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Open AccessArticle

Transcriptome Changes in Three Brain Regions during Chronic Lithium Administration in the Rat Models of Mania and Depression

1
Department of Animal Physiology, Biochemistry and Biostructure, Poznan University of Life Sciences, 60-637 Poznan, Poland
2
Department of Histology and Embryology, Poznan University of Medical Sciences, 60-781 Poznan, Poland
3
Department of Psychiatric Genetics, Poznan University of Medical Sciences, 60-806 Poznan, Poland
4
Laboratory of Neurobiology, Department of Molecular and Cellular Neurobiology, Nencki Institute, 02-093 Warsaw, Poland
5
Molecular and Cell Biology Unit, Poznan University of Medical Sciences, 60-572 Poznan, Poland
6
Department of Medical Genetics, Poznan University of Medical Sciences, 60-806 Poznan, Poland
7
Department of Adult Psychiatry, Poznan University of Medical Sciences, 60-572 Poznan, Poland
*
Authors to whom correspondence should be addressed.
Academic Editor: Anastasios Lymperopoulos
Int. J. Mol. Sci. 2021, 22(3), 1148; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms22031148
Received: 20 December 2020 / Revised: 20 January 2021 / Accepted: 22 January 2021 / Published: 24 January 2021
(This article belongs to the Special Issue Molecular Mechanisms of Mood Stabilizers)

Abstract

Lithium has been the most important mood stabilizer used for the treatment of bipolar disorder and prophylaxis of manic and depressive episodes. Despite long use in clinical practice, the exact molecular mechanisms of lithium are still not well identified. Previous experimental studies produced inconsistent results due to different duration of lithium treatment and using animals without manic-like or depressive-like symptoms. Therefore, we aimed to analyze the gene expression profile in three brain regions (amygdala, frontal cortex and hippocampus) in the rat model of mania and depression during chronic lithium administration (2 and 4 weeks). Behavioral changes were verified by the forced swim test, open field test and elevated maze test. After the experiment, nucleic acid was extracted from the frontal cortex, hippocampus and amygdala. Gene expression profile was done using SurePrint G3 Rat Gene Expression whole transcriptome microarrays. Data were analyzed using Gene Spring 14.9 software. We found that chronic lithium treatment significantly influenced gene expression profile in both mania and depression models. In manic rats, chronic lithium treatment significantly influenced the expression of the genes enriched in olfactory and taste transduction pathway and long non-coding RNAs in all three brain regions. We report here for the first time that genes regulating olfactory and taste receptor pathways and long non-coding RNAs may be targeted by chronic lithium treatment in the animal model of mania.
Keywords: manic-like behavior; depressive-like behavior; lithium; animal model; transcriptome; brain manic-like behavior; depressive-like behavior; lithium; animal model; transcriptome; brain

1. Introduction

Bipolar disorder (BD) is a severe, recurrent psychiatric condition characterized by episodes of mania or hypomania and depression. Mania presents with hyperactivity, disinhibited behavior and inflated self-esteem. On the other hand, during depression patients demonstrate anhedonia and loss of motivation and interest in usual activities [1].
Pharmacological treatment of BD includes mostly the use of mood stabilizers. The first drug fulfilling criteria for the mood stabilizer such as preventing manic and depressive episodes was lithium. Its antimanic actions in the acute episode were first discovered by Cade et al. in animal model studies [2]. Its antidepressant potential was reported in the 1970s [3,4] and later it was recommended as an augmentation of antidepressants in the treatment-resistant depression and an effective long-term prophylaxis of recurrences in mood disorders (as recently reviewed by Rybakowski [5]).
Although lithium has been for decades in clinical use, the molecular mechanism of action and normotymic potential of lithium are still not well identified. Moreover, absence of suitable animal model of bipolar disorder impedes studies on lithium action. Previous studies focused on identifying target genes and molecular pathways of lithium indicated its role in reducing inositol signaling [6], phosphorylation of AKT [7] and inhibition of GSK-3β (glycogen synthase kinase 3) signaling [6,8]. The latter two may lead to the changes in the activation of WNT pathway. These findings were possible to discover using pharmacological and pharmacogenetic studies, which showed involvement of lithium in multiple cellular signaling pathways as reviewed previously [9,10].
Animal studies of the brain transcriptome during lithium administration showed that inconsistent results are mainly due to the different duration of lithium administration (1 week, two weeks and 6 weeks), analysis of the whole brain tissue (whole brain homogenates) and the use of different animals (rat or mouse) [11,12]. Moreover, these studies used animals treated with lithium, but without manic-like or depressive-like symptoms, so it is still not known if therapeutic action of lithium in manic and depressive behavior induces changes in the gene expression of similar or different pathways depending on the baseline condition and the brain region.
Therefore, we hypothesized that chronic lithium treatment exerts its therapeutic effect by inducing changes in the brain transcriptome specific for mania and depression. The aim of this study was to investigate if chronic lithium administration in rats presenting manic-like behavior (induced by amphetamine) or depressive like behavior (induced by chronic mild stress) influences specific gene expression profiles in different brain regions (amygdala, frontal cortex and hippocampus).

2. Results

2.1. Behavioral Changes in Amphetamine-Exposed and Stress-Exposed Animals (Models of Mania and Depression)

Manic-like behavior was observed after one week of amphetamine injections using the elevated maze test as compared to the behavior of the same animals before starting the experiment (baseline) (Figure 1).
Two-week chronic mild stress protocol resulted in depressive-like behavior measured by behavioral tests (FST and OFT) in analyzed rats as compared to the behavior of the same animals before starting the experiment (baseline) (Figure 2).

2.2. Brain Transcriptome Changes in the Model of Mania and Depression

Significant changes in the behavior following chronic mild stress protocol or amphetamine corresponded to specific gene expression profiles in three brain regions. In rats presenting manic-like behavior, we found 1283 genes differentially expressed in the amygdala, 637 genes in the frontal cortex and 344 genes in the hippocampus as compared to the control group (Figure 3a,b). The altered genes in the amygdala were mainly downregulated and significantly enriched in 23 Gene Ontology (GO) terms including olfactory receptor activity and chemical response to stimuli (Table 1). The genes differentially expressed in the frontal cortex and hippocampus were not significantly enriched in any GO terms or pathways.
In rats with depressive-like behavior, we found 856 differentially expressed genes (either up- or downregulated) in the amygdala, 1910 genes in the frontal cortex and 1942 genes in the hippocampus as compared to the control rats (Figure 3c,d).
The downregulated genes in the amygdala were enriched mainly in locomotor behavior and blood circulation (8 GO terms), whereas the upregulated genes were involved, among others, in animal organ morphogenesis, response to stimuli and signal transduction (8 GO terms) (Table 2). In the frontal cortex and hippocampus the differentially expressed genes (either upregulated or downregulated) were not significantly enriched in any GO terms or pathways.

2.3. Brain Transcriptome Changes during Chronic Lithium Administration

2.3.1. Two-Week Lithium Administration

Lithium administration for two weeks resulted in significant changes in both, stress-exposed and manic-like rats in all analyzed brain regions. In rats presenting manic-like behavior and receiving lithium for two weeks, we found 4041 differentially expressed genes in the amygdala (all upregulated), 978 genes in the frontal cortex and 386 genes in the hippocampus. In the amygdala upregulated genes were enriched in 40 GO terms, mainly olfactory receptor activity, taste receptor and bitter taste receptor (Table 3). In the frontal cortex, most genes were significantly downregulated and involved in 11 GO terms, including olfactory receptor activity (Table 3). Genes differentially expressed in the hippocampus were not enriched in any GO terms.
Rats presenting depressive-like behavior showed differentially expressed 1650 genes in the amygdala, 481 genes in the frontal cortex and 304 genes in the hippocampus. The significantly altered genes in the amygdala were mainly downregulated and were involved in 206 GO terms, mainly associated with extracellular processes e.g., extracellular matrix organization (top 20 were listed in Table 4). In the frontal cortex differentially expressed genes were not significantly enriched in any GO terms between rats receiving lithium and control group. In the hippocampus, altered genes were significantly enriched in two GO terms, antigen processing and antigen binding (Table 4).

2.3.2. Four-Week Lithium Administration

In rats presenting manic-like behavior and receiving lithium for 4 weeks, we observed 2831 genes differentially expressed in the amygdala, 1964 genes in the frontal cortex and 7412 genes in the hippocampus. In the amygdala, all genes were downregulated after 4 weeks on lithium and were enriched mainly in olfactory receptor activity and bitter taste (Table 5). In the frontal cortex, downregulated genes were enriched in 31 GO terms including, e.g., olfactory receptor activity (Table 5). In the hippocampus significantly upregulated genes were enriched in 57 GO terms (mainly response to chemical stimuli, bitter taste and olfactory receptor activity) (Table 5).
Four weeks of lithium treatment in stress-exposed rats, we observed that in the amygdala 778 genes were differentially expressed, in the frontal cortex 779 genes and in the hippocampus 307 genes. In the amygdala the genes were mainly upregulated and enriched in 17 GO terms, e.g., olfactory receptor activity (Table 6), in the frontal cortex altered genes were grouped in 24 GO terms (including nucleosome assembly, DNA-protein complex) and in the hippocampus differentially expressed genes were involved in extracellular matrix/space/region and collagen fibril organization (two upregulated GO terms) (Table 6).

