin, total protein, potassium, packed cell volume, and total solids. The results of this study serve as a useful baseline for future reference interval generation in goldfish.

The rate of hypoxia induction (RHI) is an important but overlooked dimension of environmental hypoxia that may affect an organism's survival. We hypothesized that, compared with rapid RHI, gradual RHI will afford an organism more time to alter plastic phenotypes associated with O 2 uptake and subsequently reduce the critical O 2 tension ( P crit ) of the rate of O 2 uptake ( Ṁ O 2 ). We investigated this by determining P crit values for goldfish exposed to short (∼24 min), typical (∼84 min) and long (∼480 min) duration P crit trials to represent different RHIs. Consistent with our predictions, long duration P crit trials yielded significantly lower P crit values (1.0-1.4 kPa) than short and typical duration trials, which did not differ (2.6±0.3 and 2.5±0.2 kPa, respectively). Parallel experiments revealed these time-related shifts in P crit were associated with changes to aspects of the O 2 transport cascade that took place over the hypoxia exposures: gill surface areas and haemoglobin-O 2 binding affinities were significantly higher in fish exposed to gradual RHIs over 480 min than fish exposed to rapid RHIs over 60 min. Our results also revealed that the choice of respirometric technique (i.e. closed versus intermittent) does not affect P crit or routine Ṁ O 2 , despite the significantly reduced water pH and elevated CO 2 and ammonia levels measured following closed-circuit P crit trials of ∼90 min. Together, our results demonstrate that gradual RHIs result in alterations to physiological parameters that enhance O 2 uptake in hypoxic environments. An organism's innate P crit is therefore most accurately determined using rapid RHIs (<90 min) so as to avoid the confounding effects of hypoxic acclimation., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2017. Published by The Company of Biologists Ltd.)

Monocrotophos (MCP) is an organophosphorus pesticide that is median-toxic to fish. MCP pesticide resulted in an increase of 17 beta estradiol following a decrease in testosterone in male goldfish (Carassius auratus). To fully understand the mechanism of MCP pesticide that causes the imbalance between male and female hormones, we determined the levels of plasma cholesterol, spermatic steroidogenic acute regulatory protein mRNA, steroidogenesis enzyme mRNA, plasma sex hormone synthesis intermediates, and effectual hormones in male goldfish exposed to MCP pesticide at nominal concentrations of 0.01, 0.10, and 1.00 mg/L for 21 days in a semi-static exposure system. The results indicated that MCP pesticide (a) led to decreased steroidogenic acute regulatory protein mRNA levels; (b) decreased mRNA levels of cholesterol side chain cleavage enzyme and cytochrome P450 17 alpha hydroxylase, which are steroidogenesis enzymes involved in androgen synthesis; and (c) increased cytochrome P450 aromatase mRNA levels, a steroidogenesis enzyme involved in the synthesis of effectual estrogen. The present study provides evidence that MCP pesticide affects synthesis and conversion of sex steroids through multiple targets in male goldfish.

While magnesium requirements for teleost fish highlight the physiological importance of this cation for homeostasis, little is known regarding the molecular identity of transporters responsible for magnesium absorption or secretion. The recent characterization of the vertebrate magnesium transporter solute carrier 41a1 (SLC41a1) in the kidney of a euryhaline fish has provided a glimpse of possible moieties involved in piscine magnesium regulation. The present study obtained a novel SLC41a1 coding sequence for Carassius auratus and demonstrated ubiquitous expression in all tissues examined. Transcriptional regulation of SLC41a1 in response to dietary and environmental magnesium concentrations was observed across tissues. Specifically, decreased environmental magnesium correlated with decreased expression of SLC41a1 in the intestine, whereas the gill and kidney were unaffected. Dietary magnesium restriction correlated with decreased expression of SLC41a1 in the intestine and gill, while again no effects were detected in the kidney. Finally, elevated dietary magnesium correlated with increased expression of SLC41a1 in the kidney, while expression in the intestine and gill remained stable. Plasma magnesium was maintained in all treatments, and dietary assimilation efficiency increased with decreased dietary magnesium. Consumption of a single meal failed to impact SLC41a1 expression, and transcript abundance remained stable over the course of digestion in all treatments. Transcriptional regulation occurred between 7 and 14days following dietary and environmental manipulations and short-term regulation (e.g. <24h) was not observed. Overall the data supports transcriptional regulation of SLC41a1 reflecting a possible role in magnesium loss or secretion across tissues in fish., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Metabolic rate depression (MRD) has long been proposed as the key metabolic strategy of hypoxic survival, but surprisingly, the effects of changes in hypoxic O 2 tensions ( P w O 2 ) on MRD are largely unexplored. We simultaneously measured the O 2 consumption rate ( Ṁ O 2 ) and metabolic heat of goldfish using calorespirometry to test the hypothesis that MRD is employed at hypoxic P w O 2  values and initiated just below P crit , the P w O 2   below which Ṁ O 2  is forced to progressively decline as the fish oxyconforms to decreasing P w O 2 Specifically, we used closed-chamber and flow-through calorespirometry together with terminal sampling experiments to examine the effects of P w O 2  and time on Ṁ O 2 , metabolic heat and anaerobic metabolism (lactate and ethanol production). The closed-chamber and flow-through experiments yielded slightly different results. Under closed-chamber conditions with a continually decreasing P w O 2 , goldfish showed a P crit of 3.0±0.3 kPa and metabolic heat production was only depressed at P w O 2  between 0 and 0.67 kPa. Under flow-through conditions with P w O 2  held at a variety of oxygen tensions for 1 and 4 h, goldfish also initiated MRD between 0 and 0.67 kPa but maintained Ṁ O 2  to 0.67 kPa, indicating that P crit is at or below this P w O 2 Anaerobic metabolism was strongly activated at P w O 2   ≤1.3 kPa, but only used within the first hour at 1.3 and 0.67 kPa, as anaerobic end-products did not accumulate between 1 and 4 h exposure. Taken together, it appears that goldfish reserve MRD for near-anoxia, supporting routine metabolic rate at sub- P crit P w O 2  values with the help of anaerobic glycolysis in the closed-chamber experiments, and aerobically after an initial (<1 h) activation of anaerobic metabolism in the flow-through experiments, even at 0.67 kPa P w O 2 ., (© 2017. Published by The Company of Biologists Ltd.)

