Date of Defense

4-9-2004

Department

Geosciences

First Advisor

Dr. Charles Ide

Second Advisor

Dr. Anna Jelaso

Third Advisor

Dr. John Spitsbergen

Keywords

Aroclor 1254, NGF, immunocytochemistry, RT-PCR, gene expression, frog, development

Abstract

Polychlorinated biphenyls are ubiquitous, persistent environmental contaminants that cause a variety of adverse health effects wildlife and humans. PCBs are the major contaminant in the Kalamazoo River Superfund Site, in which the Environmental Institute is funded to study. Projects at the Environmental Institute are focused on developing molecular biotechnology tools that will improve environmental risk assessment. Previous acute studies involved the exposure of developing tadpoles, Xenopus laevis to PCBs through aquarium water in order to measure gene expression. Nerve growth factor (NGF) gene expression showed changes in these previous studies. NGF is a neurotrophic factor that regulates the growth and development of specific neurons. Jelaso et al. (2003) showed that tadpoles exposed to low doses (5ppb and 50ppb) of the PCB mixture, Aroclor 1254, caused significant increases in NGF gene expression in 18-day old tadpoles independent of adverse health effects. Tadpoles exposed to higher doses of PCBs (500- 700ppb) caused decreases in NGF gene expression that occurred as adverse health effects appeared in 11-day old tadpoles, and in doses of l-50ppm in 11-day old tadpoles (Jelaso et al. 2002). My thesis project is designed as an extension of these studies aimed at exposing tadpoles long-term, using a more natural means of exposure. My thesis is divided into two parts; part one involved a long-term dietary exposure of tadpoles to A1254. Changes in NGF gene expression were measured and health and development were charted. Tadpoles exposed to low doses of A1254 (24ppm) showed increases in NGF gene expression similar to previous acute, DMSO assisted studies.

Part 2 was an extension of part 1 focused on measuring NGF protein expression levels in tadpoles exposed through their diet. Immunocytochemistry (ICC) was used to characterize NGF protein expression in normal development. ICC conditions were optimized in order to reduce background staining and increase specific staining of the NGF antibody. 5, 10, and 20-day old tadpole tissue sections were stained and 10-day old tadpoles were analyzed as a guideline for the remaining sections. NGF is expressed in many tissues in 10-day old tadpoles including sensory neurons and cranial ganglia associated with modulating sensory information, motor neurons in the spinal cord, specific sensory regions in the oral cavity, eye and ear. NGF has been expressed in the cranial ganglia, eye, inner ear, oral cavity, and other sensory related tissues in mice, rats and chick embryos. This is the first study outlining the presence of NGF in the tissues of the developing frog. The next step in this study will be to determine a common functional role between tissues containing NGF.

Access Setting

Honors Thesis-Open Access

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