Research Grants/Fellowships/SBIR

2001 Progress Report: Reproductive and endocrine effects of o,p'-DDT, an environmental estrogen, and p,p'-DDE, an antiandrogen in male and female Atlantic croaker during critical periods of their reproductive life history cycles

EPA Grant Number: R826125
Title: Reproductive and endocrine effects of o,p'-DDT, an environmental estrogen, and p,p'-DDE, an antiandrogen in male and female Atlantic croaker during critical periods of their reproductive life history cycles
Investigators: Thomas, Peter
Institution: University of Texas at Austin
EPA Project Officer: Turner, Vivian
Project Period: October 1, 1997 through September 30, 2000
Project Period Covered by this Report: October 1, 2000 through September 30, 2001
Project Amount: $430,010
RFA: Endocrine Disruptors (1997) RFA Text |  Recipients Lists
Research Category: Endocrine Disruptors , Economics and Decision Sciences , Health , Safer Chemicals



Specific objectives are to determine: (1) which stages of the croaker reproductive life history cycle are disrupted by o,p'-DDT and p,p'-DDE; (2) what endocrine and reproductive effects of o,p'-DDT and p,p'-DDE are related to their estrogenicity and antiandrogenicity, respectively; (3) the sites of o,p'-DDT and p,p'-DDE action on the HPGL axis; (4) the role of the ER in mediating the actions of p,p'-DDE; and (5) whether there are sex differences in the susceptibility to o,p'-DDT and p,p'-DDE.

Progress Summary:

In Vivo Exposure to Antiandrogens

The analysis of in vivo experiments with croakers exposed in the diet to p,p'-DDE (0.01 mg and 0.05 mg/100 g bw/day) and cyproterone acetate (0.1 mg/100 g bw/day) for 14 and 24 weeks during the period of gonadal recrudescence has been completed. It was found that gonadal growth was impaired in both males and females exposed to the highest dose of p,p'-DDE, and this was accompanied by decreases in plasma levels of gonadal steroids but not in the capacity of the gonads to produce steroids in vitro. Possible neuroendocrine sites of p,p'-DDE action were investigated by measurement of plasma luteinizing hormone (LH) levels. Both basal LH secretion and luteinizing hormone-releasing hormone (LHRH)-induced LH secretion were not consistently altered following p,p'-DDE treatment. These results suggest that the decrease in plasma sex steroid levels is not mediated by alterations at higher levels of the hypothalamus-pituitary-gonadal axis. An alternative possibility is that the metabolic clearance rate of steroids is enhanced after p,p'-DDE treatment, as has been observed after organochlorine exposure in mammals. However, this was beyond the scope of the present study.

In Vivo Exposure to Androgens

Nuclear Androgen Receptor

Previously, two nuclear androgen receptor subtypes had been identified in the Atlantic croaker. The regulation of the brain androgen receptor by gonadal steroids was examined in gonadectomized male and female croakers. Gonadectomy caused a decrease in brain androgen receptor levels, which were partially restored by treatment with testosterone and estrogen, but the nonaromatizable androgen, 5"-dihydrotestosterone, was less effective. These results indicate that natural androgens and estrogens can regulate nuclear androgen receptor levels in fishes. The gonadectomized croaker model appears to be useful for investigating the influence of endocrine disrupting chemicals on receptor abundance and hence androgen action.

Androgen Downregulation of Ovarian Steroidogenesis by a Nongenomic Mechanism

Previously, an androgen nuclear receptor had been identified in croaker ovaries, which suggests that androgens also have important functions in female gonadal physiology. The effects of androgens on ovarian estradiol production in vitro were investigated. Low physiological (10 nM) concentrations of a variety of androgens inhibited gonadotropin-stimulated estradiol secretion. These actions were not reversed by co-incubation with two nuclear androgen receptor antagonists (p,p'-DDE and cyproterone acetate, antiandrogens), which suggests that this androgen action is not mediated by binding to the nuclear androgen receptor. Experiments demonstrating that this action is rapid, nongenomic, and is at the cell surface provide further evidence that this action is nonclassical and does not involve binding to the nuclear androgen receptor. Further, a high affinity androgen binding moiety has been characterized on croaker ovarian membranes, which fulfills all the criteria for its designation as an androgen membrane receptor. This is the first clear evidence for the presence of androgen membrane receptors in vertebrate ovaries. Preliminary evidence indicates that various xenobiotic chemicals that display relatively high binding affinities for the ovarian nuclear androgen receptor display little or no binding to the ovarian membrane receptor, suggesting that this may not be an important site of endocrine disruption.