Refuse leachate is likely an important source of androgens. However, common in vitro bioassays underestimate the potential androgenic activity of leachate, owing to non-receptor-mediated mechanisms that modify the balance of sex hormones and promote the accumulation of endogenous androgens. This study aimed to develop an in vivo assay by using multiple biomarkers related to testosterone synthesis and conversion for assessing the potential androgenic activity of refuse leachate sampled from a municipal solid waste treatment plant in Qingdao, China. The results indicated that exposure to leachate increased the levels of testosterone and luteinizing hormone, but decreased those of 17β-estradiol in both male and female goldfish (Carassius auratus), suggesting a potential androgenic activity. Further, Leydig cell hyperplasia and decreased gonadal P450 aromatase mRNA levels were observed; these alterations might promote the biosynthesis of testosterone and hinder the conversion of testosterone to 17β-estradiol, which in turn enhance testosterone accumulation. Exposure to leachate also resulted in reproductive impairments, including decreased gonadosomatic index and plasma vitellogenin levels of female goldfish, as well as decreased testicular enzyme activities in male goldfish. The integrated use of biochemical, molecular, and histological markers not only improved our understanding of the androgenic effects of leachate but also verified the reliability and validity of the results. Therefore, the in vivo bioassay described in this study might allow the investigation of the androgenic effects of other complex contaminant mixtures in the future., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The four experimental groups were carried out to test the response of crucian carp Carassius auratus to ammonia toxicity and taurine: group 1 was injected with NaCl, group 2 was injected with ammonium acetate, group 3 was injected with ammonium acetate and taurine, and group 4 was injected with taurine. Fish in group 2 had the highest ammonia and glutamine contents, and the lowest glutamate content in liver and brain. Serum superoxide dismutase (SOD), glutathione (GSH) activities, red cell count (RBC), white cell count (WBC), lysozyme (LYZ) activity, complement C3 content of fish in group 2 reflected the lowest, but malondialdehyde content was the highest. Importantly, serum SOD and GSH activites, RBC, WBC, and LYZ activity, C3, C4 and total immunoglobulin contents of fish in group 3 were significantly higher than those of fish in group 2. This study indicates that ammonia exerts its toxic effects by interfering with amino acid transport, inducing ROS generation, leading to malondialdehyde accumulation and immunosuppression of crucian carp. The exogenous taurine could mitigate the adverse effect of high ammonia level on fish physiological disorder., (Copyright © 2016 Elsevier Inc. All rights reserved.)

To elucidate the potential thyroid disrupting effects of refuse leachate on females, female goldfish (Carassius auratus) were exposed to 0.5% diluted leachates from each step of a leachate treatment process (i.e. raw leachate before treatment, after membrane bioreactor treatment, and the final treated leachate) for 21days. Raw leachate exposure caused disturbances in the thyroid cascade of female fish, as evidenced by the elevated plasma 3,3',5-triiodo-l-thyronine (p<0.05) and thyroid-stimulating hormone (p<0.01) levels as well as up-regulated hepatic and gonadal type I deiodinase (p<0.01), type II deiodinase (p<0.01) and thyroid receptor (p<0.05) mRNA levels. Thyroid disrupting potency decreased markedly as raw leachate progressed through the "membrane bioreactor + reverse osmosis" treatment but could still be detected in the treated leachate. As our results indicated, thyroid system in female goldfish was more sensitive to leachate exposure than that of the male fish., (Copyright © 2016 Elsevier B.V. All rights reserved.)

