Gene expression in the alcoholic brain

ABSTRACT Alcoholism inflicts great suffering on individuals and their families and constitutes a huge financial and health burden for society. Current treatments are only partially effective and not adequate because the extraordinary complexity of the condition has, to date, precluded a sufficient level of understanding for specific and successful treatments to be developed. Since alcoholism is a heterogeneous disorder there is unlikely to be a single effective therapeutic, necessitating the development of a range of options. Fortunately addiction neuroscience is a dynamic field and the quality and quantity of information that is now generated gives hope for the development of more useful treatments in the near future. Alcoholism causes long-lasting neuroadaptations resulting in poor health, brain damage, lost productivity, increased incidence of accidents and injury, violence, loss of social and occupational function and premature death. Ethanol alters brain structure and function in numerous and fundamental ways which persist during abstinence. Each of these long-term neuroadaptations is initiated by changes in gene expression which, with repeated alcohol use, become more permanent. The study of alcohol-induced changes in gene expression can aid in the identification of the molecular targets of ethanol and thereby advance our understanding of this disease. The aim of this thesis was to examine differences in gene expression in the brains of alcoholic and non-alcoholic human subjects. The first part involved the investigation of differentially expressed cDNA fragments identified by a PCR-differential display of the cortex of human alcoholics and controls. This led to the unexpected discovery of an upregulated non-coding RNA (ncRNA) in hippocampus, cerebellum and brainstem of human alcoholics and in hippocampus, cerebellum and cortex of alcohol-treated rats. RNA transcripts which do not encode protein but function as ncRNAs play a crucial role in the regulation of gene expression. Thus the increased ncRNA expression suggests that alcohol may influence the regulation of gene expression via its action on ncRNAs. In recent years the crucial importance of the amygdala in drug-seeking and relapse has been increasingly recognized. The wealth of information generated by microarrays can give a more detailed picture of the molecules which participate in the processes underlying the different forms of synaptic plasticity, so that more comprehensive and sophisticated models of the events underlying the development of alcoholism can be formed. For the second part of this work a 19K microarray was used to compare gene expression in the amygdala of alcoholic patients and control cases. The results show that alcoholism affects a broad range of genes and many systems in the amygdala including genes involved in glutamatergic function, synaptic transmission, structural plasticity, metabolism, transcription and RNA processing, energy metabolism, neurodegeneration, chromatin remodeling and the circadian cycle. The glutamate system is profoundly affected by alcoholism and in the amygdala it is involved in associative learning which plays an important role in alcoholism and other addictions. Downregulation of the excitatory amino acid transporters 1 (GLAST) and 2 (GLT-1) and the AMPA glutamate receptor 2 (GluR2) subunit revealed by the microarray were confirmed quantitatively by Western blots and qualitatively by immunohistochemistry. The decreased expression of GLAST, GLT-1 and GluR2 in the alcoholic patients may increase glutamate tone and activity in the amygdala and this may contribute to neurodegeneration as well as the expression of associative memories and anxiety which underlie continued drug-seeking and chronic relapse