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Murgatroyd, Christopher A. (2005): Impaired repression at a vasopressin promoter polymorphism in a rat model of trait anxiety and depression. Dissertation, LMU München: Faculty of Biology
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Abstract

Two inbred rat lines have been developed that show either high (HAB) or low (LAB) anxiety-related behavior. The behavioral phenotype correlates with arginine vasopressin (AVP) expression at the level of the hypothalamic paraventricular nucleus (PVN), but not supraoptic nucleus, with HAB animals overexpressing the neuropeptide in both magnocellular and parvocellular subdivisions of the PVN. This study aimed to investigate the molecular cause of the AVP overexpression in the PVN of the HAB animals. Sequencing of the AVP locus resulted in the detection of a number of single nucleotide polymorphisms (SNPs) in the promoter differing between the HAB and LAB animals. Two of the SNPs were embedded in cis-regulatory elements. Genotyping further revealed that the HAB-specific allele of the AVP gene promoter occurs in 1.5% of outbred Wistar rats. An assay was developed to address the question of differential allele-specific transcription rates between the LAB and HAB alleles using cross-mated HAB/LAB F1 animals. Results from the experiment revealed the HAB AVP promoter to be more transcriptionally active in vivo. EMSA assays confirmed that one specific SNP [A(-1276)G] conferred reduced binding of the transcriptional repressor CArG binding factor A (CBF-A) in the HAB allele. Reporter gene assays supported the view that the A(-1276)G transition in the CArG element impairs CBF-A repression in the HAB allele, which in turn relates to a weakened repression of the intact HAB AVP promoter by CBF-A. Furthermore, CBF-A is highly co-expressed in AVP-containing neurons of the PVN supporting an important role for regulation of AVP gene expression in vivo. Taken together, these results demonstrate a role for an AVP gene polymorphism and CBF-A in elevated AVP expression in the PVN of HAB rats likely to contribute to their behavioral and neuroendocrine phenotype. Phenotypic variation among organisms is central to evolutionary adaptations underlying natural and artificial selection, and also determines individual susceptibility to common diseases including cardiovascular, metabolic and age related and psychiatric diseases. Most phenotypic diversity in natural populations is characterised by differences in degree rather than in kind. In accordance with this view, HAB rats lacked changes in the coding part of the vasopressin gene, but instead were homozygous for the polymorphic promoter region. Therefore, the HAB specific AVP promoter represents a natural model for AVP overexpression and highlights, in turn, cognate molecular pathways which potentially fuel the resulting pathologies. Specifically, our finding that the SNP in position 1276 of the AVP gene promoter underlies AVP overexpression in the PVN of HAB rats, makes this SNP a potential target for further studies aimed at improving therapeutic tools. Finally, further studies are necessary to understand to which degree and in concert with which vulnerability loci vasopressin overexpression underlies the complex neuroendocrine and behavioural stigmata in the HAB line. Certainly, such studies will yield important insights into gene-gene interactions between susceptibility genes and shed light on additional pathways suitable for therapeutic interventions. In conclusion, this present work exemplifies that selective inbreeding for behavioural traits and combined phenotypic and molecular analysis of candidate genes is an important step and tool to address these issues.