Mouse experiments reveal how early-life stress may leave a lasting epigenetic imprint in dopamine-related brain circuitry, priming molecular, neuronal, and behavioral responses to stress later in life.
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In a recent study published in the journal Neuron, researchers investigated whether childhood adversity (‘early-life stress’ [ELS]) can drive epigenetic mechanisms that modulate lasting sensitivity to subsequent stress, with potential relevance to psychiatric vulnerability.
Study data (ventral tegmental area [VTA] histone modifications) were initially obtained using high-throughput mass spectrometry in a murine model. Subsequently, viral epigenome editing, transcriptomic profiling, patch-clamp electrophysiology, and behavioral observations were used to elucidate the outcomes of ELS-driven epigenetic modulations of later transcriptional, neurophysiological, and stress-related behavioral responses.
Study findings revealed that ELS enriches histone-3 lysine-4 monomethylation (H3K4me1) and the H3K4 monomethyltransferase “SETD7” in the VTA of mouse brains. Furthermore, the study established that bidirectional manipulation of Setd7 in juvenile mice showed that Setd7/H3K4me1 enrichment can be sufficient to heighten transcriptional, physiological, and behavioral responses to adult stress, while Setd7 knockdown demonstrated the necessity for key physiological and behavioral effects of ELS.
These findings identify a biological mechanism through which early-life stress may encode lasting stress hypersensitivity, and they may help inform future research on approaches to promote resilience following childhood adversity.
Background
A growing body of psychiatric evidence indicates that early-life (‘childhood’) adversity represents a major environmental risk factor for the development of depression, anxiety, and substance use disorders later in life. Clinical and preclinical evidence indicate that early-life adversity is associated with lasting alterations in neural reward circuitry, particularly in the midbrain ventral tegmental area (VTA).
Correlational evidence further suggests that these altered neural reward circuits can prime individuals to exhibit heightened sensitivity to subsequent adult stressors. Unfortunately, while previous research identified latent gene expression changes in mesocorticolimbic circuits following early stress, causal evidence for the precise chromatin-level mechanisms that maintain this transcriptional memory remained hitherto elusive.
About the study
The present study aimed to address this neurobiological gap and inform future psychiatric research by characterizing the chromatin landscape of the VTA following early-life stress and determining whether epigenetic modifications prime or enhance adult stress sensitivity.
The experimental research was conducted on murine (C57BL/6J mice) models of early-life stress (ELS), which were subjected to early-life (postnatal days P10-P17) 3-4-hour maternal separation periods and reduced nesting material.
Resultant epigenetic shifts in adult male VTA tissue were mapped using unbiased bottom-up liquid chromatography-tandem mass spectrometry (LC-MS/MS) to quantify individual and combinatorial histone modifications. These analyses identified H3K4me1 among the altered histone marks, while subsequent gene-expression analyses identified Setd7 as a prominently altered histone-modifying enzyme.
Subsequently, the study isolated the functional consequences of SETD7 modifications using viral vectors (AAV9) that were engineered for juvenile Setd7 overexpression (Setd7-OE) or neuronally expressed short hairpin RNA (shRNA)-mediated knockdown (Setd7-KD). Importantly, the overexpression construct used a ubiquitous promoter and was not specific to dopamine neurons.
The consequences of these Setd7 manipulations were evaluated in both male and female C57BL/6J mice across three domains: 1. Bulk RNA-sequencing (RNA-seq) was used to track transcriptional reactivity to adult social defeat stress, 2. Whole-cell patch-clamp electrophysiology was leveraged to measure VTA dopamine neuron excitability and hyperpolarization-activated (Ih) currents following acute variable stress, and 3. Behavioral testing, which comprised social-interaction and open-field testing, was conducted to determine whether Setd7 overexpression could mimic ELS-associated stress sensitivity and whether Setd7 knockdown could reduce it.
Study findings
LC-MS/MS results confirmed that ELS significantly altered histone tail dynamics in the adult VTA, altering numerous histone modifications, many of which were associated with permissive or open chromatin states, with H3K4me1 selected for mechanistic follow-up because of its association with open or primed chromatin and the accompanying alteration in SETD7. The authors reported that 75% of the histone modification changes were associated with permissive, open, active, primed, or poised chromatin states.
Concurrent gene expression screening revealed that Setd7 messenger RNA (mRNA) was increased in adult VTA tissue, while immunohistochemistry showed increased nuclear SETD7 protein in VTA dopamine neurons by postnatal day P21 following ELS. Setd7 manipulation experiments revealed that juvenile murine Setd7 overexpression increased VTA H3K4me1 levels by 34% (P < 0.0001) and augmented transcriptional reactivity to adult stress.
Specifically, in green fluorescent protein (GFP) controls exposed to adult stress, 72% of differentially expressed genes were downregulated, whereas 94% of genes differing between adult-stressed Setd7-OE and adult-stressed GFP mice were upregulated.
The authors also found that the transcriptional response produced by Setd7-OE differed from that previously observed after ELS, suggesting that SETD7/H3K4me1 represents only one component of the mechanisms through which ELS sensitizes responses to later stress.
Electrophysiological experiments revealed that while Setd7-OE did not alter baseline firing, it significantly heightened dopamine neuron excitability and increased depolarizing Ih currents following adult-life stress; the Ih-current comparison was significant at P = 0.023.
Behavioral evaluations showed a parallel increase in stress susceptibility, with Setd7-OE substantially increasing the proportion of stress-susceptible mice from 8.3% to 50% (P = 0.003).
Conversely, knocking down Setd7 in juvenile VTA was observed to reduce H3K4me1 levels by 37% (P = 0.0233). In ELS-exposed mice, Setd7-KD prevented stress-induced hyperexcitability in dopamine neurons, concurrently blocking the ELS-associated increase in Ih currents (P = 0.015), and reduced adult stress susceptibility from 85% to 33% (P = 0.016).
Together, these findings identify SETD7/H3K4me1 as a mechanistic target for future investigation, but do not establish gene therapy or another SETD7-targeted intervention as a viable treatment in humans.
Conclusions
The present study provides causal evidence linking the ELS-induced enrichment of SETD7 and H3K4me1 in the VTA, including SETD7 enrichment in dopaminergic neurons, to the epigenetic priming of lasting stress sensitivity in mice.
While the authors note several methodological limitations, including the non-cell-type-specific promoter used in the overexpression vector, viral expression spread into adjacent midbrain nuclei, and the reliance on different adult stress paradigms across electrophysiological and behavioral assays, the study’s findings indicate that more precise investigation of locus-specific chromatin modifications and SETD7 activity may help determine whether this mechanism can eventually inform approaches to promoting resilience following childhood adversity.
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