Citation:

VILLENEUVE, D. L., I. KNOEBL, M. D. KAHL, K. M. JENSEN, D. E. HAMMERMEISTER, K. J. GREEN, L. S. BLAKE, AND G. T. ANKLEY. RELATIONSHIP BETWEEN BRAIN AND OVARY AROMATASE ACTIVITY AND ISOFORM-SPECIFIC AROMATASE MRNA EXPRESSION IN THE FATHEAD MINNOW (PIMEPHALES PROMELAS) - JOURNAL ARTICLE. AQUATIC TOXICOLOGY. Elsevier Science Ltd, New York, NY, 76:353-368, (2006).

Impact/Purpose:

The indeterminate condition of exposure indicator research stands to change markedly with the ability to connect molecular biological technologies with cellular or tissue effects and outcomes. Three focal areas of ecological research aim to develop a sequence of approaches where "the earliest recognizable signatures of exposure" (i.e., unique patterns of up- and down-regulated genes and proteins) are identified for numerous stressors, demonstrable in case studies and incorporated into Agency, State and Regional studies supported by EMAP and other programs.

Area 1, Computational Toxicology Research: Exposure assessment has historically been based on use of chemical analysis data to generate exposure models. While biological activity of chemicals has been recognized to be important for exposure risk assessments, measurement of such activity has been limited to whole organism toxicity tests. Use of molecular approaches will: improve extrapolation between components of source-to-outcome continuum (source , exposure , dose , effect , outcome).

Using a systems modeling approach, gene and protein expression data, in small fish models (fathead minnow and zebrafish), will be integrated with metabolomic and histopathological data. This will assist in prediction of environmental transformation and chemical effects based on structural characteristics, and enhance quantitative risk assessments, including areas of uncertainty such as a basis for extrapolation of effects of endocrine disrupting chemicals, interspecies extrapolation, complex chemical mixtures and dose-response assessment.

Area 2, Ecological Research-Environmental Diagnostics: Development of molecular diagnostic indicators contributes to several of the GPRA Diagnostic Research Goals. Methods will employ DNA microarray technology and expression proteomics, focusing on species of relevance to aquatic ecosystem risk assessment. Significantly, these diagnostic indicators will open the door to understanding subcellular interactions resulting from exposure to complex chemical mixtures.

define relationship between genetic disposition of populations and degree/specificity of stressor-specific gene transcriptional response in aquatic organisms (fish and invertebrates)

identify of chemical mixture induced transcriptional "patterns" using microarrays and hyperspectral scanning - via collaboration with DOE Sandia National Labs

apply molecular indicators to watershed level stressor study, including pilot studies with targeted pesticides and toxins indicators

develop molecular indicators of exposure for invertebrates (Daphnia, Lumbriculus, Chironomus)

Area 3, Exposure Research in Endocrine Disruptors:

Subobjective 1: Develop exposure methods, measurement protocols, and models for assessment of risk management practices of endocrine disrupting compounds. As risk management approaches are identified and developed, there will be a need to identify, adapt and develop bioassay screening tools and other analytical methods to assess their efficacy. Measurements research will be performed to define management needs. This effort will entail cross-lab participation from NRMRL, NERL and NHEERL.

Subobjective 2: Determine extent of environmental and human exposures to EDCs, characterize sources and factors influencing these exposures, develop and evaluate risk management strategies to reduce exposures. In order to develop effective risk management strategies, it is important to understand the extent of exposures to endocrine disrupting compounds and factors influencing source-to-exposure-to-dose relationships.

apply molecular indicators of exposure to estrogenic compounds in selected wastewater treatment plants located in ten USEPA Regions

identify differential gene expression following exposure of fathead minnows to environmental androgens and androgen-like compounds

apply molecular indicators of exposure to

Description:

There is growing evidence that some chemicals present in the environment have the capacity to inhibit, or potentially induce, aromatase activity. This study compared aromatase activities and isoform-specific mRNA expression in brain and ovary tissue from non-exposed fathead minnows representing three different ages and stages of reproductive activity, and from fathead minnows exposed to the aromatase inhibitor fadrozole for 7 d. The goal was to determine whether measures of a single aromatase endpoint in either brain or ovary tissue would be sufficient to understand and predict system-wide effects of endocrine disrupting chemicals on aromatase activity and transcript levels. Aromatase activity in the ovary, but not brain, varied significantly with age/reproductive category, with adults held in non-reproductive conditions showing significantly lower activity than juveniles and reproductively-active adults. Significant correlations between isoform-specific transcript levels and aromatase activity were observed for ovary tissue, but those relationships were not robust for all age/reproductive categories, nor were they sustained in fadrozole-treated fish. In vitro, fadrozole inhibited the aromatase activity of brain and ovary post-mitochondrial supernatants with similar potency, despite large differences in the magnitude of activity. In vio, fadrozole altered aromatase activity and isoform-specific transcript levels in both brain and ovary tissue, but concentration-response relationships were different for each tissue. Aromatase activity and P450aromB mRNA expression in brain showed a dose-dependent decrease at concentrations greater than 5.55 ug/L. In contrast, ovary activity showed an inverted U-shaped concentration-response consistent with the interplay between increased P450aromA transcript levels in ovary and competitive inhibition of the aromatase enzyme. As a whole, results of this study did not reveal any robust correlations between brain and ovary aromatase activity and/or isoform-specific mRNA expression. However, they were consistent with the current body of evidence related to teleost aromatase regulation, suggesting that increased understanding of the biology of aromatase may facilitate system-wide understanding of effects on aromatase based on relatively few measured endpoints.

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