Fetal Alcohol Exposure Alters Proopiomelanocortin Gene Expression and Hypothalamic-Pituitary-Adrenal Axis Function via Increasing MeCP2 Expression in the Hypothalamus

Abstract

Proopiomelanocortin (POMC) is a precursor gene of the neuropeptide β-endorphin in the hypothalamus and is known to regulate various physiological functions including stress response. Several recent reports showed that fetal alcohol exposure programs the hypothalamus to produce lower levels of POMC gene transcripts and to elevate the hypothalamic-pituitary-adrenal (HPA) axis response to stressful stimuli. We investigated the role of methyl CpG binding protein (MeCP2) in the effects of prenatal ethanol on POMC gene expression and hypothalamic-pituitary-adrenal (HPA) axis function. Pregnant Sprague Dawley rats were fed between GD 7 and 21 with a liquid diet containing 6.7% alcohol, pair-fed with isocaloric liquid diet, or fed ad libitum with rat chow, and their male offsprings were used at 60 days after birth in this study. Fetal alcohol exposure reduced the level of POMC mRNA, but increased the level of DNA methylation of this gene in the arcuate nucleus (ARC) of the hypothalamus where the POMC neuronal cell bodies are located. Fetal alcohol exposed rats showed a significant increase in MeCP2 protein levels in POMC cells, MeCP2 gene transcript levels as well as increased MeCP2 protein binding on the POMC promoter in the arcuate nucleus. Lentiviral delivery of MeCP2 shRNA into the third ventricle efficiently reduced MeCP2 expression and prevented the effect of prenatal ethanol on POMC gene expression in the arcuate nucleus. MeCP2-shRNA treatment also normalized the prenatal ethanol-induced increase in corticotropin releasing hormone (CRH) gene expression in the hypothalamus and elevated plasma adrenocorticotrophic hormone (ACTH) and corticosterone hormone responses to lipopolysaccharide (LPS) challenge. These results suggest that fetal alcohol programming of POMC gene may involve recruitment of MeCP2 on to the methylated promoter of the POMC gene to suppress POMC transcript levels and contribute to HPA axis dysregulation.

Preconception Alcohol Increases Offspring Vulnerability to Stress

“…Aza treatment did not elicit any increased expression of Pomc in control samples and Crf in AF samples as they have reduced levels of methylation. We and others previously reported that hypermethylated promoters recruit methyl CpG binding proteins to repress the transcription of Pomc and tumor suppressor genes (Gangisetty et al, 2014;Lopez-Serra et al, 2008). In our experiments, Aza elicited its effect on stress regulatory genes by reducing their methylation and increasing expression.…”

DNA Methylation, Substance Use and Addiction: a Systematic Review of Recent Animal and Human Research from a Developmental Perspective

“…Three studies from the same working group found that prenatal alcohol exposure associated with increased Pomc methylation in the rat hypothalamus, which in turn related to decreased gene expression [13,15,16]. These changes were maintained transgenerationally (up to three generations), but could be rescued by gestational choline supplementation.…”

“…We do, however, highlight five genes that were investigated by multiple studies and, promisingly, showed a consistent direction of associations. In three rodent studies from the same working group [13,15,16], increased methylation and decreased expression of Pomc-a gene implicated in stress response, metabolism, and immune function-was observed in response to prenatal alcohol exposure across multiple tissues. These results highlight one mechanism through which fetal alcohol programming can occur, contributing to HPA axis dysregulation and increased addiction risk [16].…”

“…In three rodent studies from the same working group [13,15,16], increased methylation and decreased expression of Pomc-a gene implicated in stress response, metabolism, and immune function-was observed in response to prenatal alcohol exposure across multiple tissues. These results highlight one mechanism through which fetal alcohol programming can occur, contributing to HPA axis dysregulation and increased addiction risk [16]. In two other studies, the opioid receptor mu 1 (OPRM1)-a gene important for mediating drug-induced activation of reward pathways-was hypermethylated in the blood of former opiate addicts [41] and alcohol-dependent individuals [43].…”

“…The sampling of multiple tissues over time will also make it possible to establish cross-tissue variability and locate peripheral biomarkers that most closely resemble DNAm changes in neural networks underlying addiction. Furthermore, incorporating additional omics data, such as gene expression, serum levels, protein content, and enzymatic activity, will be useful for clarifying the functional relevance of observed DNAm changes at multiple biological levels [e.g., 13,16]. Importantly, the use of methyl-modifying agents (e.g., methionine, choline [15,32]) will offer valuable opportunities for testing the reversibility of drug-induced DNAm changes, identifying whether certain developmental periods are more sensitive to intervention, and examining whether normalisation of DNAm patterns parallel changes in addiction-relevant phenotypes.…”