Date of Award

January 2012

Document Type

Thesis

Degree Name

Master of Science (MS)

Department

Biomedical Sciences

First Advisor

Joyce E. Ohm

Abstract

Amphetamine and cocaine has long been known as drug of abuse. These drugs impart their physiological manifestations by regulating the reward circuitry of the brain which is controlled primarily by the dopaminergic pathway where they control the release of dopamine. It has been found that repeated cocaine administration changes histone acetylation patterns at the promoter sequences of several important genes. Sirtuin proteins are histone deactylases that regulate important biological functions have also been found to be elevated in the nucleus accumbens after cocaine administration. Although there have been several studies that have tried to elucidate the mode of action of amphetamine, it is still not known whether the drug of abuse, induces heritable histone modifications at affects each of the genes of the dopaminergic pathway by inducing changes at the promoter regions of these genes. Therefore, in order to better understand the mechanism of the action of amphetamine, the goal of the present study is to look into the effect of amphetamine in stimulation of epigenetic changes by the induction of histone modifications at the promoters of genes of the dopaminergic pathway. We chose Caenorhabditis elegans (C. elegans) as our experimental model system for our study. Dopamine is synthesized in eight sensory neurons (4 CEP, 2 ADE and 2 PDE) and modulates locomotion, learning and egg laying activity. All the known dopaminergic components involved in DA synthesis, vesicle storage, release and reuptake are highly conserved between the worm and mammals.

In order to identify the histone modifications that are affected by amphetamine administration, immunoblot followed by mRNA expression was performed which revealed that amphetamine indeed affects the expression of histone modifications. For, gene specific investigation, chromatin immunoprecipitation was employed. Chromatin immunoprecipitation revealed that amphetamine treatment down regulates the expression of dat-1, the dopamine transporter and cat-1, vesicular monoamine transporter by reducing the enrichment of tri-methylated H3K4. Amphetamine treatment also increases the expression of cat-2, tyrosine hydroxylase by reducing the enrichment of tri-methylated H3K27.Our results also showed that amphetamine treatment has no effect on the expression of bas-1, the aromatic amino acid decarboxylase. We further confirmed our results by mRNA expression. These results indicate the role of amphetamine in inducing changes in the histone modifications, though further research is needed to confirm the observations.

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