In considering the behavioral variation that occurs in response to variations in the social environment, the mechanisms underlying persistent increases or decreases in neurotransmitter release, receptor levels and hormonal activation described in the previous sections must be determined. Advances in molecular biology have identified processes through which dynamic yet stable alterations in the activity of genes can be induced. The epigenetic regulation of transcription is a critical feature of the link between genotype and phenotype and refers to those factors which control accessibility of DNA to transcription and which can alter the levels of gene expression (either silencing genes or increasing transcriptional activity) without altering the sequence of DNA. The molecular mechanisms through which these epigenetic effects are achieved are diverse, including histone protein modifications and DNA methylation (Feng et al., 2007
; Razin, 1998
). Within the cell nucleus, DNA is wrapped around a core of histone proteins which can undergo multiple post-translational modifications including methylation, acetylation and ubiquination (Peterson and Laniel, 2004
; Zhang and Reinberg, 2001
). These modifications alter the dynamic interactions between the histones and DNA which either reduce or enhance the accessibility of DNA. In particular, histone acetylation is associated with increased transcriptional activity whereas histone deacetylation or methylation is typically associated with transcriptional repression. Acetylation of histones is mediated by the enzyme histone acetyltransferase (HAT) whereas histone deacetylase (HDAC) promotes removal of the acetyl group from the histone tails. Thus, through alterations in the conformation of histones, the accessibility of DNA can be rapidly and reversibly altered. In contrast, DNA methylation represents what is generally considered a more stable and enduring modification to the activity of genes. DNA methylation occurs when cytosine nucleotides, usually located in CpG islands, within DNA become converted to 5-methylcytosine. This process is mediated by methyltransferases which either promote maintenance (i.e.
DNMT1) or de novo
DNA methylation (i.e.
DNMT3) (Feng et al., 2007
; Razin, 1998
; Turner, 2001
). The conversion to 5-methylcytosine does not alter the DNA sequence but does reduce the likelihood that that sequence of DNA will be transcribed. Methylated DNA attracts methyl-binding proteins, such as MeCP2, which further reduce the accessibility of the gene and is associated with transcriptional repression (Fan and Hutnick, 2005
). The stability of DNA methylation patterns within the genome permits the stable regulation of gene expression associated with cellular differentiation. Importantly, during cell division, both DNA and DNA methylation patterns are inherited by daughter cells, thus allowing differentiated cells to transmit their phenotype to the next generation of cells (Fukuda and Taga, 2005
There is emerging evidence for the role of epigenetics in understanding the persistent effects of environmental exposures on the activity of genes with implications for brain and behavior. Moreover, these epigenetic effects are not limited to nutritional and toxicological exposures, as there is support for the hypothesis that the sustained effects of variations in social experience can be linked to epigenetic variation, particularly in DNA methylation patterns. The link between variations in postnatal maternal care, altered DNA methylation, and long-term changes in behavior in rodents, supports the role of this molecular mechanism in shaping individual differences in neurobehavioral outcomes. As described in the previous sections, natural variations in postnatal maternal LG in the rat are associated with changes in numerous receptor pathways, with effects on hippocampal GR being implicated in the high levels of HPA reactivity observed amongst offspring who receive low levels of LG (Liu et al., 1997
). Analysis of the GR 17
promoter region suggests that low levels of LG are associated with increased GR 17
methylation, decreased GR expression and an increased HPA response to stress. Time course analysis has indicated that these maternally induced epigenetic profiles emerge during the postnatal period and are sustained into adulthood (Weaver et al., 2004
). The pathways through which these effects are achieved are currently being elucidated and it appears likely that maternal LG mediated up-regulation of NGFI-A in infancy may be critical to activating GR transcription and maintaining low levels of DNA methylation within the GR 17
promoter amongst the offspring of High LG dams (Weaver et al., 2007
) (see ). Similarly, increased levels of ERα observed in the MPOA of the offspring of High LG dams may be induced through LG-mediated activation of the promoter region of this gene. Analysis of levels of DNA methylation within the 1B promoter region of the ERα gene in MPOA tissue indicates that High LG is associated with decreased promoter methylation whereas Low LG is associated with increased promoter methylation, leading to reduced gene expression and an attenuated response to hormonally-primed behaviors (Champagne et al., 2006
). Chromatin immunoprecipitation assays demonstrate that this differential DNA methylation has consequences for the binding of transcription factors such as STAT5a to the 1B promoter. Maternal LG is associated with increased levels of STAT5a during the postnatal period and the increased levels of this factor may lead to sustained activation of transcription and reduced DNA methylation.
