Grantee Research Project Results
2000 Progress Report: Neuron Deficits and Degeneration Induced by Methylmercury
EPA Grant Number: R827096Title: Neuron Deficits and Degeneration Induced by Methylmercury
Investigators: Charleston, Jay S. , Kavanagh, Terrance J
Institution: University of Washington , Institute of Neurotoxicology and Neurological Disorders
EPA Project Officer: Hahn, Intaek
Project Period: August 1, 1998 through July 31, 2000 (Extended to July 31, 2001)
Project Period Covered by this Report: August 1, 1999 through July 31, 2000
Project Amount: $255,001
RFA: Exploratory Research - Environmental Biology (1998) RFA Text | Recipients Lists
Research Category: Biology/Life Sciences , Human Health , Aquatic Ecosystems
Objective:
The broad, long-term objective of this research project is to determine potential mechanism(s) by which low-dose methylmercury (MeHg) exposure alters perinatal brain development. The experimental design seeks to test the hypothesis that perinatal exposure to low levels of MeHg reduces neuronal cell number precursors (resulting in decreased neuron number) and disrupts the normal pattern of cell migration (resulting in ectopic placement of neurons) during organogenesis of the brain. Studies to examine the mechanisms that may be involved in reducing neuron numbers and disrupting cell migration are proposed. These mechanisms include investigation of the effects that low-level perinatal MeHg exposure has on: (1) changes in levels of polymerized microtubules; (2) changes in levels of expression of neural cell adhesion molecules (NCAM); (3) disruption of glutathione (GSH) and GSH cycle-related enzymes; and (4) increased levels of apoptosis within the developing brain.
These studies, featuring a mouse model, combine state-of-the-art unbiased stereological analysis of changes in cell numbers (neurons, astrocytes, microglia, and oligodendrocytes within four specific brain sites-cerebral cortex, hippocampus, thalamus, and cerebellum) with quantitative fluorescence immunocytochemistry, quantitative assessment of apoptosis, and quantitative analytical assay of total mercury within the brain tissue. The goal of this research project is to provide a level of sensitivity beyond that provided by traditional neuropathologic procedures. The premise of the proposed research is that the effects of chronic, low-level MeHg exposure on brain structure during development are most likely subtle in nature, requiring the use of sensitive quantitative methods to evaluate changes in specific cell types. The proposed studies in this application will help establish a rodent model for more mechanistic studies of low-level MeHg neurotoxicity.
Study Overview
This study has been designed to evaluate the consequences of very low perinatal MeHg exposure on the development of the central nervous system in a mouse model. Exposure levels were designed to provide 0.00, 0.5 , 1.0, and 4.0 ppm exposure to the dams, resulting in gestational and nursing of pups. The exposure design created a 4 x 4 tissue matrix derived from four exposure durations and four exposure doses. The exposure durations resulted in tissues generated on gestation day (GD) 14, GD 18 (immediately prebirth), postnatal day (PND) 14, and PND 21. Approximately 384 brain samples (regions) are being quantified in respect to cell number of neurons, astrocytes, microglia, and oligodendrocytes; a similar number of brain sites are being used for quantitative image analysis of target epitopes.
Progress Summary:
Current progress has been summarized in the Table below. For the purpose of this report, the project has been divided into the following categories: in-life (dosing and tissue collection), MeHg tissue quantification, MeHg dosing solution confirmation, stereological analysis, and quantitative fluorescence/image analysis.
Phase | Status |
In-life | Completed |
Mercury tissue analysis | Completed |
Dosing solution analysis | Completed |
Tissue collection | Completed |
Tissue processing (fixation, embedment) | Completed |
Stereology | In progress-approximately 50% completed |
GSH localization | Not completed-sensitivity and technical difficulties with collection of fresh tissue has proven to not work in this experimental design. |
GFAP quantitative fluorescence | GFAP staining-image capture has been completed, image analysis in progress |
NCAM quantitative image analysis | Not completed |
GSH Quantitative image analysis | Not completed |
Future Activities:
The research plan will continue. Progress on the remaining work items has been hampered somewhat due to difficulties in establishing the INND as a new research institute. Several logistical problems associated with this new research institute have required more effort than anticipated. However, we are confident that continued good progress on this project will occur. Immediate goals include continuing stereology work and quantitative image analysis of NCAM- and GSH-related targets.
Journal Articles:
No journal articles submitted with this report: View all 1 publications for this projectSupplemental Keywords:
methylmercury, MeHg, perinatal exposure, brain development, neural cell adhesion molecules, glutathione, enzymes, apoptosis, mouse model, neurotoxicity., RFA, Scientific Discipline, Health, Toxics, Water, National Recommended Water Quality, Environmental Chemistry, Chemistry, Epidemiology, Susceptibility/Sensitive Population/Genetic Susceptibility, genetic susceptability, Biology, Mercury, cerebral cortex, mercury uptake, teratogen, lead, cerebellum, dose response, exposure, neurotoxicity, methylmercury, scanning laser cytometry, neurodevelopmental toxicity, brain development, water quality, hippocampus, mercury induced degeneration, pregnancy, mercury concentrations, neuron deficits, environmental hazard exposuresProgress and Final Reports:
Original AbstractThe perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.