2.3.3. Molecular Pathways in the Brain during Chronic Lithium Treatment

When we combined together gene sets differentially expressed in rats with depressive-like behavior receiving lithium for two and four weeks, we observed only a few shared genes between these two time points either in the amygdala or frontal cortex and hippocampus as compared to the control rats (Figure 4). These genes were not enriched in any GO terms and pathways.
In rats with manic-like behavior, we observed that chronic lithium treatment (2 and 4 weeks) significantly influenced the expression of the genes from the olfactory and taste transduction pathway (mainly different olfactory and taste receptors) and long non-coding RNAs in all three brain regions in rats receiving lithium as compared to water-receiving animals (Figure 5). Gene-set enrichment analysis showed that the shared genes between 2 and 4 weeks showed different gene expression profile: those upregulated after 2-week lithium treatment were downregulated after 4 weeks in the amygdala and frontal cortex. In the hippocampus, a 4-week lithium treatment upregulated all the genes.

2.4. Brain Transcriptome Changes in Lithium Responders

We also compared the gene expression profile before and after chronic lithium administration (2 and 4 weeks) in lithium responders. These rats showed changes in behavioral tests (open field test and elevated maze test) corresponding to reduced manic or depressive symptoms, e.g., reduced number of visits and time spent in open arms in manic rats after lithium and increased exploration time and number of line crossing in depressive rats after lithium (Figure 6).
Gene expression analysis in rats responding to lithium showed that in after chronic lithium administration gene expression was mainly upregulated in the amygdala and frontal cortex, but downregulated in the hippocampus in manic rats responding to lithium as compared to rats before lithium treatment. Interestingly, depressive rats responding to lithium showed the opposite trend, with mostly downregulated gene expression in the amygdala and frontal cortex, but upregulated in the hippocampus as compared to depressive rats before lithium. These genes were significantly enriched in GO terms related i.a. to olfactory receptor pathway, sensory perception of smell, G-protein coupled receptor and detection of stimulus (Table 7 and Table 8).

3. Discussion

The main observation of this study shows that the brain transcriptome is influenced by chronic lithium administration. The important finding is that the differentially expressed genes specific for the mania model are involved in olfactory receptor transduction and taste receptor pathways and show altered expression of long non-coding RNA genes.
Comparing depressive-like and manic-like animals, we found that in the depression model, the gene expression profile is more region-specific (different pathways regulated upon lithium in the amygdala, frontal cortex and hippocampus) and time-dependent (2 weeks versus 4 weeks). In the model of mania, on the other hand, lithium significantly affected olfactory receptor and bitter taste receptor pathways, independent of the brain region or time of lithium administration suggesting their role in antimanic action. The genes from these pathways also showed significantly altered expression in “manic” rats before lithium administration. Our observation is consistent with the previous human studies in bipolar disorder patients that presented several olfactory and gustatory dysfunctions [13] and the sensory enhancement and dysregulation of taste were often reported during an acute manic episode in BD patients [14].
Previous animal studies of lithium showed that mice chronically fed with lithium accumulated it mainly in neurogenic brain regions including olfactory bulb [15]. Similarly, chronic lithium effects in the neurogenic brain region (hippocampus) were also observed in human BD studies showing that lithium influenced hippocampal volume and structural plasticity in BD patients [16,17,18]. These findings suggest that chronic lithium treatment targets olfactory bulb and alters associated pathways (e.g., olfactory transduction).
On the contrary, in “depressive” rats we observed the upregulated expression of genes regulating olfactory receptor activity after 4-week lithium administration in the amygdala, which is consistent with the previous observations that bipolar patients during depressive episode experienced sensory blunting and olfactory deficits [14]. Thus chronic lithium treatment seems to restore olfactory function in rats presenting depressive-like behavior. Moreover, previous study showed that olfactory bulbectomy induced depressive-like behavior in rodents [19], in particular when taking into account the close anatomical links between the olfactory system and the brain circuits involved in memory [20] and emotion [21]. The recent studies reported the presence of sensory changes during mood swings in BD [22] and that olfactory assessment may be useful to screen unipolar and bipolar depression [23]. These genes involved in olfactory receptors activity were also differentially expressed in either manic and depressive rats responding to chronic lithium treatment and showed the opposite effect that also depended on tissue: in the amygdala they were upregulated after chronic lithium in mania, but downregulated in depression; in the hippocampus they showed downregulated expression in mania, but upregulated in depression after chronic lithium. This opposite gene expression profiles between amygdala and hippocampus after chronic lithium treatment may be further supported by results of the recent study that showed the opposite temporal profiles of protein expression between amygdala and hippocampal neurons in long-term response to acute stress [24].
Our findings from the animal model indicating the involvement of the taste receptor pathway were further supported by the observations in our clinical sample of bipolar and unipolar patients showing altered expression of taste receptor genes in peripheral blood leukocytes during depressive episode (Dmitrzak-Węglarz et al., under review). However, this pathway was not previously reported in the chronic lithium mechanism of action.
Another pathway significantly influenced by chronic lithium treatment in our animal mania model was bitter taste receptor pathway. Recent transcriptome study by Lee et al. [25] showed altered taste receptors gene expression during manic episodes. These genes included bitter taste receptors, TAS2R5 and TAS2R3, that were suggested as potential markers specific for the manic state [25]. These receptors were previously found downregulated (together with olfactory receptors) in the dorsolateral prefrontal cortex of schizophrenia postmortem brain tissues [26] and this downregulation influenced altered cognition. The taste alteration was also related to psychosocial and cognitive performance in bipolar patients [13]. However, the previous studies did not investigate the effects of mood stabilizers on taste-related gene expression. Our observation that chronic lithium influences the regulation of the olfactory and taste receptors’ pathway is supported by the previous case studies reporting long lasting impaired taste (dysgeusia) and smell (hyposmia) in lithium users: cluster headache patient [27] and bipolar patient [28].
Our study revealed that a number of differentially expressed genes influenced by chronic lithium treatment in the brain were long non-coding RNAs. The paper by ConLiGen showed significant associations for two long non-coding RNAs (lncRNAs) localized on chromosome 21 with lithium prophylactic efficacy supporting their involvement in modulating a clinical response [29]. Several studies reported that lncRNAs expression was dysregulated in psychiatric conditions, including BD [30] whereas Lee et al. found recently that lncRNAs were the predominant category showing upregulation during an acute manic episode [25]. Here, we report for the first time that lncRNAs expression is altered by chronic lithium treatment and suggest they may be regulators of antimanic lithium action.
Comparing the 2-week and 4-week gene expression profile, we found that the longer the treatment was, the more differentially expressed genes were observed, independent of the brain region. A previous study comparing microarrays from rat brains after 7 and 42 days of lithium treatment showed that, although plasma Li concentration reached therapeutic levels after 2 days of treatment, it required 2 weeks to reach therapeutic levels in the brain [11]. This finding suggested that the long-term lithium treatment and associated gene expression profile changes better represent the downstream Li effects, which are more relevant to its clinical effect. The other report in the rat fed for 21 days with lithium led to significant changes in the transcriptome in the frontal cortex [31].
To investigate the chronic lithium effect in the brain that mimics changes during the episode of depression or mania, we aimed to model mania and depression separately as the animal model of bipolar disorder does not exist [32]. Animal models are relevant platforms to evaluate the intracellular mechanisms in the brain associated with the pathogenesis of psychiatric disorders and drug studies, including lithium discovery [2,33,34]. In our study, we applied a chronic mild stress (CMS) protocol, the most extensively validated [35], to induce depressive-like behavior. Moreover, the CMS protocol was also suitable to model recurrent depression (a hallmark of bipolar disorder) [36]. The choice of amphetamine to mimic mania was supported by the previous preclinical studies that used repeated intraperitoneal injections of psychostimulants, such as amphetamine, to mimic acute manic episodes in rodents, including hyperactivity and risk-taking behavior due to increased sustained dopamine efflux [37,38,39,40,41,42]. Moreover, these behaviors were reversed by the administration of mood stabilizers, including lithium [40,43]. This chronic amphetamine-induced mania model showed recently good face, predictive and construct validity in modeling mania in Wistar rats [44,45,46,47].

4. Materials and Methods

4.1. Experimental Animals

All experimental procedures were performed in agreement with 3R rule and the study was approved by local ethical committee Poznan University of Life Sciences, Poland (agreement no. 22/2017, 23 June 2017). We used male Wistar rats with the baseline weight of 180 ± 10 g. The rats were housed five animals per cage with food and water available ad libitum and were maintained in a 12 h light/dark cycle (lights on at 7:00 a.m.) at a temperature of 22 ± 1 °C. All experiments were performed at the same time each day to avoid circadian variations. The animals were kept for one week of acclimatization and then were randomly divided into experimental groups with 5 animals in each group (amphetamine-exposed and control group, chronic mild stress-exposed and non-stressed rats; amphetamine-exposed rats receiving lithium or water and stress-exposed rats receiving lithium or water). After the acclimatization period, animals underwent baseline behavioral tests. The experimental design was shown in Figure 7.