In the present study, the effect of copper was examined in the common goldfish (Carassius auratus auratus). Fish were fasted and exposed to either a high (0.84μM), a low (0.34μM) or a control copper concentration (0.05μM) for 1 and 7days. Swimming performance was not affected by either fasting or copper exposure. Food deprivation alone had no effect on ionoregulation, but low plasma osmolality levels and plasma Na + were noticed in fasted fish exposed to Cu for 7days. Both gill Na + /K + -ATPase and H + -ATPase activities were undisturbed, while both kidney ATPase activities were up-regulated when challenged with the high Cu levels. Up-regulated kidney ATPase activities likely acted as compensatory strategy to enhance Na + reabsorption. However, this up-regulation was not sufficient to restore Na + to control levels in the highest exposure group., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The influence of sodium fluoride (NaF) on calcium metabolism was examined in goldfish (fresh water teleost). At 2days after administration of NaF (500ng/g body weight; 5μg/g body weight) (around 10(-5) to 10(-4)M in goldfish), we indicated that plasma calcium levels upregulated in both doses of NaF-treated goldfish. To examine the mechanism of hypercalcemia by NaF treatments, therefore, direct effects of NaF on osteoblasts and osteoclasts in goldfish were investigated by an original assay system using teleost scale which has osteoblasts, osteoclasts and bone matrix. Alkaline phosphatase activity in the scales increased with the treatment of NaF (10(-6) and 10(-5)M) during 6h of incubation. Also, tartrate-resistant acid phosphatase activity increased after exposure to NaF (10(-5)M) at the 6h of incubation. To investigate the osteoclastic activation, the mRNA expression of osteoclastogenesis related factors were examined. The receptor activator of the nuclear factor-κB ligand (RANKL) which is known as a factor for osteoclastogenesis, increased in the NaF-treated scales after 6h of incubation. The ratio of RANKL/osteoprotegerin (osteoclastogenesis inhibitory factor) significantly increased after 6h of incubation. Resulting from the increase of RANKL mRNA level, the expression of transcription-regulating factors was significantly increased. Furthermore, the expression of functional genes, cathepsin K and matrix metalloproteinase-9 mRNA, was significantly increased. In our knowledge, this is the first report concerning the effects of NaF on osteoblasts and osteoclasts in teleosts. We concluded that NaF influences calcium metabolism via osteoclastic activation in goldfish., (Copyright © 2016 Elsevier Inc. All rights reserved.)

This study investigated the effects of increasing water temperature (22-30 °C) on the physiological stress response and immunity of goldfish, Carassius auratus, and the ability of green light-emitting diode (LED) irradiation or melatonin injections to mitigate this temperature-induced stress. To evaluate the effects of either green-wavelength LED light or melatonin on stress in goldfish, we measured plasma triiodothyronine (T3), thyroxine (T4), and thyroid hormone receptor (TR) mRNA expression; plasma cortisol and glucose; and immunoglobulin M (IgM) and lysozyme mRNA expression. The thyroid hormone activities, TR mRNA expression, and plasma cortisol and glucose were higher in goldfish exposed to high-temperature water, but were lower after exposure to melatonin or green-wavelength LED light. Lysozyme mRNA expression and plasma IgM activity and protein expression were lower after exposure to high water temperatures and higher after melatonin or green-wavelength LED light treatments. Therefore, high water temperature induced stress and decreased immunity; however, green-wavelength LED light and melatonin treatments mitigated the effects of stress and enhanced immunity. The benefits of melatonin decreased with time, whereas those of green-wavelength LED treatment did not.

