Androgen receptors (ARs) mediate the physiological effects of androgens in vertebrates. In fishes... more Androgen receptors (ARs) mediate the physiological effects of androgens in vertebrates. In fishes, AR-mediated pathways can be modulated by aquatic contaminants, resulting in the masculinisation of female fish or diminished secondary sex characteristics in males. The Murray–Darling rainbowfish (Mel-anotaenia fluviatilis) is a small-bodied freshwater teleost used in Australia as a test species for environmental toxicology research. We determined concentration–response profiles for selected agonists and antagonists of rainbowfish ARa and ARb using transient transactivation assays. For both ARa and ARb, the order of potency of natural agonists was 11-ketotestosterone (11-KT) > 5a-dihydrotestosterone > tes-tosterone > androstenedione. Methyltestosterone was a highly potent agonist of both receptors relative to 11-KT. The relative potency of the veterinary growth-promoting androgen, 17b-trenbolone, varied by more than a factor of 5 between ARa and ARb. The non-steroidal anti-androgen bicalutamide exhibited high inhibitory potency relative to the structurally related model anti-androgen, flutamide. The inhibitory potency of the agricultural fungicide, vinclozolin, was approximately 1.7-fold relative to flu-tamide for ARa, but over 20-fold in the case of ARb. Fluorescent protein tagging of ARs showed that the rainbowfish ARa subtype is constitutively localised to the nucleus, while ARb is cytoplasmic in the absence of ligand, an observation which agrees with the reported subcellular localisation of AR subtypes from other teleost species. Collectively, these data suggest that M. fluviatilis ARa and ARb respond differently to environmental AR modulators and that in vivo sensitivity to contaminants may depend on the tissue distribution of the AR subtypes at the time of exposure.
The aim of the present study was to investigate the lethal and sublethal effects of prednisolone ... more The aim of the present study was to investigate the lethal and sublethal effects of prednisolone exposure on the embryonic and posthatching stage of the freshwater snail, Physa acuta. The egg masses were exposed for 14 d to prednisolone concentrations ranging from 15.6 mg/L to 1000 mg/L. Treatment with prednisolone at 125 mg/L to 1000 mg/L resulted in significant decline in growth, survival, and heart rate, as well as notable abnormalities in embryonic development. Premature embryonic hatching was observed at lower concentrations of 31.25 mg/L and 62.5 mg/L, whereas delayed hatching was seen at concentrations from 125 mg/L to 1000 mg/L. To assess impacts of prednisolone exposure on the hatched juveniles, the drug exposure was extended for another 28 d. Impairment of shell development was noted in juveniles exposed to concentrations from 62.5 mg/L to 1000 mg/L at the end of 42 d, which resulted in thin and fragile shells. The thickness of shells in snails exposed to 1000 mg/L was significantly lower in comparison to those in the 15.6-mg/L and control treatments. In addition, lower calcium concentration in shells of the exposed juvenile snails at treatments of 62.5 mg/L to 1000 mg/L consequently reduced their growth. The present study confirms that continuous exposure to prednisolone can result in deleterious effects on calcium deposition, resulting in shell thinning in the freshwater snail P. acuta. Environ Toxicol Chem 2016;35:2339–2348. # 2016 SETAC
The aim of the present study was to investigate if the effects of the androgen, dihydrotestostero... more The aim of the present study was to investigate if the effects of the androgen, dihydrotestosterone (DHT) on the sexual development in juvenile Murray rainbowfish (Melanotaenia fluviatilis) are canceled out by the anti-androgen, flutamide. Fish (60 days post hatch) were exposed to 250 ng/L of DHT, 25 g/L of flutamide (Flu-low), 250 g/L of flutamide (Flu-high), DHT + Flu low and DHT + Flu high. After 35 days of exposure, lengths and weights of the fish were measured and the condition factor (CF) calculated; vitellogenin (VTG) concentrations were measured in tail tissue; sex steroid hormones (17-estradiol [E2] and 11-keto testosterone [11-KT]) were measured in the head tissue and abdominal regions were used in histological investigation of the gonads. Treatment with DHT reduced the body-length of both male and female fish, an effect which was canceled out by low and high concentrations of flutamide. However, flutamide (low or high) could not nullify the DHT-induced reduction in the CF in either sex. The E2 levels were reduced only in female fish after exposure to DHT but returned to normal after treatment with Flu-high. DHT increased the levels of 11-KT and decreased the E2/11-KT ratio in both sexes. Flu-high, but not Flu-low, could nullify these effects. Both DHT and flutamide (low or high) induced VTG production and this effect persisted when both chemicals were co-administered. Treatment with DHT did not affect gonadal cell development in the testes. However, the female fish treated with DHT contained ovaries in early-vitellogenic stage in comparison to the pre-vitellogenic ovaries in control fish. Co-treatment with flutamide (low or high) resulted in oocyte atresia. The results from the present study suggest that treatment with Flu-high could cancel out DHT-induced effects only on the hormonal profile and body-length in both male and female fish. Juvenile fish co-treated with DHT and flutamide (low or high) had high VTG levels and low CF. In addition, the ovaries in female fish were atretic. These data represent potential adverse effects on the ability of the fish to reproduce successfully.
