Early Life Stress-Induced Epigenetic Programming of Hippocampal NPY-Y2 Receptor Gene Expression Changes in Response to Adult Stress

Abstract

Early Life Stress (ELS) can critically influence brain development and future stress responses and thus represents an important risk factor for mental health and disease. Neuropeptide Y (NPY) is discussed to be a key mediator of resilient vs. vulnerable adaptations and specifically, the NPY-Y2 receptor (Y2R) may be involved in the pathophysiology of depression due to its negative regulation of NPY-release. The present study addressed the hypotheses that ELS and adult stress (AS) affect the expression of hippocampal Y2R and that exposure to ELS induces an epigenetically mediated programming effect towards a consecutive stress exposure in adulthood. The specific aims were to investigate if (i) ELS or AS as single stressors induce changes in Y2 receptor gene expression in the hippocampus, (ii) the predicted Y2R changes are epigenetically mediated via promoter-specific DNA-methylation, (iii) the ELS-induced epigenetic changes exert a programming effect on Y2R gene expression changes in response to AS, and finally (iv) if the predicted alterations are sex-specific. Animals were assigned to the following experimental groups: (1) non-stressed controls (CON), (2) only ELS exposure (ELS), (3) only adult stress exposure (CON+AS), and (4) exposure to ELS followed by AS (ELS+AS). Using repeated maternal separation in mice as an ELS and swim stress as an AS we found that both stressors affected Y2R gene expression in the hippocampus of male mice but not in females. Specifically, upregulated expression was found in the CON+AS group. In addition, exposure to both stressors ELS+AS significantly reduced Y2R gene expression when compared to CON+AS. The changes in Y2R expression were paralleled by altered DNA-methylation patterns at the Y2R promoter, specifically, a decrease in mean DNA-methylation in the CON+AS males compared to the non-AS exposed groups and an increase in the ELS+AS males compared to the CON+AS males. Also, a strong negative correlation of mean DNA-methylation with Y2R expression was found. Detailed CpG-site-specific analysis of DNA-methylation revealed that ELS induced increased DNA-methylation only at specific CpG-sites within the Y2R promoter. It is tempting to speculate that these ELS-induced CpG-site-specific changes represent a “buffering” programming effect against elevations of Y2R expression induced by AS.

Epigenetic (re)programming of gene expression changes of CB1R and FAAH in the medial prefrontal cortex in response to early life and adolescence stress exposure

“…The interpretation of a "(re)programming" effect of ELS is further supported by findings of our previous studies in male mice on hippocampal NPY-Y2 receptor and oxytocin receptor (OxtR) gene expression. Similar to the "buffering" effect of ELS observed in the present study we found increased NPY-Y2 gene expression after a single adult swim stress exposure, whereas animals exposed to both stressors (ELS + AS) displayed expression levels similar to those in unstressed controls (Kocamaz et al, 2022). For OxtR gene expression we found evidence for an "amplifying" effect of ELS: while ELS or AS alone had no effect on gene expression, the consecutive exposure to both ELS and AS resulted in increased gene expression in the hippocampal formation (Lesse et al, 2017).…”

“…Deoxyribonucleic acid methylation levels were determined using the bisulfite-converted DNA and pyrosequencing method (Kocamaz et al, 2022). In short, genomic DNA was extracted from left mPFC using DNeasy Blood and Tissue Kit (Qiagen GmbH, Hilden, Germany) according to the manufacturer's instructions.…”

“…Guided by the "two-(or multiple) hit" concept (Daskalakis et al, 2013;Gruber et al, 2015;Lesse et al, 2017;Kocamaz et al, 2022) we hypothesized that ELS might influence the observed upregulation of CB1R and FAAH gene expression in response to FS exposure during adolescence. Our and other studies revealed that vulnerable as well as resilient adaptations may occur depending on the time point of stress exposure, and that the presumed "(re)programming" effects of the first stressor may either amplify or ameliorate/suppress the cellular and molecular responses toward subsequent stressors (Nederhof and Schmidt, 2012;Daskalakis et al, 2013;McElroy and Hevey, 2014).…”

“…Among the different epigenetic modifications such as histone modifications, microRNAs and DNA methylation, the latter is one of the best-described mechanisms with respect to epigenetic changes induced by early adverse experiences (Park et al, 2014;Sasagawa et al, 2017;Zhang et al, 2019;Kocamaz et al, 2022). This epigenetic modification represents a crucial regulator for the fine-tuning of gene expression and the classical view is that enhanced DNA-methylation (hypermethylation) leads to decreased or even silenced gene expression (Bird, 2002;Greenberg and Bourc'his, 2019).…”

“…However, the mechanisms of these complex interactions and how they may promote vulnerable or resilient adaptations are far from understood in detail. There is increasing evidence that epigenetic mechanisms may act as mediators between genetically programmed developmental processes and potentially (re-)programming early environmental influences (Bock et al, 2014;Köhler et al, 2019;Kocamaz et al, 2022;Rahman and McGowan, 2022). Thus, they are discussed to play a crucial role in translating early adverse experiences into long-lasting gene expression changes and thereby can modulate stress-related responses (Bock et al, 2014;Provençal and Binder, 2015;Radtke et al, 2015;Alyamani and Murgatroyd, 2018;Li et al, 2020).…”