4.2. Animal Model of Mania

Manic-like behavior was induced by daily intraperitoneal (i.p) injections of dextroamphetamine (d-AMPH) 2 mg/kg for 2 weeks as described previously [37,42]. The control group were animals receiving daily intraperitoneal injection with saline (0.9% NaCl, 1 mL/kg) for 14 days. After seven days, the behavior of all animals was assessed (Figure 7).

4.3. Animal Model of Depression

Animal model of depression was developed by the chronic mild stress protocol (CMS) as described previously [48]. To induce depressive behavior the animals were exposed for 4 weeks to different stress stimuli according to the CMS protocol described by Papp [49]. The control group were rats kept in standard conditions for 4 weeks that were not exposed to CMS. After two weeks of the start of the experiment, behavior of all animals was assessed (Figure 7).

4.4. Lithium Administration

The animals for the lithium study, after one week of amphetamine injections or two weeks of the chronic mild stress protocol and behavioral assessment, were randomly divided into lithium treated amphetamine-exposed group (n = 15) and amphetamine-exposed control group (rats receiving water, n = 15) and lithium-treated stress-exposed group (n = 15) and a stress-exposed control group (rats receiving water, n = 15). The animals were receiving the daily lithium solution in the dose of 1 mg/kg body mass or the same amount of water into the mouth with a syringe [50]. To minimize the side effects of lithium on kidney function, a saline bottle was provided in all lithium and control cages, as previously reported [51]. To assess the short term and long term lithium effects on brain transcriptome, the animals (n = 15 at each time in each group) were sacrificed by decapitation without euthanasia at two time points of chronic lithium administration, 14 and 30 days (long-term effect) after the first dose. The each control group (for stress-exposed and amphetamine-exposed rats) consisted of animals receiving water (Figure 7).

4.5. Behaviour Assessment

All the behavioral tests were performed during the light phase (between 08:00 a.m. and 12:00 p.m.), at room temperature in a separate quiet room. A blind evaluator assessed all behavioral parameters. To assess depressive-like or manic-like behavior we used a forced-swim test, open field test and elevated maze test at baseline and after induction of stress-induced depression or amphetamine-induced mania.
A forced swim test was performed to measure despair behavior and the hopelessness in the animal [52,53]. Each rat was placed in a glass cylinder 40 cm tall filled with water to a depth of 30 cm (24 ± 1 °C) so the rats could not support themselves by touching the bottom. Two swimming sessions were conducted: a 15-min pretest followed by a 5-min test 24 h later. For each animal, we recorded time spent immobilized (no additional activity other than that required to keep the rat’s head above the water) and active climbing time (upward-directed movements of the forepaws along the side of the swim chamber. The data were analyzed using the video tracking system (Videomot2, AnimaLab, Poznan, Poland). The water in the cylinders was replaced after each trial to remove urine or feces and to avoid confounding results.
Open field test was performed to evaluate the spontaneous locomotory activity in a new environment and exploration [54,55,56]. The rat was placed in the middle of the plastic box measuring 100 cm in diameter with 50-cm walls, divided into 25 squares of 20 × 20-cm size on the bottom (AnimaLab, Poland) and allowed to explore it for 5 min. We recorded the following parameters: time spent immobilized, time spent in center, total distance and exploration time and analyzed the data with the video tracking system (Videomot2, AnimaLab, Poland). After the end of each test, the open field was cleaned to remove any remaining materials (such as feces, urine or smell) that could interfere with the test results.
The elevated maze test was used to test the anxiety level and risk-taking behavior in the rats [42]. The apparatus consisted of two opposite open arms, two opposite closed arms and a central platform connecting the four arms. The maze was propped up 50 cm away from the ground by the maze feet. The test room was maintained with constant temperature, humidity and illumination. The rats were placed in the central platform facing one of the open arms. The number of entries into the open and closed arms and the total time spent in each arm during a 5-min exploration period were recorded by the video tracking system (Videomot2, AnimaLab, Poznań, Poland). After the end of each test, the open field was cleaned to remove any remaining materials (such as feces, urine or smell) that could interfere with the test results.

4.6. Microarray-Based Gene Expression Analysis

The different regions of brain tissue (amygdala, hippocampus, prefrontal cortex and hypothalamus) were collected immediately after decapitation (without anesthesia). The skull was opened, and the cerebral content was excised and rapidly dissected on a chilled Petri dish. The frontal cortex, hippocampus and amygdala were isolated and cleaned from the subcortical structures and white matter, immediately snap frozen in liquid nitrogen and kept frozen in −80 °C for further processing. Frozen tissues were used for RNA extraction using Nucleospin RNA/Protein kit (Macherey Nagel, Dylan, Germany). RNA integrity (RIN) was assessed using Tape Station 2200 (Agilent) and RNA concentration was measured using fluorimeter (Quantus, Promega). Total RNA from amygdala, hippocampus and frontal cortex in the starting amount of 50 ng was used for microarray experiments. We used SurePrint G3 Rat Gene Expression v2 Microarray Kit in format 8 × 60 k and one-color Low Input Quick Amp Labeling Kit (Agilent Technologies, Cedar Creek, TX, USA) to analyze gene expression profile following the standard protocol provided by the manufacturer. The hybridization signals were detected with SureScan Dx Microarray Scanner (Agilent Technologies, Santa Clara, CA, USA). The images obtained after scanning were analyzed with Agilent Feature extraction software v.12.0.3.1.

4.7. Statistical Analysis

Behavioral data were analyzed using a t-test for paired samples after checking normality of data distribution using the Shapiro–Wilk test. A cut-off value p < 0.05 was regarded as significant. Data were shown as mean ± SEM in figures and text if not otherwise stated. For behavioral measurements and gene expression data, 5 animals per experimental group were sufficient to achieve power of 80% (as calculated by the G-power calculator available at https://stats.idre.ucla.edu/other/gpower). To conform to the 3R’s rule, we included the minimal required number of animals (n = 5) in the study.
Bioinformatic analysis of gene expression data was analyzed with the use of Gene Spring 14.9 software (Agilent Technologies, Santa Clara, CA, USA). To identify differentially expressed genes, moderated t-statistics from the empirical Bayes method on normalized fluorescence signal was used for the fold of change (FC) calculation. FC was calculated in relation to the adequate control group. The list of significantly differentially expressed transcripts (p < 0.05 and FC > 2.0) was generated using statistical filtering (moderated t-test with multiple test correction FDR). Then, separate lists of up- and downregulated genes were used for functional analysis to identify Gene Ontology (GO) terms and pathways significantly enriched by the set of differentially expressed genes, either upregulated or downregulated (p < 0.05). Gene Ontology and pathway analyses were done in Gene Spring.
To identify the genes and pathways shared during lithium administration, we used Venn diagrams using a tool freely available at: http://bioinformatics.psb.ugent.be/webtools/Venn. For shared genes, we used G:Profiler tool (available at: https://biit.cs.ut.ee/gprofiler/gost) to perform functional enrichment analysis (gene set enrichment analysis) including GO and pathways from KEGG Reactome and Wiki Pathways [57]. Heat maps were drawn using the Heatmapper tool available at: http://www.heatmapper.ca.

5. Conclusions

We reported here for the first time that genes regulating olfactory and taste receptor pathways and long non-coding RNAs, that were implicated in BD pathogenesis, might be targeted by chronic lithium treatment in animals presenting manic-like behavior. Further functional studies of these pathways are warranted to elucidate the exact therapeutic molecular mechanism of lithium.

Author Contributions

D.S. participated in the study design and conceptualization, performed animal experiments, data interpretation and drafted the manuscript; P.C., P.A.K., E.P.-O., M.S. (Maciej Sassek), P.Z., E.B. and W.L. performed animal experiments, analyzed the data and revised the manuscript; K.S. and J.N. performed molecular analysis and revised the manuscript; M.S. (Magdalena Socha) and E.B.-O. optimized the microarray experiment and revised the manuscript; J.P. participated in the study design and supervised the experiments; J.T.-H. acquired the funding and supervised the experiments, L.N. and J.K.R. supervised the experiments and revised the manuscript, A.S. participated in the study design, administered the project, performed microarray experiments, analyzed and interpreted the data and revised the manuscript. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by National Science Centre, Poland, grant number 2016/21/B/NZ5/00148.

Institutional Review Board Statement

The study was approved by local ethical committee at Poznan University of Life Sciences, Poland (agreement no. 22/2017, 23 June 2017).

Data Availability Statement

The detailed data used to support the findings of this study are available from the corresponding author upon written request.

Acknowledgments

We would like to thank the staff of the Department of Medical Genetics, Poznan University of Medical Sciences for access to the laboratory equipment for microarrays experiments.

