Propiconazole inhibits steroidogenesis and reproduction in the fathead minnow (Pimephales promelas).

Conazoles are designed to inhibit cytochrome P450 (CYP) 14α-demethylase, an enzyme key to fungal cell wall formation. In vertebrates, conazoles may inhibit other CYPs, potentially disrupting processes like sex steroid synthesis. Propiconazole is a current-use pesticide that is among the first chemicals being tested in the U.S. Environmental Protection Agency endocrine disruptor screening program. Fathead minnows (Pimephales promelas) were exposed to 0, 5, 50, 500, or 1000 µg propiconazole/l in a 21-day study that evaluated apical reproductive endpoints (fecundity, fertility, hatch); measures of endocrine function and steroid synthesis, such as cholesterol, vitellogenin (VTG), and sex steroid (testosterone [T], 17ß-estradiol [E2]) concentrations in the plasma; and changes in gonadal expression of steroidogenic genes. Plasma E2 and VTG concentrations in females were reduced by exposure to propiconazole, and egg production was decreased in the 500 and 1000 µg/l treatment groups. These in vivo effects coincided with inhibition of E2 synthesis by ovary explants exposed to propiconazole in vitro. We also observed a compensatory response in females exposed to propiconazole, manifested as increased gonad weight and upregulation of genes coding for key steriodogenic proteins, including CYP19 (aromatase), CYP17 (hydroxylase/lyase), CYP11A (cholesterol side-chain-cleavage), and steroidogenic acute regulatory protein. Other than an increase in relative testis weight, effects on endocrine function in males were less pronounced than in females. This study provides important data relative to the potential endocrine activity of propiconazole in fish and, more generally, to the further delineation of pathways for the reproductive effects of steroid synthesis inhibitors in fish.

Effects of gemfibrozil on lipid metabolism, steroidogenesis, and reproduction in the fathead minnow (Pimephales promelas).

Fibrates are a class of pharmaceuticals that indirectly modulate cholesterol biosynthesis through effects on peroxisome proliferator-activated receptors. Gemfibrozil is a fibrate that has been detected in wastewater treatment plant influents, effluents, and drinking water. The objective of the present study was to assess the potential physiological and reproductive impacts of gemfibrozil on fathead minnows (Pimephales promelas). Fish were exposed to gemfibrozil in two different studies. The first was a short-term test with water concentrations of 0, 15, and 600 µg gemfibrozil/L, sampling after 2 or 8 d of exposure. Plasma cholesterol concentrations were significantly reduced in males exposed to 600 µg gemfibrozil/L for 8 d. In addition, expression of several hepatic genes important to lipid metabolism was altered, suggesting that gemfibrozil does affect lipid metabolism in fish. A 21-d study was conducted to investigate further the effects on lipid metabolism and steroidogenesis as well as to assess potential impacts of gemfibrozil on reproduction. Fish were exposed to water concentrations of 0, 1.5, 15, 600, and 1,500 µg gemfibrozil/L. Exposure to 1,500 µg gemfibrozil/L caused a modest, but not significant, reduction in fecundity. However, gemfibrozil had no consistent effect on plasma cholesterol, triglycerides, or sex steroids after 21 d of exposure. The present study showed no evidence for significant physiological or reproductive impacts of gemfibrozil at an environmentally relevant concentration of 1.5 µg/L.

Effects of a glucocorticoid receptor agonist, dexamethasone, on fathead minnow reproduction, growth, and development.

Synthetic glucocorticoids are pharmaceutical compounds prescribed in human and veterinary medicine as anti-inflammatory agents and have the potential to contaminate natural watersheds via inputs from wastewater treatment facilities and confined animal-feeding operations. Despite this, few studies have examined the effects of this class of chemicals on aquatic vertebrates. To generate data to assess potential risk to the aquatic environment, we used fathead minnow 21-d reproduction and 29-d embryo-larvae assays to determine reproductive toxicity and early-life-stage effects of dexamethasone. Exposure to 500 µg dexamethasone/L in the 21-d test caused reductions in fathead minnow fecundity and female plasma estradiol concentrations and increased the occurrence of abnormally hatched fry. Female fish exposed to 500 µg dexamethasone/L also displayed a significant increase in plasma vitellogenin protein levels, possibly because of decreased spawning. A decrease in vitellogenin messenger ribonucleic acid (mRNA) expression in liver tissue from females exposed to the high dexamethasone concentration lends support to this hypothesis. Histological results indicate that a 29-d embryo-larval exposure to 500 µg dexamethasone/L caused a significant increase in deformed gill opercula. Fry exposed to 500 µg dexamethasone/L for 29 d also exhibited a significant reduction in weight and length compared with control fry. Taken together, these results indicate that nonlethal concentrations of a model glucocorticoid receptor agonist can impair fish reproduction, growth, and development.

Effects of a short-term exposure to the fungicide prochloraz on endocrine function and gene expression in female fathead minnows (Pimephales promelas).

Prochloraz is a fungicide known to cause endocrine disruption through effects on the hypothalamic-pituitary-gonadal (HPG) axis. To determine the short-term impacts of prochloraz on gene expression and steroid production, adult female fathead minnows (Pimephales promelas) were exposed to the chemical (0 or 300 µg/L) for a time-course of 6, 12 and 24 h. Consistent with inhibition of cytochrome P450 17α-hydroxylase/17,20-lyase (CYP17) and aromatase (CYP19), known molecular targets of prochloraz, plasma 17ß-estradiol (E2) was reduced within 6 h. Ex vivo E2 production was significantly reduced at all time-points, while ex vivo testosterone (T) production remained unchanged. Consistent with the decrease in E2 levels, plasma concentrations of the estrogen-responsive protein vitellogenin were significantly reduced at 24 h. Genes coding for CYP19, CYP17, and steroidogenic acute regulatory protein were up-regulated in a compensatory manner in ovaries of the prochloraz-treated fish. In addition to targeted quantitative real-time polymerase chain reaction analyses, a 15k feature fathead minnow microarray was used to determine gene expression profiles in ovaries. From time-point to time-point, the microarray results showed a relatively rapid change in the differentially expressed gene (DEG) profiles associated with the chemical exposure. Functional analysis of the DEGs indicated changes in expression of genes associated with cofactor and coenzyme binding (GO:0048037 and 0050662), fatty acid binding (GO:0005504) and organelle organization and biogenesis (GO:0006996). Overall, the results from this study are consistent with compensation of the fish HPG axis to inhibition of steroidogenesis by prochloraz, and provide further insights into relatively rapid, system-wide, effects of a model chemical stressor on fish.