In this study with crucian carp (Carassius auratus gibelio), the effect on enrofloxacin (EF) and its metabolite ciprofloxacin (CF) and on the activity of cytochrome P450 1A (CYP1A) and cytochrome P450 3A (CYP3A) was estimated following the oral administration of rifampicin (RIF) (12mg/kg) and β-naphthoflavone (BNF) (12mg/kg), respectively. First, reversed-phase high-performance liquid chromatography (RP-HPLC) was used to detect the pharmacokinetics of EF with continual blood sampling. In RIF-treated, BNF-treated and control groups, the value of the CmaxCF/CmaxEF ratio was 4.41, 0.81 and 0.95, and the corresponding value of the AUC0-t-CF/AUC0-t-EF ratio was 3.69, 1.84 and 1.76, respectively. In the RIF-treated, BNF-treated and control groups, the MRT values of EF were 26.57, 27.45 and 30.88h, and the corresponding values for CF were 5.79, 35.18 and 38.11h, respectively. Based on these results for crucian carp, the accumulation and elimination of EF and CF in the RIF-treated group were more rapid than in BNF-treated and control groups. Second, liver microsomes were pretreated with the inducer of CYP1A for BNF and that of CYP3A for RIF, and then the enzymatic activities of CYP1A and CYP3A were measured, respectively. The activities of ethoxyresorufin-O-deethylation (EROD) and erythromycin-N-demethylation (ERND) increased significantly (P<0.05) for CYP1A and CYP3A, respectively. However, in further experiments on the formation of CF, the level of EF N-deethylation was significantly induced by RIF and inhibited by ketoconazole (KTZ) for CYP3A but had no influence for CYP1A, BNF and berberine chloride (BER). We concluded that CYP3A might be responsible for the N-deethylation of EF and because of this activity, could also serve as a toxicity biomarker in crucian carp., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The periprandial profile and effects of short- (7 days) and long-term (30 days) fasting on the ghrelinergic system were studied in goldfish (Carassius auratus). Plasma levels of acyl-ghrelin, desacyl-ghrelin, and ghrelin O-acyl transferase (GOAT) were analyzed by enzymoimmunoassays, and expression of preproghrelin, goat and growth hormone secretagogue receptors (ghs-r) was quantified by real-time PCR. Circulating levels of acyl-ghrelin and GOAT rise preprandially, supporting the role of acyl-ghrelin as a meal initiator in this teleost. Consistently, preproghrelin and ghs-r1a1 expression increases 1 h before feeding time in intestinal bulb, suggesting that this receptor subtype might be involved in the preprandial action of ghrelin in this tissue. Significant postfeeding variations are detected for preproghrelin in telencephalon, goat in telencephalon and hypothalamus, ghs-r1a1 in vagal lobe, ghs-r1a2 and ghs-r2a1 in hypothalamus and ghs-r2a2 in telencephalon and vagal lobe, especially in unfed fish. Short- and long-term fasting significantly increase preproghrelin expression in telencephalon and gut. Goat expression is upregulated by short-term fasting in telencephalon and hypothalamus, and by both short- and long-term fasting in gut. Expression of ghs-r increases by fasting in telencephalon, while an upregulation of type 2, but not type 1, receptors is observed in vagal lobe. In intestinal bulb, ghs-r1a2 transcripts increase after both short- and long-term fasting. These results show a high dependence of the ghrelinergic system on feeding and nutritional status in fish, and demonstrate for the first time a differential implication of the various components of this system suggesting different roles for the four ghrelinergic receptor subtypes.

We investigated the effect of light spectra on retinal damage and stress in goldfish using green (530 nm) and red (620 nm) light emitting diodes (LEDs) at three intensities each (0.5, 1.0, and 1.5 W/m(2)). We measured the change in the levels of plasma cortisol and H2O2 and expression and levels of caspase-3. The apoptotic response of green and red LED spectra was assessed using the terminal transferase dUTP nick end labeling (TUNEL) assay. Stress indicator (cortisol and H2O2) and apoptosis-related genes (caspase-3) decreased in green light, but increased in red light with higher light intensities over time. The TUNEL assay revealed that more apoptotic cells were detected in outer nuclear layers after exposure to red LED over time with the increase in light intensity, than the other spectra. These results indicate that green light efficiently reduces retinal damage and stress, whereas red light induces it. Therefore, red light-induced retina damage may induce apoptosis in goldfish retina., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Combination of bioassays and chemical analysis was applied to determine the genotoxic/mutagenic contamination in four different sites of the basin of Lake Sevan in Armenia. Water genotoxicity was evaluated using the single cell gel electrophoresis technique (comet assay) in erythrocytes of gibel carp (Carassius auratus gibelio), Tradescantia micronucleus (Trad-MCN) and Tradescantia stamen hair mutation (Trad-SHM) assays. Significant inter-site differences in the levels of water genotoxicity according to fish and Trad-MCN bioassays have been revealed. Two groups of locations with lower (south-southwest of the village Shorzha and Peninsula of Lake Sevan) and higher (estuaries of Gavaraget and Dzknaget rivers) levels of water genotoxicity were distinguished. Correlation analysis support the hypothesis that the observed genetic alterations in fish and plant may be a manifestation of the effects of water contamination by nitrate ions, Si, Al, Fe, Mn and Cu. Increase of DNA damage in fish also correlated with content of total phosphorus.