Pharmaceuticals can enter freshwater and affect aquatic ecosystem health. Although toxicity tests... more Pharmaceuticals can enter freshwater and affect aquatic ecosystem health. Although toxicity tests have been carried out for the commonly used pharmaceuticals, evidence-based water quality guidelines have not been derived. High-reliability water quality guideline values have been derived for 4 pharmaceuticals—carbamazepine, diclofenac, fluoxetine, and propranolol—in freshwaters using a Burr type III distribution applied to species sensitivity distributions of chronic toxicity data. Data were quality-assured and had to meet acceptability criteria for " chronic " no-observed-effect concentrations or concentrations affecting 10% of species, endpoints of population relevance (namely, effect endpoints based on development, growth, reproduction, and survival). Biomarker response data (e.g., biochemical, histological, or molecular responses) were excluded from the derivation because they are typically not directly relevant to wildlife population-related impacts. The derived guideline values for 95% species protection were 9.2 mg/L, 770 mg/L, 1.6 mg/L, and 14 mg/L for carbamazepine, diclofenac, fluoxetine, and propranolol, respectively. These values are significantly higher than the unknown reliability values derived for the European Commission, Switzerland, or Germany that are based on the application of assessment factors to the most sensitive experimental endpoint (which may include biochemical, histological, or molecular biomarker responses) of a limited data set. The guideline values derived in the present study were not exceeded in recent data for Australian rivers and streams receiving pharmaceutical-containing effluents from wastewater-treatment plants.
Androgen receptors (ARs) mediate the physiological effects of androgens in vertebrates. In fishes... more Androgen receptors (ARs) mediate the physiological effects of androgens in vertebrates. In fishes, AR-mediated pathways can be modulated by aquatic contaminants, resulting in the masculinisation of female fish or diminished secondary sex characteristics in males. The Murray–Darling rainbowfish (Mel-anotaenia fluviatilis) is a small-bodied freshwater teleost used in Australia as a test species for environmental toxicology research. We determined concentration–response profiles for selected agonists and antagonists of rainbowfish ARa and ARb using transient transactivation assays. For both ARa and ARb, the order of potency of natural agonists was 11-ketotestosterone (11-KT) > 5a-dihydrotestosterone > tes-tosterone > androstenedione. Methyltestosterone was a highly potent agonist of both receptors relative to 11-KT. The relative potency of the veterinary growth-promoting androgen, 17b-trenbolone, varied by more than a factor of 5 between ARa and ARb. The non-steroidal anti-androgen bicalutamide exhibited high inhibitory potency relative to the structurally related model anti-androgen, flutamide. The inhibitory potency of the agricultural fungicide, vinclozolin, was approximately 1.7-fold relative to flu-tamide for ARa, but over 20-fold in the case of ARb. Fluorescent protein tagging of ARs showed that the rainbowfish ARa subtype is constitutively localised to the nucleus, while ARb is cytoplasmic in the absence of ligand, an observation which agrees with the reported subcellular localisation of AR subtypes from other teleost species. Collectively, these data suggest that M. fluviatilis ARa and ARb respond differently to environmental AR modulators and that in vivo sensitivity to contaminants may depend on the tissue distribution of the AR subtypes at the time of exposure.