Conflicts of Interest

The authors declare no conflict of interest. The funding body had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

References

  1. American Psychiatric Association. Diagnostic and Statistical Manual of Mental Disorders; American Psychiatric Pub.: Washington, DC, USA, 2013. [Google Scholar]
  2. Cade, J.F. Lithium salts in the treatment of psychotic excitement. Med. J. Aust. 1949, 2, 349–352. [Google Scholar] [CrossRef]
  3. Rybakowski, J. Lithium carbonate in endogenous depression. Psychiatr. Pol. 1972, 6, 547–550. [Google Scholar]
  4. Mendels, J. Lithium in the treatment of depression. Am. J. Psychiatry 1976, 133, 373–378. [Google Scholar]
  5. Rybakowski, J.K. Lithium-past, present, future. Int J. Psychiatry Clin. Pr. 2020, 24, 330–340. [Google Scholar] [CrossRef]
  6. Harwood, A.J. Lithium and bipolar mood disorder: The inositol-depletion hypothesis revisited. Mol. Psychiatry 2005, 10, 117–126. [Google Scholar] [CrossRef]
  7. Chalecka-Franaszek, E.; Chuang, D.M. Lithium activates the serine/threonine kinase akt-1 and suppresses glutamate-induced inhibition of akt-1 activity in neurons. Proc. Natl. Acad. Sci. USA 1999, 96, 8745–8750. [Google Scholar] [CrossRef]
  8. De Sarno, P.; Li, X.; Jope, R.S. Regulation of akt and glycogen synthase kinase-3 beta phosphorylation by sodium valproate and lithium. Neuropharmacology 2002, 43, 1158–1164. [Google Scholar] [CrossRef]
  9. Severino, G.; Squassina, A.; Costa, M.; Pisanu, C.; Calza, S.; Alda, M.; Del Zompo, M.; Manchia, M. Pharmacogenomics of bipolar disorder. Pharmacogenomics 2013, 14, 655–674. [Google Scholar] [CrossRef]
  10. Alda, M. Lithium in the treatment of bipolar disorder: Pharmacology and pharmacogenetics. Mol. Psychiatry 2015, 20, 661–670. [Google Scholar] [CrossRef]
  11. Bosetti, F.; Seemann, R.; Bell, J.M.; Zahorchak, R.; Friedman, E.; Rapoport, S.I.; Manickam, P. Analysis of gene expression with cdna microarrays in rat brain after 7 and 42 days of oral lithium administration. Brain Res. Bull. 2002, 57, 205–209. [Google Scholar] [CrossRef]
  12. McQuillin, A.; Rizig, M.; Gurling, H.M. A microarray gene expression study of the molecular pharmacology of lithium carbonate on mouse brain mrna to understand the neurobiology of mood stabilization and treatment of bipolar affective disorder. Pharm. Genom. 2007, 17, 605–617. [Google Scholar] [CrossRef] [PubMed]
  13. Kazour, F.; Richa, S.; Desmidt, T.; Lemaire, M.; Atanasova, B.; El Hage, W. Olfactory and gustatory functions in bipolar disorders: A systematic review. Neurosci. Biobehav. Rev. 2017, 80, 69–79. [Google Scholar] [CrossRef] [PubMed]
  14. Parker, G.; Paterson, A.; Romano, M.; Granville Smith, I. Suprasensory phenomena in those with a bipolar disorder. Australas. Psychiatry 2018, 26, 384–387. [Google Scholar] [CrossRef] [PubMed]
  15. Zanni, G.; Michno, W.; Di Martino, E.; Tjarnlund-Wolf, A.; Pettersson, J.; Mason, C.E.; Hellspong, G.; Blomgren, K.; Hanrieder, J. Lithium accumulates in neurogenic brain regions as revealed by high resolution ion imaging. Sci. Rep. 2017, 7, 40726. [Google Scholar] [CrossRef] [PubMed]
  16. Bertolino, A.; Frye, M.; Callicott, J.H.; Mattay, V.S.; Rakow, R.; Shelton-Repella, J.; Post, R.; Weinberger, D.R. Neuronal pathology in the hippocampal area of patients with bipolar disorder: A study with proton magnetic resonance spectroscopic imaging. Biol. Psychiatry 2003, 53, 906–913. [Google Scholar] [CrossRef]
  17. Colla, M.; Schubert, F.; Bubner, M.; Heidenreich, J.O.; Bajbouj, M.; Seifert, F.; Luborzewski, A.; Heuser, I.; Kronenberg, G. Glutamate as a spectroscopic marker of hippocampal structural plasticity is elevated in long-term euthymic bipolar patients on chronic lithium therapy and correlates inversely with diurnal cortisol. Mol. Psychiatry 2009, 14, 696–704. [Google Scholar] [CrossRef] [PubMed]
  18. Giakoumatos, C.I.; Nanda, P.; Mathew, I.T.; Tandon, N.; Shah, J.; Bishop, J.R.; Clementz, B.A.; Pearlson, G.D.; Sweeney, J.A.; Tamminga, C.A.; et al. Effects of lithium on cortical thickness and hippocampal subfield volumes in psychotic bipolar disorder. J. Psychiatr. Res. 2015, 61, 180–187. [Google Scholar] [CrossRef]
  19. Poretti, M.B.; Rask-Andersen, M.; Kumar, P.; Rubiales de Barioglio, S.; Fiol de Cuneo, M.; Schioth, H.B.; Carlini, V.P. Ghrelin effects expression of several genes associated with depression-like behavior. Prog. Neuropsychopharmacol. Biol. Psychiatry 2015, 56, 227–234. [Google Scholar] [CrossRef]
  20. Savic, I.; Gulyas, B.; Larsson, M.; Roland, P. Olfactory functions are mediated by parallel and hierarchical processing. Neuron 2000, 26, 735–745. [Google Scholar] [CrossRef]
  21. Anderson, A.K.; Christoff, K.; Stappen, I.; Panitz, D.; Ghahremani, D.G.; Glover, G.; Gabrieli, J.D.; Sobel, N. Dissociated neural representations of intensity and valence in human olfaction. Nat. Neurosci. 2003, 6, 196–202. [Google Scholar] [CrossRef]
  22. Parker, G.; Paterson, A.; Romano, M.; Graham, R. Altered sensory phenomena experienced in bipolar disorder. Am. J. Psychiatry 2017, 174, 1146–1150. [Google Scholar] [CrossRef] [PubMed]
  23. Kazour, F.; Richa, S.; Abi Char, C.; Surget, A.; Elhage, W.; Atanasova, B. Olfactory markers for depression: Differences between bipolar and unipolar patients. PLoS ONE 2020, 15, e0237565. [Google Scholar] [CrossRef] [PubMed]
  24. Madan, J.S.; Gupta, K.; Chattarji, S.; Bhattacharya, A. Hippocampal and amygdalar cell-specific translation is similar soon after stress but diverge over time. Hippocampus 2018, 28, 441–452. [Google Scholar] [CrossRef] [PubMed]
  25. Lee, Y.C.; Chao, Y.L.; Chang, C.E.; Hsieh, M.H.; Liu, K.T.; Chen, H.C.; Lu, M.L.; Chen, W.Y.; Chen, C.H.; Tsai, M.H.; et al. Transcriptome changes in relation to manic episode. Front. Psychiatry 2019, 10, 280. [Google Scholar] [CrossRef] [PubMed]
  26. Ansoleaga, B.; Garcia-Esparcia, P.; Pinacho, R.; Haro, J.M.; Ramos, B.; Ferrer, I. Decrease in olfactory and taste receptor expression in the dorsolateral prefrontal cortex in chronic schizophrenia. J. Psychiatr. Res. 2015, 60, 109–116. [Google Scholar] [CrossRef]
  27. De Coo, I.F.; Haan, J. Long lasting impairment of taste and smell as side effect of lithium carbonate in a cluster headache patient. Headache 2016, 56, 1201–1203. [Google Scholar] [CrossRef] [PubMed]
  28. Terao, T.; Watanabe, S.; Hoaki, N.; Hoaki, T. Strange taste and mild lithium intoxication. BMJ Case Rep. 2011, 2011. [Google Scholar] [CrossRef]
  29. Hou, L.; Heilbronner, U.; Degenhardt, F.; Adli, M.; Akiyama, K.; Akula, N.; Ardau, R.; Arias, B.; Backlund, L.; Banzato, C.E.M.; et al. Genetic variants associated with response to lithium treatment in bipolar disorder: A genome-wide association study. Lancet 2016, 387, 1085–1093. [Google Scholar] [CrossRef]
  30. Akula, N.; Barb, J.; Jiang, X.; Wendland, J.R.; Choi, K.H.; Sen, S.K.; Hou, L.; Chen, D.T.; Laje, G.; Johnson, K.; et al. Rna-sequencing of the brain transcriptome implicates dysregulation of neuroplasticity, circadian rhythms and gtpase binding in bipolar disorder. Mol. Psychiatry 2014, 19, 1179–1185. [Google Scholar] [CrossRef]
  31. Fatemi, S.H.; Reutiman, T.J.; Folsom, T.D. The role of lithium in modulation of brain genes: Relevance for aetiology and treatment of bipolar disorder. Biochem. Soc. Trans. 2009, 37, 1090–1095. [Google Scholar] [CrossRef]
  32. Kato, T.; Kasahara, T.; Kubota-Sakashita, M.; Kato, T.M.; Nakajima, K. Animal models of recurrent or bipolar depression. Neuroscience 2016, 321, 189–196. [Google Scholar] [CrossRef] [PubMed]
  33. Beyer, D.K.E.; Freund, N. Animal models for bipolar disorder: From bedside to the cage. Int. J. Bipolar Disord. 2017, 5, 35. [Google Scholar] [CrossRef] [PubMed]
  34. Lan, A.; Einat, H. Questioning the predictive validity of the amphetamine-induced hyperactivity model for screening mood stabilizing drugs. Behav. Brain Res. 2019, 362, 109–113. [Google Scholar] [CrossRef] [PubMed]
  35. Willner, P. Reliability of the chronic mild stress model of depression: A user survey. Neurobiol. Stress 2017, 6, 68–77. [Google Scholar] [CrossRef] [PubMed]
  36. Remus, J.L.; Jamison, D.; Johnson, J.D. An animal model of recurrent depression: Sensitized depression-like behavior when rats are re-exposed to chronic mild stress. Brain Behav. Immun. 2013, 32, e4–e5. [Google Scholar] [CrossRef]
  37. Frey, B.N.; Martins, M.R.; Petronilho, F.C.; Dal-Pizzol, F.; Quevedo, J.; Kapczinski, F. Increased oxidative stress after repeated amphetamine exposure: Possible relevance as a model of mania. Bipolar Disord. 2006, 8, 275–280. [Google Scholar] [CrossRef] [PubMed]
