Patricia Braun and Emily Petruccelli will Present their Research Wednesday, 3/11/2015
Genome-wide analysis of differential DNA methylation due to social defeat stress mouse model
Early life stress is a significant risk factor for anxiety and depressive disorders, and DNA methylation (DNAm) plays an important role in the intervening pathways. The extent to which DNAm is altered across the genome in these disorders is unknown. We sought to identify the genes and regulatory sequences altered by stress in a mouse model of the disorders. Using a social defeat stress paradigm, we assessed genome-wide DNAm changes in mouse brain. We used two cohorts of stressed mice (N=7 and 7) and non-stressed controls (N=8 and 12), for discovery and validation, respectively. DNA was extracted from the dentate gyrus using a punch technique. DNA from the first cohort was examined in the discovery phase using the target enrichment system Methyl-Seq, which captures ~220,000 CpG-rich regulatory regions of the genome. DNA from the second cohort was used for validation with bisulfite pyrosequencing. Analysis of the Methyl-Seq data was done using GENESPRING and MethylKit. Thirty-three regions were significant in the discovery phase. Of these, 31 did not validate. In the remaining two regions, we detected differentially methylated loci between stressed and control groups. These loci represent an intronic region of Drosha (encoding a microRNA processing protein) and an intergenic region on chromosome X (IntchX). However when these two regions were further interrogated with a new cohort of mice (N=16 stressed and non-stressed), only IntchX replicated. IntchX included eight differentially methylated CpGs over 150 bps within an evolutionary conserved region. Because only one differentially methylated region due to social defeat stress region was replicated, further analysis of the Methyl-Seq data with follow-up bisulfite pyrosequencing will be performed to elucidate additional regions. Moreover, the nucleus accumbens, the brain region essential to reward, will be investigated for DNAm differences induced from the social defeat stress paradigm.
Sleep Abnormalities in a Drosophila Model of Human GEFS+
Despite an established link between epilepsy and sleep behavior, it remains unclear exactly how specific epileptogenic mutations affect sleep and how sleep influences epileptic seizures. Drosophila is an attractive model for studying the underlying mechanisms of this seizure/sleep relationship as it is routinely used to examine the genetic basis of seizure susceptibility and sleep behavior. Sun et al (2012) recently created a knock-in fly model of human Generalized Epilepsy with Febrile Seizures Plus (GEFS+), a wide spectrum disorder characterized by fever-associated seizing in childhood and lifelong affliction. GEFS+ flies carry a mutation in the voltage-gated sodium channel (Nav) gene, mimicking a disease-causing human Nav mutation (SCN1AK1270T) and display a semidominant heat-induced seizure phenotype as a result of abnormal electrophysiology in inhibitory GABAergic neurons. We found that GEFS+ mutation also dominantly modifies sleep behavior, with mutants exhibiting rapid sleep onset at dusk and increased nighttime sleep as compared to controls. This sleep profile was observed regardless of sex, mating status, and genetic background. Mutants’ exaggerated sleep was more resistant to carbamazepine (CBZ), a drug that reduces Drosophila GABAA receptor activity, and could be suppressed by either constant or acute scotophase light. We further observed that GEFS+ flies have normal circadian rhythm in free-running dark conditions, but significantly lack homeostatic rebound following sleep deprivation. Intriguingly, sleep deprivation treatment increased the heat-induced seizure susceptibility of control flies, but reduced the seizure severity of GEFS+ mutants. Ongoing experiments are addressing the potential significance of GABAergic inhibition on wake-promoting PDF+ neurons in GEFS+ mutant sleep and the impact of seizing on subsequent sleep behavior. Our findings thus far have characterized the sleep architecture of Drosophila harboring a human GEFS+ mutation and provided unique insight into the relationship between sleep and epilepsy.