Variations in early life maternal stimulation can alter offspring gene expression via epigenetic mechanisms
An epigenetic response to variations in the quality and frequency of mother-infant interactions has also been demonstrating using postnatal maternal separation and handling. Daily and prolonged maternal separation has effects on a broad range of neurotransmitter and neuropeptide systems, as described in the previous sections, and in a recent study, significant increase in AVP mRNA in the parvocellular neurons of the PVN which persist for a year following this early social experience were confirmed in mice (Murgatroyd et al., 2009
). Demonstrating the functional importance of this increased expression, the HPA hyperactivity and behavioral indices of anxiety-like and depressive responses observed amongst maternally separated mice could be attenuated if the mice were treated with a V1Br antagonist. Within the AVP gene, there are four regions rich in CpG islands that could potentially regulate gene expression through DNA methylation. Analysis of PVN tissue reveals that at one of these four regions (CGI3) maternally separated males have significantly reduced DNA methylation compared to control males at 6 weeks, 3 months and 1 year of age. Furthermore, this hypomethylation is significantly correlated with increased mRNA expression, and these effects are brain region specific as no changes in AVP mRNA or DNA methylation exist between maternally separated and control males in the SON. Analysis of the time-course of the molecular changes involved in this differential methylation suggests that short-term activation of the methyl binding protein, MeCP2, may be a critical factor within the pathways leading to AVP hypomethylation and increased AVP mRNA levels within the PVN (see ). Conversely, in response to brief postnatal maternal separation (handling), reductions in CRH mRNA in the parvocellular neurons of the PVN can be observed as earlier as PN9 and at this developmental time-point, PVN levels of the vesicular transporter of glutamate (vGlut2) (a marker for the activity of excitatory glutametergic inputs into CRH neurons) is also significantly lower in handled pups (Korosi et al., 2010
). However, these alterations in vGlut2 levels are transient and are not observed at later stages of development and, while they may be responsible for short-term changes in CRH mRNA, they cannot explain the long-term changes. Within the regulatory region of the CRH gene resides a binding element (NRSE) for the repressor neuron-restrictive silencer factor (NRSF) (Seth and Majzoub, 2001
). When bound, this factor recruits co-factors and other enzymes/proteins involved in epigenetic regulation leading to the repression of gene expression (Zheng et al., 2009
). Amongst handled offspring, protein levels of NRSF are dramatically higher in PVN tissue at PN9 and throughout adulthood, suggesting a possible mechanism for the initiation and maintenance of reduced CRH gene expression in response to handling-induced stimulation of mother-infant interactions (see ). Overall, these studies suggest that both up- and down-regulation of gene expression relevant to emotional phenotypes can be achieved through the recruitment of epigenetic mechanisms in response to early life social experiences.
In considering the neurobiological targets that are modulated by the social environment, one of the common downstream mediators of many of the neural systems described in this review is BDNF. For example, BDNF plays a critical role in the synaptic maturation triggered by GABA signaling (Represa and Ben-Ari, 2005
), and increases in BDNF are thought to mediate changes in structural plasticity in the brain associated with elevated levels of estradiol (Sato et al., 2007
) and glutamate (Marini et al., 1998
). Activation of the HPA response to stress is associated with increases in BDNF (Tapia-Arancibia et al., 2004
) and CRH mediated increases in BNDF can be inhibited through antagonism of CRH1r (Bayatti et al., 2005
). Interactions between BDNF and the mesolimbic dopamine system may modulate the response to reward, effects of psychostimulants, and depression (Berton et al., 2006
; Nestler and Carlezon, 2006
; Thomas et al., 2008
), and BDNF acts during development to shape dopaminergic, serotonergic, and GABAergic neuronal populations (Eaton et al., 1995
; Lu et al., 2009
; Zhou et al., 1996
). Epigenetic effects on BDNF of both postnatal and adult social experiences have been explored, and suggest that this activity of this gene may be highly susceptible to environmental regulation. In a rodent model of postnatal abuse, in which females engage in an increased frequency of rough handling, dragging, dropping, and stepping on pups (Roth and Sullivan, 2005
), offspring exposed to these abusive social interactions are found to have reduced expression of BDNF in the prefrontal cortex in adulthood associated with increased DNA methylation within the BDNF IV promoter region (Roth et al., 2009
). In adulthood, social defeat has likewise been demonstrated to alter BDNF levels. BNDF gene expression is significantly decreased in the hippocampus of socially defeated male mice and this effect appears to be mediated by specific decreases in the BDNF III and IV transcripts (Tsankova et al., 2006
). These effects are observed a month following exposure to the social stress, indicating a persistent effect on gene expression. Chromatin immunoprecipitation assay analysis indicates increased histone H3-K27 dimethylation at the BDNF III and IV promoters amongst socially defeated males which may account for the reduced BDNF expression. Histone deacetylase (HDAC5) mRNA levels are also found to be decreased in socially defeated males (Tsankova et al., 2006
) and HDAC5 appears to be important in mediating the effects of anti-depressant treatment in males exposed to chronic social stress (Renthal et al., 2007
). The differential levels histone H3-K27 dimethylation are also found across the genome within the NAc, both in response to chronic social defeat and prolonged adult social isolation (Wilkinson et al., 2009
). Analysis of histone acetylation in the NAc indicates that H3-K14 acetylation is initially decreased and then increased following chronic social defeat associated with decreases in HDAC2 levels. Interestingly, post-mortem analysis of brain tissue from depressed patients indicates increases in H3-K14 acetylation and decreased HDAC2 levels similar to those observed in socially defeated mice (Covington et al., 2009
), suggesting that there may be an environmentally induced-epigenetic substrate associated with mood disorders in humans.
It is important to note that studies exploring the relationship between social experiences, epigenetic variation, gene expression, and behavioral outcomes have relied primarily on a target gene approach, rather than exploring the global effects of these manipulations. Given the very broad influences of social experiences occurring at different developmental time-points that are illustrated even amongst the subset of targets we have explored in this review, there are likely multiple systems that are regulated through DNA methylation and histone modifications. Investigation of epigenetic regulation within these systems may yield important new insights into the processes through which long-term variations in phenotype are achieved.