Goldfish (Carassius auratus) vitellogenin (Vtg) is an efficient biomarker for estrogen contamination in aquatic environments. In this study, Vtg and lipovitellin (Lv) were purified from the plasma of 17β-estradiol (E2)-induced male goldfish and unfertilized eggs of females, and were used to generate polyclonal antibodies against Vtg (anti-Vtg) and Lv (anti-Lv), respectively. SDS-PAGE and Western blot were performed to confirm the specificity of the two antibodies and the immunological similarity between Vtg and Lv. As anti-Lv recognized more antigen epitopes than anti-Vtg, it was used to develop a sandwich enzyme-linked immunosorbent assay (ELISA) for goldfish Vtg with purified Lv as the standard. The detection limit of the assay was 1.82ng/mL, and the working range was 3.9-250ng/mL. The use of Lv instead of Vtg as the standard provided greater precision and strengthened the robustness of the sandwich ELISA. Western blot and the Lv-based ELISA were used to detect Vtg inductions in surface mucus and plasma of E2-induced goldfish. The surface mucus Vtg level in E2-induced males was significantly higher than that in the control males and E2-induced females, and was much closer to the plasma Vtg level in E2-induced males than that in E2-induced females. Therefore, the surface mucus Vtg level of male goldfish may be a reliable indicator of estrogenic activity in the aquatic environment., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Goldfish (Carassius auratus) represents a good model to detect the estrogenic effects of chemicals, and vitellogenin (Vtg) is a vital indicator of estrogenic activity. The heterologous anti-carp Vtg antibody has previously been used for goldfish Vtg detection. Here, we report the preparation of an anti-goldfish Vtg antibody to improve the sensitivity and specificity of goldfish Vtg immunoassays. Vtg was purified from the plasma of 17β-estradiol (E2)-induced goldfish by gel filtration followed by anion-exchange chromatography. It was characterized as a phospholipoglycoprotein with an apparent molecular weight of ~460 kDa and separated into three major polypeptides corresponding to ~130, ~106, and ~81 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). A polyclonal antibody against goldfish Vtg was raised in rabbits and found to be specific for goldfish Vtg through immunoelectrophoresis and Western blot. A sensitive sandwich enzyme-linked immunosorbent assay (ELISA) was developed for the quantification of plasma Vtg, with a detection limit of 3.6 ng/mL and a detection range from 7.8 to 250 ng/mL. The intra- and inter-assay coefficients of variations were 2.4-6.8% and 6.7-10.8%, respectively. Additionally, we qualitatively and quantitatively detected the induction of Vtg in male fish exposed to 0.01, 0.01, and 1.00 mg/L monocrotophos pesticide by Western blot and ELISA. The homologous sandwich ELISA based on the anti-goldfish Vtg antibody could provide a valuable tool for the study of estrogenic effects of exogenous chemicals on goldfish., (Copyright © 2014 Elsevier Inc. All rights reserved.)

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.

To understand how gonadotropin-inhibitory hormone (GnIH) regulates goldfish GH cell functions, we monitored GH release and expression during early, mid-, and/or late gonadal recrudescence. In vivo and in vitro responses to goldfish (g) GnIH were different, indicating direct action at the level of pituitary, as well as interactions with other neuroendocrine factors involved in GH regulation. Injection of gGnIH consistently reduced basal serum GH levels but elevated pituitary gh mRNA levels, indicating potential dissociation of GH release and synthesis. Goldfish GnRH (sGnRH and cGnRHII) injection differentially stimulated serum GH and pituitary gh mRNA levels with some seasonal differences; these responses were reduced by gGnIH. In contrast, in vitro application of gGnIH during 24-h static incubation of goldfish pituitary cells generally elevated basal GH release and attenuated sGnRH-induced changes in gh mRNA, while suppressing basal gh mRNA levels at mid- and late recrudescence but elevating them at early recrudescence. gGnIH attenuated the GH release responses to sGnRH during static incubation at early, but not at mid- and late recrudescence. In cell column perifusion experiments examining short-term GH release, gGnIH reduced the cGnRHII- and sGnRH-stimulated secretion at late recrudescence but inhibited tha action of cGnRHII only during mid-recrudescence. Interestingly, a reduction of basal GH release upon perifusion with gGnIH during late recrudescence was followed by a rebound increase in GH release upon gGnIH removal. These results indicate that gGnIH exerts complex effects on basal and GnRH-stimulated goldfish GH cell functions and can differentially affect GH release and mRNA expression in a seasonal reproductive manner., (© 2014 Society for Endocrinology.)

This study aims to assess effects of 96 h goldfish exposure to 1, 10 and 100 mg/L of the herbicide, 2,4-dichlorophenoxyacetic acid (2,4-D), on metabolic indices and free radical process markers in white muscle of a commercial fish, the goldfish Carassius auratus L. Most oxidative stress markers and antioxidant enzymes were not affected at 2,4-D fish treatment. 2,4-D fish exposure induced the elevated levels of total (by 46% and 40%) and reduced (by 77% and 73%) glutathione in muscles of goldfish of 10 mg/L 2,4-D and recovery (after 100 mg/L of 2,4-D exposure) groups, respectively. However, in muscles of 100 mg/L 2,4-D exposed goldfish these parameters were depleted (by 47% and 64%). None of investigated parameters of protein and carbohydrate metabolisms changed in white muscles of 2,4-D exposed fish, with exception of lactate dehydrogenase activity, which was slightly (by 11-15%) elevated in muscles of goldfish exposed to 10-100 mg/L of 2,4-D, but also recovered. Thus, the short term exposure of goldfish to the selected concentrations of 2,4-D does not substantially affect their white muscle, suggesting the absence of any effect under the environmentally relevant concentrations., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

