Impaired repression at a vasopressin promoter polymorphism in a rat model of trait anxiety and depression

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.