Transcriptome Dynamics in Mouse Amygdala under Acute and Chronic Stress Revealed by Thiol-labeled RNA Sequencing
Both acute and chronic stress have significant impact on brain functions. The amygdala is essential in mediating stress responses, but how its transcriptomic dynamics change under stress remains elusive. To overcome the difficulties in detecting subtle stress-induced changes by evaluating total RNA using classic RNA sequencing, we conducted thiol-labeled RNA sequencing (SLAM-seq). We injected 4-thiouridine (4sU) into mouse amygdala followed by SLAM-seq to detect nascent mRNA induced by acute and chronic restraint stress, and found that SLAM-seq could label actively transcribed genes in the major neuronal and glial subtypes. Using SLAM-seq, we found that chronic stress led to higher turnover of a group of genes associated with myelination, and this finding is confirmed by immunostaining which showed increased myelination in the chronically stressed amygdala. Additionally, genes detected by SLAM-seq and RNA-seq only partially overlapped, with SLAM-seq particularly sensitive to transcriptional changes in genes with high basal transcription levels. Thus, by applying SLAM-seq in vivo, we obtained a rich dataset of nascently transcribed genes in the amygdala under stress, and revealed distinct transcriptional dynamics associated with acute and chronic stress. Overall design: To investigate transcriptomic dynamics of amygdala under stress, we established an in vivo SLAM-seq protocol by injecting 4-thiouridine (4sU) or DMSO into mouse amygdala, and subjected mice to two stress conditions. We then performed SLAM-seq analysis (T-C conversion rates for all genes detected) and gene expression profiling analysis using data obtained from RNA-seq of 2 different stress conditions and 1 no-stress control condition.