Exploring mechanisms of lead toxicity in the zebrafish model organism.

摘要

The use of the heavy metal lead (Pb) has resulted in widespread human exposures. Active efforts to reduce use and exposures have been successful in reducing Pb levels in the general population. However, studies show that adverse effects to the developing nervous system occur at levels once considered safe. Despite what is known about the effects of Pb exposure, the mechanisms of toxicity have not yet been completely elucidated. In particular, the question as to whether developmental exposure to Pb might influence the onset of diseases later in life has not been addressed. The studies in this dissertation were designed to further develop the use of the zebrafish model organism for use in toxicology experiments with a focus on developmental Pb exposure. A protocol was developed to assay gene expression alterations induced by Pb exposure on a global level using microarray analysis and to interpret these results with gene ontology analysis. Exposure to 100 ppb Pb in aquaria water starting shortly after fertilization throughout 72 hours post-fertilization resulted in the expression change of 90 genes. The genes were heavily enriched with neurological functions and with associations with neurological diseases. In contrast, only 30 genes were altered at 120 hours-post fertilization and these genes were not enriched in neurological function. Among the genes altered by Pb exposure was reelin (reln), a gene that plays critical roles throughout and following neuronal development. The alteration in the expression of this gene was further investigated by quantitative PCR and in situ hybridization at different developmental time points and at various exposure conditions. It was demonstrated that the expression of reln was only altered in a short critical window between 60 and 72 hours post-fertilization. To further our understanding of the effect of Pb on neurodegenerative disorders, genes associated with Alzheimer's and Parkinson's disease were analyzed in the zebrafish. Analysis included sequence comparison, as well as expression localization during development and in the adult brain. These data provide a framework for future analysis of the influence of Pb or other environmental contaminants on disease pathogenesis.