  38. Szabo, S.T.; Machado-Vieira, R.; Yuan, P.; Wang, Y.; Wei, Y.; Falke, C.; Cirelli, C.; Tononi, G.; Manji, H.K.; Du, J. Glutamate receptors as targets of protein kinase c in the pathophysiology and treatment of animal models of mania. Neuropharmacology 2009, 56, 47–55. [Google Scholar] [CrossRef]
  39. Feier, G.; Valvassori, S.S.; Varela, R.B.; Resende, W.R.; Bavaresco, D.V.; Morais, M.O.; Scaini, G.; Andersen, M.L.; Streck, E.L.; Quevedo, J. Lithium and valproate modulate energy metabolism in an animal model of mania induced by methamphetamine. Pharm. Biochem. Behav. 2013, 103, 589–596. [Google Scholar] [CrossRef]
  40. Zhou, Z.; Wang, Y.; Tan, H.; Bharti, V.; Che, Y.; Wang, J.F. Chronic treatment with mood stabilizer lithium inhibits amphetamine-induced risk-taking manic-like behaviors. Neurosci. Lett. 2015, 603, 84–88. [Google Scholar] [CrossRef]
  41. Valvassori, S.S.; Resende, W.R.; Dal-Pont, G.; Sangaletti-Pereira, H.; Gava, F.F.; Peterle, B.R.; Carvalho, A.F.; Varela, R.B.; Dal-Pizzol, F.; Quevedo, J. Lithium ameliorates sleep deprivation-induced mania-like behavior, hypothalamic-pituitary-adrenal (hpa) axis alterations, oxidative stress and elevations of cytokine concentrations in the brain and serum of mice. Bipolar Disord. 2017, 19, 246–258. [Google Scholar] [CrossRef]
  42. Valvassori, S.S.; Gava, F.F.; Dal-Pont, G.C.; Simoes, H.L.; Damiani-Neves, M.; Andersen, M.L.; Boeck, C.R.; Quevedo, J. Effects of lithium and valproate on erk/jnk signaling pathway in an animal model of mania induced by amphetamine. Heliyon 2019, 5, e01541. [Google Scholar] [CrossRef] [PubMed]
  43. Valvassori, S.S.; Tonin, P.T.; Varela, R.B.; Carvalho, A.F.; Mariot, E.; Amboni, R.T.; Bianchini, G.; Andersen, M.L.; Quevedo, J. Lithium modulates the production of peripheral and cerebral cytokines in an animal model of mania induced by dextroamphetamine. Bipolar Disord. 2015, 17, 507–517. [Google Scholar] [CrossRef] [PubMed]
  44. Menegas, S.; Dal-Pont, G.C.; Cararo, J.H.; Varela, R.B.; Aguiar-Geraldo, J.M.; Possamai-Della, T.; Andersen, M.L.; Quevedo, J.; Valvassori, S.S. Efficacy of folic acid as an adjunct to lithium therapy on manic-like behaviors, oxidative stress and inflammatory parameters in an animal model of mania. Metab. Brain Dis. 2020, 35, 413–425. [Google Scholar] [CrossRef] [PubMed]
  45. Varela, R.B.; Resende, W.R.; Dal-Pont, G.C.; Gava, F.F.; Nadas, G.B.; Tye, S.J.; Andersen, M.L.; Quevedo, J.; Valvassori, S.S. Role of epigenetic regulatory enzymes in animal models of mania induced by amphetamine and paradoxical sleep deprivation. Eur J. Neurosci. 2020. [Google Scholar] [CrossRef] [PubMed]
  46. Valvassori, S.S.; Tonin, P.T.; Dal-Pont, G.C.; Varela, R.B.; Cararo, J.H.; Garcia, A.F.; Gava, F.F.; Menegas, S.; Soares, J.C.; Quevedo, J. Coadministration of lithium and celecoxib reverses manic-like behavior and decreases oxidative stress in a dopaminergic model of mania induced in rats. Transl. Psychiatry 2019, 9, 297. [Google Scholar] [CrossRef]
  47. Bristot, G.; Ascoli, B.M.; Scotton, E.; Gea, L.P.; Pfaffenseller, B.; Kauer-Sant’Anna, M. Effects of lithium on inflammatory and neurotrophic factors after an immune challenge in a lisdexamfetamine animal model of mania. Braz. J. Psychiatry 2019, 41, 419–427. [Google Scholar] [CrossRef] [PubMed]
  48. Willner, P.; Muscat, R.; Papp, M. Chronic mild stress-induced anhedonia: A realistic animal model of depression. Neurosci. Biobehav. Rev. 1992, 16, 525–534. [Google Scholar] [CrossRef]
  49. Papp, M. Models of affective illness: Chronic mild stress in the rat. Curr. Protoc. Pharm. 2012, 57. [Google Scholar] [CrossRef]
  50. Atcha, Z.; Rourke, C.; Neo, A.H.; Goh, C.W.; Lim, J.S.; Aw, C.C.; Browne, E.R.; Pemberton, D.J. Alternative method of oral dosing for rats. J. Am. Assoc. Lab. Anim. Sci. 2010, 49, 335–343. [Google Scholar]
  51. Chen, G.; Rajkowska, G.; Du, F.; Seraji-Bozorgzad, N.; Manji, H.K. Enhancement of hippocampal neurogenesis by lithium. J. Neurochem. 2000, 75, 1729–1734. [Google Scholar] [CrossRef]
  52. Porsolt, R.D.; Bertin, A.; Jalfre, M. Behavioral despair in mice: A primary screening test for antidepressants. Arch. Int. Pharm. 1977, 229, 327–336. [Google Scholar]
  53. Willner, P.; Bergman, J.; Vanderschuren, L.; Ellenbroek, B. Pharmacological approaches to the study of social behaviour. Behav. Pharm. 2015, 26, 501–504. [Google Scholar] [CrossRef] [PubMed]
  54. Jindal, A.; Mahesh, R.; Bhatt, S. Etazolate rescues behavioral deficits in chronic unpredictable mild stress model: Modulation of hypothalamic-pituitary-adrenal axis activity and brain-derived neurotrophic factor level. Neurochem. Int. 2013, 63, 465–475. [Google Scholar] [CrossRef] [PubMed]
  55. Belovicova, K.; Bogi, E.; Csatlosova, K.; Dubovicky, M. Animal tests for anxiety-like and depression-like behavior in rats. Interdiscip. Toxicol. 2017, 10, 40–43. [Google Scholar] [CrossRef]
  56. Sun, H.L.; Zhou, Z.Q.; Zhang, G.F.; Yang, C.; Wang, X.M.; Shen, J.C.; Hashimoto, K.; Yang, J.J. Role of hippocampal p11 in the sustained antidepressant effect of ketamine in the chronic unpredictable mild stress model. Transl. Psychiatry 2016, 6, e741. [Google Scholar] [CrossRef]
  57. Raudvere, U.; Kolberg, L.; Kuzmin, I.; Arak, T.; Adler, P.; Peterson, H.; Vilo, J. G:Profiler: A web server for functional enrichment analysis and conversions of gene lists (2019 update). Nucleic Acids Res. 2019, 47, W191–W198. [Google Scholar] [CrossRef]
Figure 1. The comparison of behavioral changes before and after amphetamine injection in the elevated maze test: (a) number of visits in open arms, (b) time spent in open arms, (c) distance passed by the animals (paired t-test, * defines p < 0.05).
Figure 1. The comparison of behavioral changes before and after amphetamine injection in the elevated maze test: (a) number of visits in open arms, (b) time spent in open arms, (c) distance passed by the animals (paired t-test, * defines p < 0.05).
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Figure 2. The results of behavioral changes after chronic mild stress protocol: (ad) are parameters measured in an open filed test and (e,f) are parameters measured by a forced swim test (paired t-test, * defines p < 0.05).
Figure 2. The results of behavioral changes after chronic mild stress protocol: (ad) are parameters measured in an open filed test and (e,f) are parameters measured by a forced swim test (paired t-test, * defines p < 0.05).
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Figure 3. Differentially expressed genes (up- and downregulated) in three analyzed brain regions (amygdala, frontal cortex and hippocampus) in amphetamine-exposed (a,b, respectively) and stress-exposed rats (c,d, respectively) as compared to the control group.
Figure 3. Differentially expressed genes (up- and downregulated) in three analyzed brain regions (amygdala, frontal cortex and hippocampus) in amphetamine-exposed (a,b, respectively) and stress-exposed rats (c,d, respectively) as compared to the control group.
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Figure 4. Genes differentially expressed in rats depressive-like after 2 weeks and 4 weeks on lithium treatment in the amygdala, frontal cortex and hippocampus. Shared genes between these time points were indicated by purple color.
Figure 4. Genes differentially expressed in rats depressive-like after 2 weeks and 4 weeks on lithium treatment in the amygdala, frontal cortex and hippocampus. Shared genes between these time points were indicated by purple color.
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Figure 5. Gene-set enrichment analysis indicated that the genes from olfactory transduction pathway are influenced by chronic lithium administration in manic-like rats in analyzed brain areas (amygdala, frontal cortex and hippocampus).
Figure 5. Gene-set enrichment analysis indicated that the genes from olfactory transduction pathway are influenced by chronic lithium administration in manic-like rats in analyzed brain areas (amygdala, frontal cortex and hippocampus).
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Figure 6. The comparison of behavioral changes before and after lithium administration in: (ac) rats with manic-like behavior using elevated maze test and (df) rats with depressive-like behavior using open field test (one-way analysis of variance).
Figure 6. The comparison of behavioral changes before and after lithium administration in: (ac) rats with manic-like behavior using elevated maze test and (df) rats with depressive-like behavior using open field test (one-way analysis of variance).
