The functional cooperation of 5−HT1A5-HT_{1A} and mGlu4R in HEK-293 cell line

Background The serotonin 5-HT1A receptor (5-HT1AR) and metabotropic glutamate receptor 4 (mGlu4) have been implicated as sites of antipsychotic drug action. 5-HT1AR belongs to the A class of G protein-coupled receptors (GPCRs); mGlu4 is a representative of class C GPCRs. Both receptors preferentially couple with Gi protein to inhibit cAMP formation. The present work aimed to examine the possibility of mGlu4 and 5-HT1A receptor cross-talk, the phenomenon that could serve as a molecular basis of the interaction of these receptor ligands observed in behavioral studies. Methods First, in vitro studies were performed to examine the pharmacological modulation of interaction of the mGlu4 and 5-HT1A receptors in the T-REx 293 cell line using SNAP- or HALO-tag and cAMP accumulation assay. Next, the colocalization of these two receptors was examined in some regions of the mouse brain by applying RNAScope dual fluorescence in situ hybridization, immunohistochemical labeling, and proximity ligation assay (PLA). Results The ex vivo and in vitro results obtained in the present work suggest the existence of interactions between mGlu4 and 5-HT1A receptors. The changes were observed in cAMP accumulation assay and were dependent on expression and activation of mGlu4R in T-REx 293cell line. Moreover, the existence of spots with proximity expression of both receptors were showed by PLA, immunofluorescence labeling and RNAscope methods. Conclusion The existence of interactions between mGlu4 and 5-HT1A receptors may represent another signaling pathway involved in the development and treatment psychiatric disorders such as schizophrenia or depression

Habenula as a possible target for treatment-resistant depression phenotype in Wistar Kyoto rats

The mechanisms of treatment-resistant depression (TRD) are not clear and are difficult to study. An animal model resembling human TRD is the Wistar Kyoto rat strain. In the present study, we focused on selecting miRNAs that differentiate rats of the WKY strain from Wistar Han (WIS) rats in two divisions of the habenula, the lateral and medial (LHb and MHb, respectively). Based on our preliminary study and literature survey, we identified 32 miRNAs that could be potentially regulated in the habenula. Six miRNAs significantly differentiated WKY rats from WIS rats within the MHb, and three significantly differentiated WKY from WIS rats within the LHb. Then, we selected relevant transcripts regulated by those miRNAs, and their expression in the habenular nuclei was investigated. For mRNAs that differentiated WKY rats from WIS rats in the MHb (Cdkn1c, Htr7, Kcnj9, and Slc12a5), their lower expression correlated with a higher level of relevant miRNAs. In the LHb, eight mRNAs significantly differentiated WKY from WIS rats (upregulated Htr4, Drd2, Kcnj5, and Sstr4 and downregulated Htr2a, Htr7, Elk4, and Slc12a5). These data indicate that several important miRNAs are expressed in the habenula, which differentiates WKY rats from WIS rats and in turn correlates with alterations in the expression of target transcripts. Of particular note are two genes whose expression is altered in WKY rats in both LHb and MHb: Slc12a5 and Htr7. Regulation of KCC2 via the 5-HT7 receptor may be a potential target for the treatment of TRD

Analysis of region-specific changes in gene expression upon treatment with citalopram and desipramine reveals temporal dynamics in response to antidepressant drugs at the transcriptome level

RATIONALE: The notion that the onset of action of antidepressant drugs (ADs) takes weeks is widely accepted; however, the sequence of events necessary for therapeutic effects still remains obscure. OBJECTIVE: We aimed to evaluate a time-course of ADs-induced alterations in the expression of 95 selected genes in 4 regions of the rat brain: the prefrontal and cingulate cortices, the dentate gyrus of the hippocampus, and the amygdala. METHODS: We employed RT-PCR array to evaluate changes during a time-course (1, 3, 7, 14, and 21 days) of treatments with desipramine (DMI) and citalopram (CIT). In addition to repeated treatment, we also conducted acute treatment (a single dose of drug followed by the same time intervals as the repeated doses). RESULTS: Time-dependent and structure-specific changes in gene expression patterns allowed us to identify spatiotemporal differences in the molecular action of two ADs. Singular value decomposition analysis revealed differences in the global gene expression profiles between treatment types. The numbers of characteristic modes were generally smaller after CIT treatment than after DMI treatment. Analysis of the dynamics of gene expression revealed that the most significant changes concerned immediate early genes, whose expression was also visualized by in situ hybridization. Transcription factor binding site analysis revealed an over-representation of serum response factor binding sites in the promoters of genes that changed upon treatment with both ADs. CONCLUSIONS: The observed gene expression patterns were highly dynamic, with oscillations and peaks at various time points of treatment. Our study also revealed novel potential targets of antidepressant action, i.e., Dbp and Id1 genes. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00213-012-2714-0) contains supplementary material, which is available to authorized users