We investigated relationships among branchial unidirectional Na(+) fluxes, ammonia excretion, urea excretion, plasma ammonia, plasma cortisol, and gill transporter expression and function in three freshwater fish differing in their sensitivity to high environmental ammonia (HEA). The highly ammonia-sensitive salmonid Oncorhynchus mykiss (rainbow trout), the less ammonia-sensitive cyprinid Cyprinus carpio (common carp) and the highly ammonia-resistant cyprinid Carassius auratus (goldfish) were exposed chronically (12-168 h) to 1 mmol l(-1) ammonia (as NH4HCO3; pH 7.9). During HEA exposure, carp and goldfish elevated ammonia excretion (JAmm) and Na(+) influx rates ( ) while trout experienced higher plasma ammonia (TAmm) and were only able to restore control rates of JAmm and . All three species exhibited increases in Na(+) efflux rate ( ). At the molecular level, there was evidence for activation of a 'Na(+)/NH4(+) exchange metabolon' probably in response to elevated plasma cortisol and TAmm, though surprisingly, some compensatory responses preceded molecular responses in all three species. Expression of Rhbg, Rhcg (Rhcg-a and Rhcg-b), H(+)-ATPase (V-type, B-subunit) and Na(+)/K(+)-ATPase (NKA) mRNA was upregulated in goldfish, Rhcg-a and NKA in carp, and Rhcg2, NHE-2 (Na(+)/H(+) exchanger) and H(+)-ATPase in trout. Branchial H(+)-ATPase activity was elevated in goldfish and trout, and NKA activity in goldfish and carp, but NKA did not appear to function preferentially as a Na(+)/NH4(+)-ATPase in any species. Goldfish alone increased urea excretion rate during HEA, in concert with elevated urea transporter mRNA expression in gills. Overall, goldfish showed more effective compensatory responses towards HEA than carp, while trout were least effective.

Aloes are now considered a very interesting source of bioactive compounds among which phytosterols should play a major role. The present study is an attempt to investigate the hypocholesterolemic activity of Aloe vera associated with its impact on the reproductive status of juvenile goldfish. Therefore, the short- and long-term effects of feeding supplementary diet containing aloe components (20 mg aloe/g diet; 2%) on plasma lipids, plasma vitellogenin, and hepatic estrogen receptor α/β1 mRNA levels in goldfish were examined. Results of GC-MS for phytosterols show high abundance of β-sitosterol in freeze-dried powder of Aloe vera whole leaves. Moreover, a 2% aloe powder dietary supplement was not found estrogenic in juvenile goldfish after either 7- or 30-day treatment, but was consistent in plasma hypocholesterolemic effects following long-term exposure. The present data further support that plasma cholesterol modulation induced by phytosterols may not be related to estrogen-like activity.

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.

A long-term feeding experiment was conducted to investigate the inclusion of soybean meal in diets for goldfish (Carassius auratus) on fish reproduction. In the present study, 20 weeks after hatching, goldfish with an initial average weight of 2±0.03g (mean±SD) were divided into 12 groups (three tanks per dietary treatment) and fed 400gkg(-1) crude protein diets. The four experimental diets were as follows: diet 1, fish meal (FM); diet 2, 35% soybean meal (SBM35%); diet 3, 65% soybean meal (SBM65%); diet 4, 100% soybean meal (SBM100%). After feeding with experimental diets, the impact on reproduction was investigated. In both males and females, the plasma testosterone (T) was significantly decreased, while 17β-estradiol (E2) levels were significantly increased. Levels of 17α, hydroxyprogesterone. (17-OH-P) did not differ as a result of soybean meal feeding in either males or females. The average number of eggs spawned and sperm quality were reduced on feeding with soybean inclusion. Histological examination showed impact on oocyte maturation progress and spermatogenesis process in female and male fish, respectively. In addition, feeding goldfish with soybean meal until maturation caused reduction in fertilization and hatching rates in parallel to increasing soybean meal inclusion. The results demonstrated that inclusion of soybean meal might cause sex hormone biosynthesis disruption and reproductive impairments in fish, ultimately decreased fertilization as well as hatching rates in the offspring., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Feeding and swimming can influence ion balance in fish. Therefore we investigated their impact on ionoregulation and its hormonal control in goldfish and common carp. As expected due to the osmorespiratory compromise, exhaustive swimming induced increases in gill Na(+)/K(+) ATPase (NKA) activity in both species, resulting in stable levels of plasma ions. In contrast to our expectations, this only occurred in fed fish and feeding itself increased NKA activity, especially in carp. Fasting fish were able to maintain ion balance without increasing NKA activity, we propose that the increase in NKA activity is related to ammonia excretion rather than ion uptake per se. In goldfish, this increase in NKA activity coincided with a cortisol elevation whilst no significant change was found in carp. In goldfish, high conversion of plasma T4 to T3 was found in both fed and fasted fish resulting in low T4/T3 ratios, which increased slightly due to exhaustive swimming. In starved carp the conversion seemed much less efficient, and high T4/T3 ratios were observed. We propose that thyroid hormone regulation in carp was more related to its role in energy metabolism rather than ionoregulation. The present research showed that both species, whether fed or fasted, are able to sufficiently adapt their osmorepiratory strategy to minimise ions losses whilst maintaining gas exchange under exhaustive swimming., (Copyright © 2013 Elsevier Inc. All rights reserved.)