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Figure 7. The experimental design of the study showing the (a) procedure of inducing manic-like behavior, (b) procedure of inducing depressive-like behavior, (c) lithium administration in rats with manic-like behavior and (d) lithium administration in animals showing depressive-like behavior. CMS—chronic mild stress protocol; EMT—elevated maze test, FST—forced swim test, OFT—open field test.
Figure 7. The experimental design of the study showing the (a) procedure of inducing manic-like behavior, (b) procedure of inducing depressive-like behavior, (c) lithium administration in rats with manic-like behavior and (d) lithium administration in animals showing depressive-like behavior. CMS—chronic mild stress protocol; EMT—elevated maze test, FST—forced swim test, OFT—open field test.
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Table 1. Gene Ontology (GO) terms enriched from significantly downregulated genes between rats with manic-like behavior and control rats in the amygdala.
Table 1. Gene Ontology (GO) terms enriched from significantly downregulated genes between rats with manic-like behavior and control rats in the amygdala.
GO AccessionGO TermCorr pNo. Genes
GO:0007186G-protein coupled receptor signaling pathway0.000117
GO:0007600sensory perception0.000103
GO:0051606detection of stimulus0.00088
GO:0050906detection of stimulus involved in sensory perception0.00084
GO:0038023signaling receptor activity0.000120
GO:0060089molecular transducer activity0.000120
GO:0009593detection of chemical stimulus0.00081
GO:0050907detection of chemical stimulus involved in sensory perception0.00079
GO:0004888transmembrane signaling receptor activity0.000111
GO:0004930G-protein coupled receptor activity0.00093
GO:0004871signal transducer activity0.000126
GO:0007606sensory perception of chemical stimulus0.00082
GO:0004984olfactory receptor activity0.00074
GO:0050911detection of chemical stimulus involved in sensory perception of smell0.00074
GO:0007608sensory perception of smell0.00075
GO:0007165signal transduction0.000201
GO:0003008system process0.000123
GO:0050877nervous system process0.000106
GO:0023052signaling0.000209
GO:0007154cell communication0.000212
GO:0050896response to stimulus0.001287
GO:0051716cellular response to stimulus0.004238
GO:0005833hemoglobin complex0.0195
Table 2. GO terms significantly enriched from differentially expressed genes between rats with depressive-like behavior and control group in the amygdala.
Table 2. GO terms significantly enriched from differentially expressed genes between rats with depressive-like behavior and control group in the amygdala.
GO AccessionGO TermCorr pNo. Genes
Downregulated
GO:0007610behavior0.00226
GO:0007188adenylate cyclase-modulating G-protein coupled receptor signaling pathway0.00212
GO:0007187G-protein coupled receptor signaling pathway, coupled to cyclic nucleotide second messenger0.00312
GO:0044459plasma membrane part0.00453
GO:0003013circulatory system process0.00417
GO:0008015blood circulation0.00417
GO:0007626locomotory behavior0.00414
GO:0007193adenylate cyclase-inhibiting G-protein coupled receptor signaling pathway0.0377
Upregulated
GO:0023052signaling0.000102
GO:0007154cell communication0.000102
GO:0007165signal transduction0.00095
GO:0038023signaling receptor activity0.00155
GO:0060089molecular transducer activity0.00155
GO:0004888transmembrane signaling receptor activity0.00152
GO:0004871signal transducer activity0.00657
GO:0007186G-protein coupled receptor signaling pathway0.00649
GO:0003008system process0.00657
GO:0051606detection of stimulus0.01737
GO:0050896response to stimulus0.022126
GO:0004930G-protein coupled receptor activity0.02840
GO:0009593detection of chemical stimulus0.02834
GO:0050906detection of stimulus involved in sensory perception0.04534
GO:0009653anatomical structure morphogenesis0.05045
Table 3. Differentially expressed genes enriched in GO terms in amphetamine-exposed rats after two weeks of lithium treatment.
Table 3. Differentially expressed genes enriched in GO terms in amphetamine-exposed rats after two weeks of lithium treatment.
GO AccessionGO TermCorr pNo. Genes
Amygdala upregulated
GO:0038023signaling receptor activity0.000367
GO:0060089molecular transducer activity0.000367
GO:0004888transmembrane signaling receptor activity0.000352
GO:0007186G-protein coupled receptor signaling pathway0.000344
GO:0007600sensory perception0.000310
GO:0004930G-protein coupled receptor activity0.000299
GO:0007606sensory perception of chemical stimulus0.000285
GO:0051606detection of stimulus0.000281
GO:0050906detection of stimulus involved in sensory perception0.000273
GO:0009593detection of chemical stimulus0.000268
GO:0050907detection of chemical stimulus involved in sensory perception0.000264
GO:0007608sensory perception of smell0.000256
GO:0004984olfactory receptor activity0.000247
GO:0050911detection of chemical stimulus involved in sensory perception of smell0.000247
GO:0004871signal transducer activity0.000379
GO:0050877nervous system process0.000330
GO:0003008system process0.000363
GO:0031224intrinsic component of membrane0.000640
GO:0016021integral component of membrane0.000627
GO:0007165signal transduction0.000509
GO:0042221response to chemical0.000507
GO:0044425membrane part0.000687
GO:0005886plasma membrane0.000521
GO:0005549odorant binding0.00068
GO:0071944cell periphery0.000528
GO:0023052signaling0.000523
GO:0007154cell communication0.000533
GO:0032501multicellular organismal process0.000669
GO:0050896response to stimulus0.000731
GO:0004252serine-type endopeptidase activity0.00044
GO:0051716cellular response to stimulus0.000597
GO:0008236serine-type peptidase activity0.00044
GO:0017171serine hydrolase activity0.00044
GO:0008527taste receptor activity0.00015
GO:0050912detection of chemical stimulus involved in sensory perception of taste0.00117
GO:0050913sensory perception of bitter taste0.00216
GO:0033038bitter taste receptor activity0.00213
GO:0001580detection of chemical stimulus involved in sensory perception of bitter taste0.00415
GO:0004175endopeptidase activity0.00561
GO:0050909sensory perception of taste0.00719
Frontal cortex downregulated
GO:0038023signaling receptor activity0.02280
GO:0060089molecular transducer activity0.02280
GO:0007608sensory perception of smell0.02252
GO:0004984olfactory receptor activity0.02250
GO:0050911detection of chemical stimulus involved in sensory perception of smell0.02250
GO:0009593detection of chemical stimulus0.02652
GO:0050907detection of chemical stimulus involved in sensory perception0.02651
GO:0051606detection of stimulus0.02955
GO:0007606sensory perception of chemical stimulus0.03354
GO:0050906detection of stimulus involved in sensory perception0.03352
GO:0004888transmembrane signaling receptor activity0.03672
Table 4. Differentially expressed genes enriched in GO terms in stress-exposed rats after two weeks of lithium treatment.
Table 4. Differentially expressed genes enriched in GO terms in stress-exposed rats after two weeks of lithium treatment.
GO AccessionGO TermCorr pNo. Genes
Amygdala (top 20 out of 206)
GO:0005578proteinaceous extracellular matrix0.00041
GO:0031012extracellular matrix0.00034
GO:0005576extracellular region0.000117
GO:0044421extracellular region part0.000108
GO:0005615extracellular space0.000104
GO:0044420extracellular matrix component0.00019
GO:0009888tissue development0.00062
GO:0030198extracellular matrix organization0.00021
GO:0009653anatomical structure morphogenesis0.00068
GO:0043062extracellular structure organization0.00021
GO:0007275multicellular organism development0.000112
GO:0030199collagen fibril organization0.00011
GO:0072001renal system development0.00024
GO:0005581collagen trimer0.00013
GO:0032502developmental process0.000122
GO:0048513animal organ development0.00087
GO:0048646anatomical structure formation involved in morphogenesis0.00039
GO:0009887animal organ morphogenesis0.00041
GO:0048731system development0.000104
Hippocampus
GO:0002474antigen processing and presentation of peptide antigen via MHC class I0.0244
GO:0042605peptide antigen binding0.0244
Table 5. Differentially expressed genes enriched in GO terms in amphetamine-exposed rats after four weeks of lithium treatment.
Table 5. Differentially expressed genes enriched in GO terms in amphetamine-exposed rats after four weeks of lithium treatment.
GO AccessionGO TermCorr pNo. Genes
Amygdala downregulated
