In the present study, we used three strains of mice with various susceptibility to stress: mice with knock-out of the gene encoding norepinephrine transporter (NET-KO), which are well characterized as displaying a stress-resistant phenotype, as well as two strains of mice displaying two different stress-coping strategies, i.e., C57BL/6J (WT in the present study) and SWR/J. The procedure of restraint stress (RS, 4 h) was applied, and the following behavioral experiments (the forced swim test and sucrose preference test) indicated that NET-KO and SWR/J mice were less sensitive to RS than WT mice. Then, we aimed to find the miRNAs which changed in similar ways in the serum of NET-KO and SWR/J mice subjected to RS, being at the same time different from the miRNAs found in the serum of WT mice. Using Custom TaqMan Array MicroRNA Cards, with primers for majority of miRNAs expressed in the serum (based on a preliminary experiment using the TaqMan Array Rodent MicroRNA A + B Cards Set v3.0, Thermo Fisher Scientific, Waltham, MA, USA) allowed the identification of 21 such miRNAs. Our further analysis focused on miR-1 and miR-155 and their targets-these two miRNAs are involved in the regulation of BDNF expression and can be regarded as biomarkers of stress-resilience.Cells 2020, 9, 917 2 of 17 are regarded as endogenous "hubs" (as Issler and Chen have put it) for the fine tuning of target gene expression or an "expression switch", and can provide deeper insight into complex biological processes underlying stress response.In the present study, we used three strains of mice differentially reacting to stress: mice with knock-out of the gene encoding norepinephrine transporter (NET-KO), which are well characterized as displaying a stress-resistant phenotype [7-9], as well as two other strains of mice displaying different stress-coping strategies, i.e., C57BL/6J (WT in the present study) and swiss SWR/J, described extensively by Szklarczyk and co-workers [10]. The different reaction to stress of these strains of mice has been confirmed by measuring corticosterone level. The NET-KO mice display a slower increase in corticosterone level after stress compared with WT animals, while corticosterone level was not changed at 2 h after restraint stress in the blood of SWR/J mice [8,10]. We applied the procedure of restraint stress (RS), which has been shown to induce an uncontrollable aversive situation that produces both physical and psychological consequences leading to neuronal and behavioral alterations [11]. The aim of following molecular studies was to find biomarkers for different behavioral responses to RS among miRNAs expressed in the serum of three genotypes under study. This goal was achieved, as we were able to identify a group of miRNAs differentiating the stress response of NET-KO and SWR/J mice from WT animals. The most interesting were the alterations in the miR-1 and miR-155 levels, which are involved in regulation of BDNF expression, which in turn affects important biochemical processes.
Materials and Methods
AnimalsH...