This study investigated the effects of 2,4-dichlorophenoxyacetic acid (2,4-D), a widely used herbicide, on the metabolism of goldfish, Carassius auratus, using only vital (non-lethal) approaches. After 96 h exposure to 1, 10 or 100 mg/L of 2,4-D selected hematological (total hemoglobin and hematocrit) and biochemical (glucose content, aspartate transaminase and acetylcholinesterase activities) parameters were unchanged in blood of exposed fish. At 100 mg/L of 2,4-D lymphocyte numbers decreased by 8%, whereas promyelocyte and metamyelocyte numbers increased by 7- and 2-fold, respectively. Exposure to 100 mg/L of 2,4-D also elevated carbonyl protein levels (by 2-fold), triglyceride content (by 43%) and alanine transaminase activity (by 46%) in goldfish plasma. All of these hematological and biochemical parameters reverted to control values after a 96 h recovery period. These data indicate that 2,4-D has toxicological effects on goldfish that can be monitored with multiple diagnostic tests using non-lethal blood testing., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Intensive use of pesticides, particularly dithiocarbamates, in agriculture often leads to contamination of freshwater ecosystems. To our knowledge, the mechanisms of toxicity to fish by the carbamate fungicide Tattoo that contains mancozeb [ethylenebis(dithiocarbamate)] have not been studied. The present study aimed to evaluate the effects of Tattoo on goldfish gills and blood, tissues that would have close early contact with the pollutant. Exposure of goldfish Carassius auratus to 3, 5 or 10mgL(-1) of Tattoo for 96h resulted in moderate lymphopenia (by 8 percent) with a concomitant increase in both stab (by 66-88 percent) and segmented (by 166 percent) neutrophils. An increase in the content of protein carbonyl groups in blood (by 137-184 percent) together with decreased levels of protein thiols (by 23 percent) and an enhancement of lipid peroxide concentrations (by 29 percent) in gills after exposure to 10mgL(-1) of Tattoo demonstrated the induction of mild oxidative stress in response to Tattoo exposure. At the same time, the activities of selected antioxidant enzymes were enhanced in gills: superoxide dismutase by 18-25 percent and catalase by 27 percent. A 34 percent increment in low molecular mass thiol concentrations (mainly represented by glutathione) also occurred in gills and could be related to increased activity (by 13-30 percent) of glucose-6-phosphate dehydrogenase. The results indicate that Tattoo exposure perturbs free radical processes, i.e. induces mild oxidative stress and enhances the activity of certain antioxidant and associated enzymes in goldfish gills. It is clear that goldfish respond to the presence of waterborne pesticide by adjusting antioxidant defenses through upregulation of activities of antioxidant and associated enzymes., (Copyright © 2012 Elsevier Inc. All rights reserved.)

The present study investigated the effects of triiodothyronine (T3) on pituitary gonadotropin (GTH) subunits, thyroid stimulating hormone (TSH) β subunit, and growth hormone (GH) mRNA levels, as well as gonadal steroid secretion during different stages of reproduction in goldfish. Goldfish pituitary cells cultured with T3 exhibited lower tshβ mRNA levels in all reproductive stages and lower luteinising hormone β (lhβ) mRNA levels in early recrudescence, whereas gh and fshβ mRNA levels were not altered. T3 injections significantly reduced circulating oestrogen (OE2) concentrations in early and mid recrudescent male goldfish, but were without effect on the circulating level of OE2 in female fish. T3 injections also reduced circulating levels of testosterone in both male and female goldfish during the mid stage of gonadal recrudescence. In vitro culture of goldfish ovarian follicles at the late stage of gonadal recrudescence, in the presence of T3, resulted in reduced OE2 secretion; no consistent effect of T3 on testosterone secretion was observed in cultured goldfish ovarian follicles and testis. These findings support the hypothesis that T3 impairs reproduction by inhibiting production of gonadal steroids and pituitary luteinising hormone production in goldfish., (Copyright © 2012 Wiley Periodicals, Inc.)

The pharmacokinetics of enrofloxacin (EF) was investigated after single intravenous (i.v.) and oral (p.o.) administration of 10 mg/kg body weight (b.w.) in 300 healthy allogynogenetic silver crucian carp at 24-26°C. The plasma concentrations of EF and its metabolite ciprofloxacin (CF) were determined by high-performance liquid chromatography. After i.v. administration, the plasma concentration-time data were described by an open two-compartment model. The elimination half-life (T(1/2β)), area under the concentration-time curve (AUC) and total body clearance of EF were 63.5 h, 239.6 μg·h/mL and 0.04 L/h/kg, respectively. Following p.o. administration, the plasma concentration-time data showed a double peak-shaped curve, indicating the possibility of enterohepatic recirculation of EF in allogynogenetic silver crucian carp. The maximum plasma concentration (C(max)), T(1/2β) and AUC of EF were 4.5 μg/mL, 62.7 h and 205.9 μg·h/mL, respectively. Absorption of EF was very good with a bioavailability (F) of 86%, which could be correlated with the unique structure of the alimentary canal in allogynogenetic silver crucian. CF, an active metabolite of EF, was not detected in this study., (© 2011 Blackwell Publishing Ltd.)

