Background Neurons containing proopiomelanocortin (POMC) derived peptides, known to control stress axis, metabolic and immune functions, have a lower function in patients with a family history of alcoholism, raising the possibility that alcohol effects on the POMC system may transmit through generations. Here we describe epigenetic modifications of Pomc gene that transmit through generation via male germline and may be critically involved in alcoholism-inherited diseases. Methods Whether an epigenetic mechanism is involved in causing a Pomc expression deficit in fetal alcohol exposed rats is studied by determining Pomc gene methylation, expression and functional abnormalities and their normalization following suppression of DNA methylation or histone acetylation. Additionally, transgenerational studies were conducted to evaluate the germline-transmitted effect of alcohol. Results Fetal alcohol exposed male and female rat offspring showed a significant deficit in POMC neuronal functions. Associated with this was an increased methylation status of several CpG dinucleotides in the proximal part of the Pomc promoter region and altered level of histone modifying proteins and DNA methyltransferases levels in POMC neurons. Suppression of histone deacetylation and DNA methylation normalized Pomc expression and functional abnormalities. Fetal alcohol-induced Pomc gene methylation, expression and functional defects persisted in the F2 and F3 male but not in female germline. Additionally, the hypermethylated Pomc gene was detected in sperms of fetal alcohol exposed F1 offspring that was transmitted through F3 generation via male germline. Conclusions Trangenerational epigenetic studies should spur new insight into the biological mechanisms that influence the sex-dependent difference in genetic risk of alcoholism-inherited diseases.

Background Prenatal exposure to ethanol reduces the expression of hypothalamic proopiomelanocortin (POMC) gene, known to control various physiological functions including the organismal stress response. In this study, we determined whether the changes in POMC neuronal functions are associated with altered expressions of histone-modifying and DNA-methylating enzymes in POMC-producing neurons, since these enzymes are known to be involved in regulation of gene expression. In addition, we tested whether gestational choline supplementation prevents the adverse effects of ethanol on these neurons. Methods Pregnant rat dams were fed with alcohol-containing liquid diet or control diet during gestational days 7 and 21 with or without choline, and their male offspring rats were used during the adult period. Using double-immunohistochemistry, real-time reverse transcription polymerase chain reaction (RT-PCR) and methylation specific RT-PCR, we determined protein and mRNA levels of histone-modifying and DNA-methylating enzymes, and the changes in POMC gene methylation and expression in the hypothalamus of adult male offspring rats. Additionally, we measured the basal and lipopolysaccharide (LPS)-induced corticosterone levels in plasma by enzyme-linked immunoabsorbent assay. Results Prenatal ethanol treatment suppressed hypothalamic levels of protein and mRNA of histone activation marks (H3K4me3, Set7/9, acetylated H3K9, phosphorylated H3S10) increased the repressive marks (H3K9me2, G9a, Setdb1) and DNA methylating enzyme (Dnmt1) and the methyl-CpG-binding protein (MeCP2). The treatment also elevated the level of POMC gene methylation, while it reduced levels of POMC mRNA and β-EP, and elevated corticosterone response to LPS. Gestational choline normalized the ethanol-altered protein and the mRNA levels of H3K4me3, Set7/9, H3K9me2, G9a, Setdb1, Dnmt1 and MeCP2. It also normalizes the changes in POMC gene methylation and gene expression, β-EP production and the corticosterone response to LPS. Conclusions These data suggest that prenatal ethanol modulates histone and DNA methylation in POMC neurons that may be resulting in hypermethylation of POMC gene and reduction of POMC gene expression. Gestational choline supplementation prevents the adverse effects of ethanol on these neurons.

Recent evidence suggests that physical and mental health are influenced by an intricate interaction between genes and environment. Environmental factors have been shown to modulate neuronal gene expression and function by epigenetic mechanisms. Exposure to these factors including nutrients during sensitive periods of life could program brain development and have long-lasting effects on mental health. Studies have shown that early nutritional intervention that includes methyl-donors improves cognitive functions throughout life. Choline is a micronutrient and a methyl donor that is required for normal brain growth and development. It plays a pivotal role in maintaining structural and functional integrity of cellular membranes. It also regulates cholinergic signaling in the brain via the synthesis of acetylcholine. Via its metabolites, it participates in pathways that regulate methylation of genes related to memory and cognitive functions at different stages of development. Choline-related functions have been dysregulated in some neurodegenerative diseases suggesting choline role in influencing mental health across the lifespan.

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.

Neurodegenerative diseases are a major public health problem worldwide with a wide spectrum of symptoms and physiological effects. It has been long reported that the dysregulation of the cholinergic system and the adrenergic system are linked to the etiology of Alzheimer’s disease. Cholinergic neurons are widely distributed in brain regions that play a role in cognitive functions and normal cholinergic signaling related to learning and memory is dependent on acetylcholine. The Locus Coeruleus norepinephrine (LC-NE) is the main noradrenergic nucleus that projects and supplies norepinephrine to different brain regions. Norepinephrine has been shown to be neuroprotective against neurodegeneration and plays a role in behavior and cognition. Cholinergic and adrenergic signaling are dysregulated in Alzheimer’s disease. The degeneration of cholinergic neurons in nucleus basalis of Meynert in the basal forebrain and the degeneration of LC-NE neurons were reported in Alzheimer’s disease. The aim of this review is to describe current literature on the role of the cholinergic system and the adrenergic system (LC-NE) in the pathology of Alzheimer’s disease and potential therapeutic implications.