The impact of noradrenergic neurotoxin DSP-4 and noradrenaline transporter knockout (NET-KO) on the activity of liver cytochrome P450 3A (CYP3A) in male and female mice

Background Our earlier studies have shown that the brain noradrenergic system regulates cytochrome P450 (CYP) in rat liver via neuroendocrine mechanism. In the present work, a comparative study on the effect of intraperitoneal administration of the noradrenergic neurotoxin DSP-4 and the knockout of noradrenaline transporter (NET-KO) on the CYP3A in the liver of male and female mice was performed. Methods The experiments were conducted on C57BL/6J WT and $NET^{–/–}$ male/female mice. DSP-4 was injected intraperitoneally as a single dose (50 mg/kg ip.) to WT mice. The activity of CYP3A was measured as the rate of 6β-hydroxylation of testosterone in liver microsomes. The CYP3A protein level was estimated by Western blotting. Results DSP-4 evoked a selective decrease in the noradrenaline level in the brain of male and female mice. At the same time, DSP-4 reduced the CYP3A activity in males, but not in females. The level of CYP3A protein was not changed. The NET knockout did not affect the CYP3A activity/protein in both sexes. Conclusions The results with DSP-4 treated mice showed sex-dependent differences in the regulation of liver CYP3A by the brain noradrenergic system (with only males being responsive), and revealed that the NET knockout did not affect CYP3A in both sexes. Further studies into the hypothalamic–pituitary–gonadal hormones in DSP-4 treated mice may explain sex-specific differences in CYP3A regulation, whereas investigation of monoaminergic receptor sensitivity in the hypothalamic/pituitary areas of $NET^{–/–}$ mice will allow for understanding a lack of changes in the CYP3A activity in the NET-KO animals

Restraint Stress in Mice Alters Set of 25 miRNAs Which Regulate Stress- and Depression-Related mRNAs

In the present study, we aim to identify the effect of restrain stress (RS) on the expression of miRNAs in mouse serum. We used three genotypes of animals (mice with knock-out of the gene-encoding norepinephrine transporter, NET-KO; C57BL/6J, and SWR/J) which had previously been shown to display different sensitivity to RS, and focused on miRNAs which were altered by RS in the serum of all three genotypes. An analysis of miRNAs expression allowed for the identification of a set of 25 differentially expressed miRNAs; 10 were down-regulated compared to an appropriate control group of animals, while 15 were up-regulated. The application of DIANA-miRPath v. 3.0 allowed for the identification of selected pathways (KEGG) and Gene Ontology (GO) categories that were significantly controlled by these miRNAs, while miRWalk v. 3.0—the platform that used the machine learning based algorithm, TaRPmiR—was used to find their targets. The results indicate that 25 miRNAs, identified as altered upon RS in three genotypes of mice, are responsible for regulation of mRNA-encoding proteins that are key for the main hypotheses of depression; therefore, they may help to understand the link between stress and depression at the molecular level

What Do the Animal Studies of Stress Resilience Teach Us?

Long-lasting stress factors, both biological and psychological, are commonly accepted as the main cause of depressive disorders. Several animal models, using various stressful stimuli, have been used to find biochemical and molecular alterations that could help us understand the etiopathogenesis of depression. However, recent sophisticated studies indicate that the most frequently used animal models of stress only capture a portion of the molecular features associated with complex human disorders. On the other hand, some of these models generate groups of animals resilient to stress. Studies of the mechanisms of stress resilience bring us closer to understanding the process of adapting to aversive stimuli and the differences between stress-susceptible vs. resilient phenotypes. Especially interesting in this context is the chronic mild stress (CMS) experimental paradigm, most often using rats. Studies using this animal model have revealed that biochemical (e.g., the dopamine D2 receptor) and molecular (e.g., microRNA) alterations are dynamic (i.e., depend on stress duration, 2 vs. 7 weeks) and much more pronounced in stress-resilient than stress-susceptible groups of animals. We strongly suggest that studies aimed at understanding the molecular and biochemical mechanisms of depression must consider these dynamics. A good candidate to serve as a biomarker in such studies might be serum microRNA, since it can be obtained relatively easily from living individuals at various time points

Copyright © 2007 by Institute of Pharmacology Polish Academy of Sciences

Effect of citalopram in the modified forced swim test in rat