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The expression of the Calcium/Calmodulin-Dependent Protein Kinase I gamma (encoded by the Camk1g gene) depends on the activation of glucocorticoid receptors (GR) and is strongly regulated by stress. Since Camk1g is primarily expressed in neuronal cells of the limbic system in the brain, we hypothesized that it could be involved in signaling mechanisms that underlie the adaptive or maladaptive responses to stress. Here, we find that restraint-induced stress and the GR agonist dexamethasone robustly increase the expression of Camk1g in neurons of the amygdalar nuclei in the mouse brain. To assess the functional role of Camk1g expression, we performed a virally induced knock-down of the transcript. Mice with bilateral amygdala-specific Camk1g knock-down showed increased anxiety-like behaviors in the light-dark box, and an increase in freezing behavior after fear-conditioning, but normal spatial working memory during exploration of a Y-maze. Thus, we confirm that Camk1g is a neuron-specific GR-regulated transcript, and show that it is specifically involved in behaviors related to anxiety, as well as responses conditioned by aversive stimuli.

Three strains of mice with various susceptibilities to restraint stress (RS), i.e., mice with a knocked out norepinephrine transporter gene (NET-KO), SWR/J and C57BL/6J (WT) mice were shown to serve as a good model to study the molecular mechanisms underlying different stress-coping strategies. We identified 14 miRNAs that were altered by RS in the PFC of these mice in a genotype-dependent manner, where the most interesting was let-7e. Further in silico analysis of its potential targets allowed us to identify five mRNAs (Bcl2l11, Foxo1, Pik3r1, Gab1 and Map2k4), and their level alterations were experimentally confirmed. A next-generation sequencing (NGS) approach, which was employed to find transcripts differentially expressed in the PFC of NET-KO and WT mice, showed that, among others, two additional mRNAs were regulated by mmu-let-7e, i.e., mRNAs that encode Kmt2d and Inf2. Since an increase in Bcl2l11 and Pik3r1 mRNAs upon RS in the PFC of WT mice resulted from the decrease in mmu-let-7e and mmu-miR-484 regulations, we postulated that MAPK, FoxO and PI3K-Akt signaling pathways were associated with stress resilience, although via different, genotype-dependent regulation of various mRNAs by let-7e and miR-484. However, a higher level of Kmt2d mRNA (regulated by let-7e) that was found with NGS analysis in the PFC of NET-KO mice indicated that histone methylation was also important for stress resilience.

The glucocorticoid receptor (GR, also known as NR3C1) coordinates molecular responses to stress. It is a potent transcription activator and repressor that influences hundreds of genes. Enhancers are non-coding DNA regions outside of the core promoters that increase transcriptional activity via long-distance interactions. Active GR binds to pre-existing enhancer sites and recruits further factors, including EP300, a known transcriptional coactivator. However, it is not known how the timing of GR-binding-induced enhancer remodeling relates to transcriptional changes. Here we analyze data from the ENCODE project that provides ChIP-Seq and RNA-Seq data at distinct time points after dexamethasone exposure of human A549 epithelial-like cell line. This study aimed to investigate the temporal interplay between GR binding, enhancer remodeling, and gene expression. By investigating a single distal GR-binding site for each differentially upregulated gene, we show that transcriptional changes follow GR binding, and that the largest enhancer remodeling coincides in time with the highest gene expression changes. A detailed analysis of the time course showed that for upregulated genes, enhancer activation persists after gene expression changes settle. Moreover, genes with the largest change in EP300 binding showed the highest expression dynamics before the peak of EP300 recruitment. Overall, our results show that enhancer remodeling may not directly be driving gene expression dynamics but rather be a consequence of expression activation.

Cocaine induces neuronal changes as well as non-neuronal (astrocytes, microglia, oligodendroglia) mechanisms, but these changes can also be modulated by various types of drug abstinence. Due to the very complex and still incompletely understood nature of cocaine use disorder, understanding of the mechanisms involved in addictive behavior is necessary to further search for effective pharmacotherapy of this disease. The aim of this study was to investigate changes at the gene and protein levels associated with glial cell activity after cocaine exposure, as well as during early cocaine abstinence (3 days) with extinction training or in home cage isolation. Cocaine self-administration significantly decreased myelin regulatory factor (MYRF) and cyclic nucleotide phosphodiesterase (CNP) expression in the hippocampus as well as pleckstrin (PLEK) and T-lymphocyte activation antigen (CD86) in the rat striatum. Depending on cocaine abstinence conditions, microglial PLEK expression was increased through extinction training but did not change in the home cage isolation. In addition, downregulation of gene expression associated with oligodendrocytes (CNP, MYRF) and microglia regulator of G protein signaling 1 (RGS1) was observed in the hippocampus, regardless of the type of drug abstinence, while downregulation of myelin and lymphocyte protein (MAL) expression was found only in rats exposed to abstinence in the home cage. Taken together, the presented results strongly suggest that cocaine abstinence evokes significant changes in gene expression associated with the proper functioning of glial cells, suggesting their significant involvement in adaptive changes in the brain associated with cocaine exposure. Interestingly, drug abstinence conditions are important factors influencing observed changes at the transcript levels of selected genes, which may be of clinical interest.