The aim of the present study was to investigate the lethal and sublethal effects of prednisolone ... more The aim of the present study was to investigate the lethal and sublethal effects of prednisolone exposure on the embryonic and posthatching stage of the freshwater snail, Physa acuta. The egg masses were exposed for 14 d to prednisolone concentrations ranging from 15.6 mg/L to 1000 mg/L. Treatment with prednisolone at 125 mg/L to 1000 mg/L resulted in significant decline in growth, survival, and heart rate, as well as notable abnormalities in embryonic development. Premature embryonic hatching was observed at lower concentrations of 31.25 mg/L and 62.5 mg/L, whereas delayed hatching was seen at concentrations from 125 mg/L to 1000 mg/L. To assess impacts of prednisolone exposure on the hatched juveniles, the drug exposure was extended for another 28 d. Impairment of shell development was noted in juveniles exposed to concentrations from 62.5 mg/L to 1000 mg/L at the end of 42 d, which resulted in thin and fragile shells. The thickness of shells in snails exposed to 1000 mg/L was significantly lower in comparison to those in the 15.6-mg/L and control treatments. In addition, lower calcium concentration in shells of the exposed juvenile snails at treatments of 62.5 mg/L to 1000 mg/L consequently reduced their growth. The present study confirms that continuous exposure to prednisolone can result in deleterious effects on calcium deposition, resulting in shell thinning in the freshwater snail P. acuta. Environ Toxicol Chem 2016;35:2339–2348. # 2016 SETAC
The aim of the present study was to investigate if the effects of the androgen, dihydrotestostero... more The aim of the present study was to investigate if the effects of the androgen, dihydrotestosterone (DHT) on the sexual development in juvenile Murray rainbowfish (Melanotaenia fluviatilis) are canceled out by the anti-androgen, flutamide. Fish (60 days post hatch) were exposed to 250 ng/L of DHT, 25 g/L of flutamide (Flu-low), 250 g/L of flutamide (Flu-high), DHT + Flu low and DHT + Flu high. After 35 days of exposure, lengths and weights of the fish were measured and the condition factor (CF) calculated; vitellogenin (VTG) concentrations were measured in tail tissue; sex steroid hormones (17-estradiol [E2] and 11-keto testosterone [11-KT]) were measured in the head tissue and abdominal regions were used in histological investigation of the gonads. Treatment with DHT reduced the body-length of both male and female fish, an effect which was canceled out by low and high concentrations of flutamide. However, flutamide (low or high) could not nullify the DHT-induced reduction in the CF in either sex. The E2 levels were reduced only in female fish after exposure to DHT but returned to normal after treatment with Flu-high. DHT increased the levels of 11-KT and decreased the E2/11-KT ratio in both sexes. Flu-high, but not Flu-low, could nullify these effects. Both DHT and flutamide (low or high) induced VTG production and this effect persisted when both chemicals were co-administered. Treatment with DHT did not affect gonadal cell development in the testes. However, the female fish treated with DHT contained ovaries in early-vitellogenic stage in comparison to the pre-vitellogenic ovaries in control fish. Co-treatment with flutamide (low or high) resulted in oocyte atresia. The results from the present study suggest that treatment with Flu-high could cancel out DHT-induced effects only on the hormonal profile and body-length in both male and female fish. Juvenile fish co-treated with DHT and flutamide (low or high) had high VTG levels and low CF. In addition, the ovaries in female fish were atretic. These data represent potential adverse effects on the ability of the fish to reproduce successfully.
Pharmaceuticals can enter freshwater and affect aquatic ecosystem health. Although toxicity tests... more Pharmaceuticals can enter freshwater and affect aquatic ecosystem health. Although toxicity tests have been carried out for the commonly used pharmaceuticals, evidence-based water quality guidelines have not been derived. High-reliability water quality guideline values have been derived for 4 pharmaceuticals—carbamazepine, diclofenac, fluoxetine, and propranolol—in freshwaters using a Burr type III distribution applied to species sensitivity distributions of chronic toxicity data. Data were quality-assured and had to meet acceptability criteria for " chronic " no-observed-effect concentrations or concentrations affecting 10% of species, endpoints of population relevance (namely, effect endpoints based on development, growth, reproduction, and survival). Biomarker response data (e.g., biochemical, histological, or molecular responses) were excluded from the derivation because they are typically not directly relevant to wildlife population-related impacts. The derived guideline values for 95% species protection were 9.2 mg/L, 770 mg/L, 1.6 mg/L, and 14 mg/L for carbamazepine, diclofenac, fluoxetine, and propranolol, respectively. These values are significantly higher than the unknown reliability values derived for the European Commission, Switzerland, or Germany that are based on the application of assessment factors to the most sensitive experimental endpoint (which may include biochemical, histological, or molecular biomarker responses) of a limited data set. The guideline values derived in the present study were not exceeded in recent data for Australian rivers and streams receiving pharmaceutical-containing effluents from wastewater-treatment plants.
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