GO:0038023signaling receptor activity0.000228
GO:0060089molecular transducer activity0.000228
GO:0004888transmembrane signaling receptor activity0.000212
GO:0007608sensory perception of smell0.000149
GO:0004984olfactory receptor activity0.000144
GO:0050911detection of chemical stimulus involved in sensory perception of smell0.000144
GO:0007606sensory perception of chemical stimulus0.000159
GO:0007600sensory perception0.000186
GO:0050907detection of chemical stimulus involved in sensory perception0.000147
GO:0050906detection of stimulus involved in sensory perception0.000151
GO:0051606detection of stimulus0.000157
GO:0009593detection of chemical stimulus0.000147
GO:0004930G-protein coupled receptor activity0.000172
GO:0004871signal transducer activity0.000237
GO:0007186G-protein coupled receptor signaling pathway0.000196
GO:0050877nervous system process0.000199
GO:0031224intrinsic component of membrane0.000451
GO:0016021integral component of membrane0.000441
GO:0003008system process0.000220
GO:0044425membrane part0.002488
GO:0005886plasma membrane0.003365
GO:0023052signaling0.007366
GO:0071944cell periphery0.008369
GO:0007165|GO:0023033signal transduction0.009346
GO:0007154cell communication0.011373
GO:0005549odorant binding0.02040
Frontal cortex downregulated
GO:0007606sensory perception of chemical stimulus0.000154
GO:0004984olfactory receptor activity0.000140
GO:0050911detection of chemical stimulus involved in sensory perception of smell0.000140
GO:0009593detection of chemical stimulus0.000145
GO:0050907detection of chemical stimulus involved in sensory perception0.000143
GO:0007608sensory perception of smell0.000142
GO:0007600sensory perception0.000171
GO:0050906detection of stimulus involved in sensory perception0.000144
GO:0051606detection of stimulus0.000148
GO:0004888transmembrane signaling receptor activity0.000183
GO:0004930G-protein coupled receptor activity0.000154
GO:0038023signaling receptor activity0.000190
GO:0060089molecular transducer activity0.000190
GO:0050877nervous system process0.000177
GO:0007186G-protein coupled receptor signaling pathway0.000172
GO:0004871signal transducer activity0.000197
GO:0003008system process0.000195
GO:0031224intrinsic component of membrane0.000340
GO:0016021integral component of membrane0.000334
GO:0071944cell periphery0.000287
GO:0005886plasma membrane0.000281
GO:0042221response to chemical0.000269
GO:0044425membrane part0.000364
GO:0032501multicellular organismal process0.001352
GO:0005549odorant binding0.00134
GO:0036156inner dynein arm0.0064
GO:0007165signal transduction0.006252
GO:0004252serine-type endopeptidase activity0.01922
GO:0023052signaling0.030261
GO:0007154cell communication0.030267
GO:0008236serine-type peptidase activity0.04323
Hippocampus downregulated
GO:0001580detection of chemical stimulus involved in sensory perception of bitter taste0.0001394
GO:0001594trace-amine receptor activity0.0001323
GO:0003008system process0.0001316
GO:0004252serine-type endopeptidase activity0.0001313
GO:0004866endopeptidase inhibitor activity0.0001212
GO:0004867serine-type endopeptidase inhibitor activity0.0001119
GO:0004869cysteine-type endopeptidase inhibitor activity0.0001105
GO:0004871signal transducer activity0.0001083
GO:0004888transmembrane signaling receptor activity0.0001062
GO:0004930G-protein coupled receptor activity0.0001053
GO:0004984olfactory receptor activity0.0001037
GO:0005179hormone activity0.000888
GO:0005549odorant binding0.000877
GO:0005886plasma membrane0.000877
GO:0007154cell communication0.000850
GO:0007165signal transduction0.000845
GO:0007186G-protein coupled receptor signaling pathway0.000835
GO:0007600sensory perception0.000791
GO:0007606sensory perception of chemical stimulus0.000769
GO:0007608sensory perception of smell0.000740
GO:0008527taste receptor activity0.000720
GO:0009593detection of chemical stimulus0.000697
GO:0010466negative regulation of peptidase activity0.000684
GO:0010951negative regulation of endopeptidase activity0.000675
GO:0016020membrane0.000673
GO:0016021integral component of membrane0.000652
GO:0016503pheromone receptor activity0.000644
GO:0017171serine hydrolase activity0.000644
GO:0019236response to pheromone0.000173
GO:0023052signaling0.0001413
GO:0030414peptidase inhibitor activity0.0001507
GO:0030545receptor regulator activity0.00041
GO:0031224intrinsic component of membrane0.00041
GO:0032501multicellular organismal process0.0001583
GO:0033038bitter taste receptor activity0.00027
GO:0038023signaling receptor activity0.00024
GO:0042221response to chemical0.0001644
GO:0042742defense response to bacterium0.00029
GO:0044425membrane part0.0001726
GO:0048018receptor ligand activity0.00026
GO:0050789regulation of biological process0.00026
GO:0050794regulation of cellular process0.00032
GO:0050877nervous system process0.00057
GO:0050896response to stimulus0.00054
GO:0050906detection of stimulus involved in sensory perception0.00054
GO:0050907detection of chemical stimulus involved in sensory perception0.00013
GO:0050909sensory perception of taste0.00040
GO:0050911detection of chemical stimulus involved in sensory perception of smell0.00190
GO:0050912detection of chemical stimulus involved in sensory perception of taste0.00157
GO:0050913sensory perception of bitter taste0.00194
GO:0051606detection of stimulus0.00163
GO:0051716cellular response to stimulus0.00260
GO:0060089molecular transducer activity0.01023
GO:0061134peptidase regulator activity0.01154
GO:0061135endopeptidase regulator activity0.01449
GO:0065007biological regulation0.02528
GO:0071944cell periphery0.04353
Table 6. Differentially expressed genes enriched in GO terms in stress-exposed rats after four weeks of lithium treatment.
Table 6. Differentially expressed genes enriched in GO terms in stress-exposed rats after four weeks of lithium treatment.
GO AccessionGO TermCorr pNo. Genes
Amygdala upregulated
GO:0038023signaling receptor activity0.00063
GO:0060089molecular transducer activity0.00063
GO:0004888transmembrane signaling receptor activity0.00059
GO:0007600sensory perception0.00054
GO:0050907detection of chemical stimulus involved in sensory perception0.00043
GO:0007606sensory perception of chemical stimulus0.00045
GO:0009593detection of chemical stimulus0.00043
GO:0051606detection of stimulus0.00045
GO:0050906detection of stimulus involved in sensory perception0.00043
GO:0007608sensory perception of smell0.00041
GO:0004984olfactory receptor activity0.00040
GO:0050911detection of chemical stimulus involved in sensory perception of smell0.00040
GO:0007186G-protein coupled receptor signaling pathway0.00056
GO:0004871signal transducer activity0.00263
GO:0003008system process0.00363
GO:0050877nervous system process0.00355
GO:0004930 G-protein coupled receptor activity0.00346
Frontal cortex upregulated
GO:0000786nucleosome0.0005
GO:0006334nucleosome assembly0.0005
GO:0044815DNA packaging complex0.0005
GO:0045653negative regulation of megakaryocyte differentiation0.0003
GO:0031497chromatin assembly0.0005
GO:0006333chromatin assembly or disassembly0.0005
GO:0034728nucleosome organization0.0005
GO:0006323DNA packaging0.0005
GO:0065004protein-DNA complex assembly0.0005
GO:0006335DNA replication-dependent nucleosome assembly0.0003
GO:0034723DNA replication-dependent nucleosome organization0.0003
GO:0032993protein-DNA complex0.0005
GO:0071824protein-DNA complex subunit organization0.0005
GO:0000788nuclear nucleosome0.0003
GO:0006336DNA replication-independent nucleosome assembly0.0003
GO:0045652regulation of megakaryocyte differentiation0.0003
GO:0034724DNA replication-independent nucleosome organization0.0003
GO:0071103DNA conformation change0.0015
GO:0051290protein heterotetramerization0.0023
GO:0051291protein heterooligomerization0.0064
GO:0030492hemoglobin binding0.0082
GO:0006352DNA-templated transcription. initiation0.0183
GO:0045638negative regulation of myeloid cell differentiation0.0183
GO:0000785chromatin0.0475
Hippocampus upregulated
GO:0005578proteinaceous extracellular matrix0.02210
GO:0031012extracellular matrix0.00913
Table 7. Differentially expressed genes enriched in GO terms in amphetamine-exposed rats responding to chronic lithium in comparison to rats before lithium.
Table 7. Differentially expressed genes enriched in GO terms in amphetamine-exposed rats responding to chronic lithium in comparison to rats before lithium.
Go AccessionGO TermCorr pNo. Genes
upregulated in the amygdala
GO:0007606sensory perception of chemical stimulus0.000232
GO:0009593detection of chemical stimulus0.000220
