Variation in somatic and ovarian development: predicting susceptibility of amphibians to estrogenic contaminants.
ABSTRACT Although amphibian sex determination is genetic, it can be manipulated by exogenous hormone exposure during sexual differentiation. The timing of sexual differentiation varies among anuran amphibians such that species may or may not be a tadpole during this period, and therefore, may or may not be exposed to aquatic contaminants. Estrogenic contamination is present in amphibian habitats worldwide. We examined three species with varying somatic and ovarian developmental rates to assess their susceptibility to estrogenic contaminants. American toads (Bufo americanus), gray treefrogs (Hyla versicolor), and Southern leopard frogs (Ranasphenocephala) were exposed as larvae to 17-beta-estradiol (10(-7)M), three concentrations of a widespread herbicide (1, 3, 30 ppb atrazine), or a solvent control (ethanol). Somatic and ovarian developmental stages as well as time to metamorphosis were recorded. Toads and treefrogs were examined at three weeks and metamorphosis, while leopard frogs were examined at three, six, and nine weeks as well as at metamorphosis. Our results demonstrate that each species displays heterochronic somatic and ovarian development. Further, the more rapid of the two rates determines the susceptibility to estrogenic contaminants. These results suggests that amphibians with shorter larval periods, and therefore quicker somatic developmental rates (i.e. American toads, gray treefrogs), are more susceptible to somatic treatment effects (i.e. prolonged time to metamorphosis) due to estrogenic contaminants. Moreover, the results suggest that amphibians with relatively rapid ovarian development (i.e. Southern leopard frogs) are more susceptible to gonadal treatment effects caused by estrogenic contaminants.
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ABSTRACT: We examined the effects of atrazine (0–20 mg/L) on embryos, larvae, and adult anuran amphibian species in the laboratory. Atrazine treatments did not affect hatchability of embryos or 96-h posthatch mortality of larvae of Rana pipiens, Rana sylvatica, or Bufo americanus. Furthermore, atrazine had no effect on swimming speed (measured for R. pipiens only). However, there was a dose-dependent increase in deformed larvae of all three species with increasing atrazine concentration. In adult R. pipiens, atrazine increased buccal and thoracic ventilation, indicating respiratory distress. However, because atrazine had no affect on hemoglobin, this respiratory distress was probably not indicative of reduced oxygen-carrying capacity of the blood. Frogs exposed to the highest atrazine concentration stopped eating immediately after treatment began and did not eat during the 14-d experiment. However, no decreases in mass were measured even for frogs that were not eating, probably because of compensatory fluid gain from edema. Atrazine concentrations found to be deleterious to amphibian embryos and adults are considerably higher than concentrations currently found in surface waters in North America. Therefore, direct toxicity of atrazine is probably not a significant factor in recent amphibian declines.Environmental Toxicology and Chemistry 03/2001; 20(4):769 - 775. · 2.62 Impact Factor
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ABSTRACT: Several environmental chemicals are known to have estrogenic activity by interacting with development and functions of endocrine systems in nearly all classes of vertebrates. In order to get a better insight of potential estrogenic effects on amphibians caused by environmental pollution this study aims to develop a model for investigating endocrine disruptors using the amphibian Xenopus laevis. In that model the potential estrogenic activity of endocrine disruptors is determined at several levels of investigation: (I) binding to liver estrogen receptor; (II) estrogenicity in vitro by inducing vitellogenin synthesis in primary cultured hepatocytes; and (III) in vivo effects on sexual development. Here we deal with establishing methods to assay estrogenic activity of environmental chemicals in vitro and in vivo. In vitro we used a semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) technique to determine mRNA-induction of the estrogenic biomarker vitellogenin in primary cultured hepatocytes of male Xenopus laevis. Time courses of vitellogenin-mRNA in the presence and absence of 10(-6) M 17 beta-estradiol (E2) resulted in a marked loss of mRNA from controls after 2 days while E2 treatment kept vitellogenin-mRNA at a relatively stable level. After 36 h of incubation estrogenic activities of E2, 4-nonylphenol (NP), and 2,2-bis-(4-hydroxyphenyl)-propan (bisphenol A) at concentrations ranging from 10(-10) to 10(-5) M were assayed by RT-PCR of vitellogenin-mRNA and showed the following ranking of dose-dependent potency: E2 > NP > bisphenol A. These in vitro results were confirmed further by in vivo experiments determining sexual differentiation of Xenopus laevis after exposure to E2 and environmental chemicals during larval development. Concentrations of 10(-7) and 10(-8) M E2 as well as 10(-7) M of NP or bisphenol A caused a significant higher number of female phenotypes compared to controls indicating a similar ranking of estrogenic potencies in vivo as in vitro. In addition, butylhydroxyanisol and octylphenol, both showed feminization at 10(-7) M while octylphenol was also effective at 10(-8) M. In summary these results demonstrate for the first time the use of a semiquantitative RT-PCR technique for screening estrogenicity by assaying mRNA induction of the estrogenic biomarker vitellogenin in vitro. The combination of this newly developed method with classical exposure experiments is necessary for determination of the biological significance of estrogenic chemicals.Science of The Total Environment 02/1999; 225(1-2):59-68. · 3.26 Impact Factor
Article: Complex Life Cycles11/2003; 11:67-93.