Studies on Three Estrogen Nuclear Receptors

Previously, cDNAs encoding three distinct estrogen receptors had been identified in the Atlantic croaker: ER alpha, ER beta, and ER gamma. This was the first evidence for three receptor subtypes in any vertebrate species. The distribution of the three ER subtypes was investigated in croaker brain tissues by in situ hybridization. Differences were observed in the distribution of ER beta and ER gamma in several brain regions, including those involved in the regulation of the reproductive endocrine system. For example, ER beta was detected in the magnocellular neurons of the preoptic area of the anterior hypothalamus, whereas ER gamma was absent. The ligand binding domains of the three receptors have been transfected in an Escherichia coli expression system and the steroid binding characteristics of the recombinant proteins investigated in steroid competition assays. Interestingly, several differences in the binding affinities of ER beta and ER gamma for estrogens have been observed, including a much lower affinity of ER gamma for estriol, an estrogen of physiological importance in mammals.

Future Activities:

The differing patterns of ER subtype expression in reproductive tissues as well as their steroid (and possible xenoestrogen) binding affinities will be considered when examining the effects of xenoestrogen exposure and in interpreting the results of these studies.

Journal Articles on this Report : 5 Displayed | Download in RIS Format

Other project views: All 18 publications 5 publications in selected types All 5 journal articles
Type Citation Project Document Sources
Journal Article Hawkins MB, Thornton JW, Crews D, Skipper JK, Dotte A, Thomas P. Identification of a third distinct estrogen receptor and reclassification of estrogen receptors in vertebrates (submitted for publication). R826125 (2001)
not available
Journal Article Larsson DG, Sperry TS, Thomas P. Regulation of androgen receptors in Atlantic croaker brains by testosterone and estradiol. General and Comparative Endocrinology 2002;128(3):224-230 R826125 (2001)
not available
Journal Article Loomis AK, Thomas P. Binding characteristics of estrogen receptor (ER) in Atlantic croaker (Micropogonias undulatus) testes: Different affinity for estrogens and xenobiotics from that of hepatic ER. Biology of Reproduction 1999;61(1):51-60. R826125 (2001)
not available
Journal Article Loomis AK, Thomas P. Effects of estrogens and xenoestrogens on androgen production by Atlantic croaker testes in vitro: Evidence for a nongenomic action mediated by an estrogen membrane receptor. Biology of Reproduction 2000;62(4):995-1004. R826125 (1999)
R826125 (2001)
not available
Journal Article Thomas P. Chemical interference with genomic and nongenomic actions of steroids in fishes: role of receptor binding. Marine Environmental Research 2000;50(1-5):127-134. R826125 (1999)
R826125 (2001)
not available
Supplemental Keywords:

fish reproduction, endocrine disruptors, androgen receptor, estrogen receptor, xenoestrogens, DDT, sex differences, estuarine, sex, reproductive effects., RFA, Health, Scientific Discipline, Toxics, Geographic Area, Environmental Chemistry, Health Risk Assessment, pesticides, State, Endocrine Disruptors - Environmental Exposure & Risk, endocrine disruptors, Biochemistry, Children's Health, Biology, Endocrine Disruptors - Human Health, puberty, fish, anti-androgen, embryo survival, xenobiotic, endocrine disrupting chemicals, steroid, animal models, DDT, developmental processes, toxicity, estrogen response, reproductive processes, Atlantic Croaker, biological effects, DDE, estrogen receptors, California (CA)

Progress and Final Reports:
Original Abstract
1999 Progress Report