GO:0050907detection of chemical stimulus involved in sensory perception0.000216
GO:0051606detection of stimulus0.000232
GO:0050906detection of stimulus involved in sensory perception0.000220
GO:0004930G-protein coupled receptor activity0.000252
GO:0007608sensory perception of smell0.000210
GO:0004984olfactory receptor activity0.000205
GO:0050911detection of chemical stimulus involved in sensory perception of smell0.000205
GO:0007186G-protein coupled receptor signaling pathway0.000291
GO:0007600sensory perception0.000261
GO:0004888transmembrane signaling receptor activity0.000293
GO:0038023signaling receptor activity0.000306
GO:0060089molecular transducer activity0.000306
GO:0004871signal transducer activity0.000317
GO:0050877nervous system process0.000275
GO:0003008system process0.000306
GO:0031224intrinsic component of membrane0.000556
GO:0016021integral component of membrane0.000548
GO:0007165signal transduction0.000436
GO:0032501multicellular organismal process0.000596
GO:0007154cell communication0.000464
GO:0023052signaling0.000452
GO:0044425membrane part0.000596
GO:0005886plasma membrane0.000447
GO:0042221response to chemical0.000429
GO:0071944cell periphery0.000455
GO:0030414peptidase inhibitor activity0.00035
GO:0005549odorant binding0.00150
GO:0004252serine-type endopeptidase activity0.00134
GO:0004866endopeptidase inhibitor activity0.00132
GO:0061135endopeptidase regulator activity0.00332
GO:0061134peptidase regulator activity0.00535
GO:0010466negative regulation of peptidase activity0.01138
GO:0008236serine-type peptidase activity0.01934
GO:0017171serine hydrolase activity0.02534
GO:0050896response to stimulus0.026631
GO:003024carbohydrate binding0.04138
GO:0010951negative regulation of endopeptidase activity0.04435
GO:0004869cysteine-type endopeptidase inhibitor activity0.04615
upregulated in the frontal cortex
GO:0004930G-protein coupled receptor activity0.00044
GO:0004888transmembrane signaling receptor activity0.00049
GO:0007186G-protein coupled receptor signaling pathway0.00049
GO:0038023signaling receptor activity0.00151
GO:0060089molecular transducer activity0.00151
GO:0004871signal transducer activity0.00353
downregulated in the hippocampus
GO:0007186G-protein coupled receptor signaling pathway0.00087
GO:0004888transmembrane signaling receptor activity0.00085
GO:0038023signaling receptor activity0.00089
GO:0060089molecular transducer activity0.00089
GO:0004930G-protein coupled receptor activity0.00068
GO:0004871|signal transducer activity0.00092
GO:0007600sensory perception0.00367
GO:0050906detection of stimulus involved in sensory perception0.00754
GO:0050907detection of chemical stimulus involved in sensory perception0.00852
GO:0051606detection of stimulus0.00956
GO:0007606sensory perception of chemical stimulus0.01155
GO:0009593detection of chemical stimulus0.01152
GO:0003008system process0.01186
GO:0004984olfactory receptor activity0.01449
GO:0050911detection of chemical stimulus involved in sensory perception of smell0.01449
GO:0007608sensory perception of smell0.03149
GO:0050877nervous system process0.07171
Table 8. Differentially expressed genes enriched in GO terms in stress-exposed rats responding to chronic lithium treatment in comparison to rats before lithium.
Table 8. Differentially expressed genes enriched in GO terms in stress-exposed rats responding to chronic lithium treatment in comparison to rats before lithium.
Go AccessionGO TermCorr pNo. Genes
downregulated in the amygdala
GO:0044425membrane part0.0001150
GO:0032501|multicellular organismal process0.0001107
GO:0031224intrinsic component of membrane0.0001098
GO:0016021integral component of membrane0.0001090
GO:0007154cell communication0.000941
GO:0023052signaling0.000930
GO:0007165signal transduction0.000909
GO:0042221response to chemical0.000895
GO:0071944cell periphery0.000881
GO:0005886plasma membrane0.000874
GO:0004871signal transducer activity0.000773
GO:0038023signaling receptor activity0.000762
GO:0060089molecular transducer activity0.000762
GO:0004888transmembrane signaling receptor activity0.000749
GO:0007186G-protein coupled receptor signaling pathway0.000736
GO:0003008system process0.000720
GO:0050877nervous system process0.000683
GO:0007600sensory perception0.000667
GO:0004930G-protein coupled receptor activity0.000652
GO:0007606sensory perception of chemical stimulus0.000632
GO:0051606detection of stimulus0.000593
GO:0050906detection of stimulus involved in sensory perception0.000586
GO:0009593detection of chemical stimulus0.000578
GO:0050907detection of chemical stimulus involved in sensory perception0.000578
GO:0007608sensory perception of smell0.000565
GO:0004984olfactory receptor activity0.000556
GO:0050911detection of chemical stimulus involved in sensory perception of smell0.000556
GO:0005549odorant binding0.000164
GO:0051716cellular response to stimulus0.0001009
GO:0050896response to stimulus0.0001174
GO:0016503pheromone receptor activity0.00048
GO:0019236response to pheromone0.00047
GO:0016020membrane0.0001232
GO:0050789regulation of biological process0.0001346
GO:0065007biological regulation0.0001417
GO:0050794regulation of cellular process0.0001286
GO:0033038bitter taste receptor activity0.00019
GO:0008527taste receptor activity0.00020
GO:0001580detection of chemical stimulus involved in sensory perception of bitter taste0.00021
GO:0050909sensory perception of taste0.00027
GO:0050912detection of chemical stimulus involved in sensory perception of taste0.00022
GO:0050913sensory perception of bitter taste0.00021
GO:0030414peptidase inhibitor activity0.00047
GO:0004866endopeptidase inhibitor activity0.00044
GO:0061135endopeptidase regulator activity0.00144
GO:0004252serine-type endopeptidase activity0.00145
GO:0008227G-protein coupled amine receptor activity0.00322
GO:0017171serine hydrolase activity0.00349
GO:0008236serine-type peptidase activity0.00448
GO:0061134peptidase regulator activity0.00647
GO:0004867serine-type endopeptidase inhibitor activity0.01625
GO:0019373epoxygenase P450 pathway0.03010
GO:0010466negative regulation of peptidase activity0.03750
downregulated in the frontal cortex
GO:0004930G-protein coupled receptor activity0.00050
GO:0007606sensory perception of chemical stimulus0.00045
GO:0009593detection of chemical stimulus0.00043
GO:0038023signaling receptor activity0.00061
GO:0060089molecular transducer activity0.00061
GO:0007600sensory perception0.00051
GO:0050907detection of chemical stimulus involved in sensory perception0.00042
GO:0007608sensory perception of smell0.00041
GO:0004984olfactory receptor activity0.00040
GO:0050911detection of chemical stimulus involved in sensory perception of smell0.00040
GO:0051606detection of stimulus0.00044
GO:0050906detection of stimulus involved in sensory perception0.00042
GO:0004888transmembrane signaling receptor activity0.00056
GO:0007186G-protein coupled receptor signaling pathway0.00154
GO:0003008system process0.00163
GO:0004871signal transducer activity0.00262
GO:0050877nervous system process0.00254
GO:0016021integral component of membrane0.006112
GO:0031224intrinsic component of membrane0.007113
upregulated in the hippocampus
GO:0004930G-protein coupled receptor activity0.000118
GO:0007606sensory perception of chemical stimulus0.000108
GO:0050907detection of chemical stimulus involved in sensory perception0.000102
GO:0009593detection of chemical stimulus0.000103
GO:0050906detection of stimulus involved in sensory perception0.000104
GO:0004984olfactory receptor activity0.00098
GO:0050911detection of chemical stimulus involved in sensory perception of smell0.00098
GO:0051606detection of stimulus0.000105
GO:0007608sensory perception of smell0.00098
GO:0004888transmembrane signaling receptor activity0.000129
GO:0038023signaling receptor activity0.000135
GO:0060089molecular transducer activity0.000135
GO:0007186G-protein coupled receptor signaling pathway0.000127
GO:0007600sensory perception0.000115
GO:0004871signal transducer activity0.000136
GO:0050877nervous system process0.000119
GO:0003008system process0.000127
GO:0016021integral component of membrane0.000217
GO:0031224intrinsic component of membrane0.000219
GO:0007165signal transduction0.000173
GO:0023052signaling0.000178
GO:0044425membrane part0.000225
GO:0007154cell communication0.000179
GO:0042221response to chemical0.000167
GO:0071944cell periphery0.000174
GO:0005886plasma membrane0.000171
GO:0032501multicellular organismal process0.000214
GO:0005549odorant binding0.00025
GO:0051716cellular response to stimulus0.000193
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