Bioinformatic analysis of long-lasting transcriptional and translational changes in the basolateral amygdala following acute stress

Abstract

Stress profoundly impacts the brain and increases the risk of developing a psychiatric disorder. The brain's response to stress is mediated by a number of pathways that affect gene expression and protein function throughout the cell. Understanding how stress achieves such dramatic effects on the brain requires an understanding of the brain's stress response pathways. The majority of studies focused on molecular changes have employed repeated or chronic stress paradigms to assess the long-term consequences of stress and have not taken an integrative genomic and/or proteomic approach. Here, we determined the lasting impact of a single stressful event (restraint) on the broad molecular profile of the basolateral amygdala complex (BLC), a key brain region mediating emotion, memory and stress. Molecular profiling performed thirty days post-restraint consisted of small RNA sequencing, RNA sequencing and quantitative mass spectrometry and identified long-lasting changes in microRNA (miRNA), messenger RNA (mRNA) and proteins. Alignment of the three datasets further delineated the regulation of stress-specific pathways which were validated by qPCR and Western Blot analysis. From this analysis, mir-29a-5p was identified as a putative regulator of stress-induced adaptations in the BLC. Further, a number of predicted mir-29a-5p targets are regulated at the mRNA and protein level. The concerted and long-lasting disruption of multiple molecular pathways in the amygdala by a single stress event is expected to be sufficient to alter behavioral responses to a wide array of future experiences, including exposure to additional stressors.

The Acute Stress Response in the Multiomic Era

“…Transcriptomics has been a mainstay technique in neuroscience labs for many years, and several studies have shown that acute stress leads to robust changes in gene expression in the hippocampus (2,58,85,(96)(97)(98) and amygdala (99,100).…”

microRNA mir-598-3p mediates susceptibility to stress-enhancement of remote fear memory

“…Total RNA from fresh frozen bilateral tissue punches was obtained using the miRVANA PARIS RNA extraction kit (Life Technologies, Carlsbad, CA), as previously reported [5,48]. cDNA libraries of miRNAs were created from 20ng of total RNA with the mirCURY LNA RT Kit (Qiagen, Germantown, MD).…”

“…Spectrometry and Proteomics Resource Laboratory as previously described [5] and data was mined from a quantitative mass spectrometry run on BLA tissue collected from male mice 30 days post-SELF training (reported in Sillivan et al, resubmitted). Candidate proteins were identified as those that had at least 0.5 log2 fold change between treatment groups.…”

“…Additionally, memories that drive unwanted flashbacks of trauma in PTSD are unique from normal episodic memories in that they are intrusive and more life-like in nature [4]. Stress, even one acute episode, can impose lasting changes on the brain [5][6][7]. But how stress-induced changes influence or interact with remote memory mechanisms is not wellcharacterized.…”

“…One goal of the experiments reported here is to gain a better understanding of the role of miRNAs in the persistence of remote memory. We focused our analyses on the basolateral amygdala (BLA) because of the known contribution of this region to fear learning and memory storage at remote time points [16][17][18][19] We recently demonstrated that exposure to a single acute stressor (120 minutes of restraint stress) induces a change in the molecular profile of the BLA, including differential expression of 18 miRNAs an entire month after stress [5]. Differential miRNA expression has also been reported in the serum of patients with PTSD [20] [21] and between rats selectively bred for high or low stress reactivity [22].…”