Differential Activity by Polymorphic Variants of a Remote Enhancer that Supports Galanin Expression in the Hypothalamus and Amygdala: Implications for Obesity, Depression and Alcoholism

Abstract

The expression of the galanin gene (GAL) in the paraventricular nucleus (PVN) and in the amygdala of higher vertebrates suggests the requirement for highly conserved, but unidentified, regulatory sequences that are critical to allow the galanin gene to control alcohol and fat intake and modulate mood. We used comparative genomics to identify a highly conserved sequence that lay 42 kb 5' of the human GAL transcriptional start site that we called GAL5.1. GAL5.1 activated promoter activity in neurones of the PVN, arcuate nucleus and amygdala that also expressed the galanin peptide. Analysis in neuroblastoma cells demonstrated that GAL5.1 acted as an enhancer of promoter activity after PKC activation. GAL5.1 contained two polymorphisms; rs2513280(C/G) and rs2513281(A/G), that occurred in two allelic combinations (GG or CA) where the dominant GG alelle occurred in 70-83 % of the human population. Intriguingly, both SNPs were found to be in LD (R(2) of 0.687) with another SNP (rs2156464) previously associated with major depressive disorder (MDD). Recreation of these alleles in reporter constructs and subsequent magnetofection into primary rat hypothalamic neurones showed that the CA allele was 40 % less active than the GG allele. This is consistent with the hypothesis that the weaker allele may affect food and alcohol preference. The linkage of the SNPs analysed in this study with a SNP previously associated with MDD together with the functioning of GAL5.1 as a PVN and amygdala specific enhancer represent a significant advance in our ability to understand alcoholism, obesity and major depressive disorder.

Magnetic Field-Assisted Gene Delivery: Achievements and Therapeutic Potential

“…Magnetofection has been also used for effective gene transfer in cultures of multipotent rat neural precursor cells and rat oligodendrocyte precursors [21,22]. In primary cultures of rat hypothalamic neurons, magnetofection was used to transfect the CG and CA alleles of an enhancer sequence related to galanin expression [23]. Magnetofection in mouse embryonic motor neurons was used to transfect a plasmid encoding the gene for a fluorescent protein fused to the spinal muscular atrophy-disease protein Smn.…”

Brain galanin system genes interact with life stresses in depression-related phenotypes

“…3. The GAL gene is widely expressed in the human brain [e.g., LC, forebrain, amygdala, and hypothalamus (26,51)], but its involvement in the development of depression is not well understood. Although previous studies indicated that it might have a sex-or estrogen-dependent effect (49), in our study the galanin system genetic variants significantly influenced the depressionrelated phenotypes both in males and in females, with similar magnitude of effect size, providing evidence that the excessive stress effect is not mediated by sex.…”

“…Furthermore, recent functional studies provided the first evidence that polymorphisms in a highly conserved genetic region upstream from the GAL gene regulates GAL expression in brain areas, such as the amygdala and hypothalamus, implicated in the pathogenesis of depression (51,52).…”

Distinct features of neurotransmitter systems in the human brain with focus on the galanin system in locus coeruleus and dorsal raphe

“…Thus, galanin interacts with 5-HT1A receptors, and galanin antagonists and agonists have anxiolytic and antidepressive effects in animal experiments (144)(145)(146)(147)(148)(149)(150)(151). Interestingly, association of genes encoding galanin and/or GalR3 has been reported for psychiatric phenotypes (152)(153)(154), including panic disorder (155), depression-related parameters (156)(157)(158), and nicotine dependence (159,160).…”