Fish suffer from anemia and hypovolemic hypotensive shock after in vivo exposure with microcystins.However, except for in vivo causes for anemia and hypotension, an in vitro study of fish erythrocytes exposed to MC is necessary. For a better understanding of hematology toxicity of MC, the main aim of the present study was to investigate the toxic effects of microcystin on fish erythrocytes in vitro. Crucian carp erythrocytes were incubated in vitro with microcystin-LR (MC-LR) at doses of 0, 1, 10, 100 and 1,000 nM.The level of lipid peroxidate significantly increased in MC-LR treatment groups. Glutathione decreased after exposure to MC-LR. The activities of antioxidative enzymes, including superoxide dismutase, catalase,glutathione peroxidase and glutathione-S-transferase,were significantly increased after exposure with MC-LR.The hemolysis was significantly increased, while the activities of acetylcholinesterase, Na?–K?-ATPase and Ca2?–Mg2?-ATPase were significantly decreased. In addition, pathological alterations in agglomerated and jagged erythrocytes were observed in blood smears. The findings indicate that damages to erythrocytes should also be responsible for anemia and hypotensive shock or even death.

Thyrotropin (TSH) is a pituitary glycoprotein hormone heterodimer that binds to its G-protein coupled receptor (TSH-R) at the thyroid to promote the synthesis and secretion of thyroid hormone. Very little is known about TSH-TSH-R interactions in teleost fish. Mammalian gonadotropins have been reported to have an intrinsic ability to activate teleost fish TSH-Rs, suggesting the TSH-R in teleost fish is more promiscuous than in other vertebrates. In this study we utilized the goldfish T(4)-release response and recombinant human TSH analogs as in vivo tools to evaluate the structural constraints on hormone-receptor interactions. We found that four positively charged lysines substituted for neutral or negatively charged amino acids within positions 11-20 of the glycoprotein hormone subunit α (GSUα) significantly increased biological activity of hTSH in fish, as it does in mammals. We further found that bovine follicle stimulating hormone but not luteinizing hormone, whose GSUα subunits also contain four lysine or arginine amino acid residues in the N-terminal portion of GSUα, was thyrotropic in goldfish, suggesting gonadotropin β subunit contributes to the heterothyrotropic activity. Though recombinant human FSH did not produce a dose-dependent increase in T(4), thyrotropic activity could be acquired with the addition of positively charged amino acids at the N-terminal portion of its GSUα, confirming the importance of the charge on those amino acids for activation of the goldfish TSH-R. These studies demonstrate that mammalian glycoprotein hormone analogs can be utilized to evaluate the conservation of receptor binding and activation mechanisms between fish and mammals., (Published by Elsevier Inc.)

The composition of leucocytes of the head kidney is studied in the crucian carps (Carassius auratus) either contaminated or uncontaminated with Digramma interrupta. The composition of leucocytes in the pronephros of the crucian carp from Lake Baikal basin has a lymphoid character. Compared to the crucian carp from the European part of Russia, in the fish from Baikal the granulocytopoetic processes are more pronounced. This is proved by the high content of young forms of granulocytes. In the fish infected with digramma, the immune suppression of proliferation of blasts and young forms of eosinophils was revealed. On the other hand, the inflammatory and humoral specific immune reactions are enhanced. Partial suppression of the immune response of C. auratus to invasion by D. interrupta facilitates development of the parasite.

The pharmacokinetics of florfenicol (FF) was studied in plasma after a single dose (40 mg/kg) of intramuscular (i.m.) or oral gavage (p.o.) administration to crucian carp (Carassius auratus cuvieri) in freshwater at 25 °C. Ten fish per sampling point were examined after treatment. The data were fitted to two-compartment open models follow both routes of administration. The estimates of total body clearance (CL(b) ), volume of distribution (V(d) /F), and absorption half-life (T(1/2(ka)) ) were 0.067 L/h/kg and 0.145 L/h/kg, 2.21 L/kg and 1.04 L/kg, 2.75 and 1.54/h following i.m. and p.o. administration, respectively. After i.m. injection, the elimination half-life (T(1/2(β)) ) was calculated to be 38.2h, the maximum plasma concentration (C(max) ) to be 16.82 μg/mL, the time to peak plasma FF concentration (T(max) ) to be 1.50 h, and the area under the plasma concentration-time curve (AUC) to be 597.4 μg/mL·h. Following p.o. administration, the corresponding estimates were 2.17 h, 29.32 μg/mL, 1.61 h, and 276.1 μg/mL·h., (© 2011 Blackwell Publishing Ltd.)