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.

Norepinephrine transporter (NET) knock-out upregulates dopamine and serotonin transporters in the mouse brain.

The noradrenaline, serotonin and dopamine transporters are three main transporters, which are the target of the antidepressant drugs. In the present study we demonstrate that the life-long deletion of the noradrenaline transporter (NET) induced up-regulation of two other monoamine transporters, dopamine and serotonin (DAT and SERT, respectively). An increase in the binding of [(3)H]paroxetine to the SERT and [(3)H]GBR12935 to the DAT was observed in various brain regions of NET-KO mice, without alterations of mRNA encoding these transporters, as measured by in situ hybridization. This important finding impacts the interpretation of previous data indicating the supersensitizity of NET-KO mice for psychostimulants or stronger effect of citalopram in behavioral tests. While using the NET-KO mice in various psychopharmacological studies is very important, one has to be aware that these mice lack NET from the earliest period of their existence, thus compensatory alterations do take place and have to be considered when it comes to interpretation of the obtained results.

Changes in the level of calcyon mRNA in the brain of rats exposed to cocaine, self-administered or received passively.

The level of mRNA encoding calcyon (measured by in situ hybridization), one of the dopamine receptor interacting proteins, has been examined in the rat brain in the established animal model used to study the mechanisms of cocaine addiction (cocaine self-administration involving a yoked procedure). Two weeks of cocaine self-administration (maintenance) did not affect the level of calcyon mRNA, regardless of the way cocaine was delivered, except for tuberculum olfactorium, where calcyon mRNA was increased after cocaine treatment. In the reinstatement phase of the experiment cocaine alone induced an increase in the calcyon mRNA expression in most of the brain region studied (caudate putamen; tuberculum olfactorium; paraventricular thalamic nucleus; ventromedial hypothalamic nucleus and paraventricular hypothalamic nucleus) but only in the yoked saline control group. In other words, these results show that the single dose of cocaine (10 mg/kg) was able to induce an alteration in the level of calcyon mRNA in these rats which never before experienced any cocaine administration. The most significant effects were observed in the ventromedial hypothalamic nucleus and paraventricular hypothalamic nucleus. Interestingly, a similar effect was observed when the reinstatement of cocaine-seeking behaviour was evoked by cue (conditioned stimuli) that indicates that no cocaine was necessary to induce the changes in the level of calcyon mRNA expression. This effect was significant in tuberculum olfactorium, ventromedial hypothalamic nucleus and paraventricular hypothalamic nucleus. Such a result together with the brain areas involved in these effects might suggest the role of calcyon similar to the CART peptides and special vulnerability of calcyon expression rather to acute than chronic stimuli.

[Fluorescence studies of G-protein coupled receptors oligomerization]. / Fluorescencyjne metody badania oligomeryzacji receptorów zwiazanych z bialkami G.

Fluorescence methods based on the resonance energy transfer phenomenon are currently extensively used to study biological processes, particularly interactions of G-protein coupled receptors. The development of molecular engineering has made it feasible to produce fluorescently tagged fusion proteins, which constitute a model system for studies of receptors oligomerization. It is widely accepted, that physical interactions between receptors might be of great importance for development of potential therapeutic factors, which influence signal transduction across the membrane. Here, we report on a variety of fluorescence approaches commonly used to investigate receptors oligomerization. Comparative studies reveal that even a relatively simple method, such as steady-state fluorescence spectroscopy could be useful in verifying receptor interactions. However, more sophisticated techniques as confocal microscopy and fluorescence lifetime microscopy not only provide quantifiable data of high relevance but also allow observations of dynamic processes in complex biological systems.