Molecular Profiling of the Lateral Habenula in a Rat Model of Depression

<div><p>Objective</p><p>This study systematically investigated the effect of chronic mild stress and response to antidepressant treatment in the lateral habenula at the whole genome level.</p><p>Methods</p><p>Rat whole genome expression chips (Affymetrix) were used to detect gene expression regulations in the lateral habenula of rats subjected to chronic mild stress (mild stressors exchanged twice a day for 8 weeks). Some rats received antidepressant treatment during fifth to eights week of CMS. The lateral habenula gene expression profile was studied through the gene ontology and signal pathway analyses using bioinformatics. Real-time quantitative polymerase chain reaction (RT-PCR) was used to verify the microarray results and determine the expression of the <i>Fcrla, Eif3k, Sec3l1, Ubr5, Abca8a, Ankrd49, Cyp2j10, Frs3, Syn2, and Znf503</i> genes in the lateral habenula tissue.</p><p>Results</p><p>In particular we found that stress and antidepressant treatment affected intracellular cascades like growth factor receptor signaling, G-protein-coupled receptor signaling, and Wnt signaling – processes involved in the neuroplastic changes observed during the progression of depression and antidepressant treatment.</p><p>Conclusion</p><p>The present study suggests an important role of the lateral habenula in the development of depression-like conditions and correlates to previous studies demonstrating a significant role of the lateral habenula in depressive-like conditions and antidepressant treatment.</p></div

Figure modified from the BRAIN ATLAS [31].

<p>Pictures show 13 different substructures analysed at 3 rostrocaudal levels, within each structure, throughout different brain regions. In grey areas affected in stress-susceptible rats; in dark grey with stripes brain regions affected in both CMS phenotypes; in light grey areas affected in stress resilient rats. In beige, areas not affected by the CMS. DG, dendate gyrus; Cg1, cingulate cortex—area 1; IL, infralimbic cortex; PrL, prelimbic cortex; LO, lateral orbital cortex; VO, ventral orbital cortex; Pir, piriform cortex; PVA, paraventricular thalamic nucleus—anterior part; BLA, basolateral amygdaloid nucleus—anterior part; MHb, medial habenula; LHb, lateral habenula; CA3, field CA3 of hippocampus; VLGMC, ventral lateral geniculate nucleus magnocellular part; * p < 0.05; ** p < 0.01; *** p < 0.001.</p

Neuronal substrates underlying stress resilience and susceptibility in rats

<div><p>Background</p><p>Stress and stressful life events have repeatedly been shown as causally related to depression. The Chronic Mild Stress rat model is a valid model of stress-induced depression. Like humans, rats display great heterogeneity in their response to stress and adversity. Hence some individuals are stress-sensitive and prone to develop depression-like behaviour in response to modest stressors, while others are stress-resilient and remain essentially symptom free.</p><p>Objectives</p><p>Compared to the large body of research, which describes stress-induced maladaptive neurobiological changes, relatively little attention has been devoted to understand resiliency to stress. The aim of the present study was to identify changes in neuronal activity, associated with stress-resilient and stress-susceptible chronic mild stress endophenotypes, by examining <i>c-Fos</i> expression in 13 different brain areas. Changes in <i>c-Fos</i> expression have been reported as associated to stressful conditions.</p><p>Methods</p><p>Stress-induced modulation of neuronal activation patterns in response to the chronic mild stress paradigm was mapped using the immediate early gene expression <i>c-Fos</i> as a marker. Quantification of the <i>c-Fos</i>-like immunoreactivity responses was done by semi-automated profile counting procedures and design-based stereology.</p><p>Results</p><p>Exposure to chronic mild stress significantly altered <i>c-Fos</i> expression in a total of 6 out of 13 investigated areas. Chronic mild stress was found to suppress the <i>c-Fos</i> response within the magnocellular ventral lateral geniculate nucleus of both stress subgroups. In the the lateral and ventral orbital cortices of stress-resilient rats, the <i>c-Fos</i> like immunoreactivity response was also repressed by stress exposure. On the contrary the <i>c-Fos</i> response within the amygdala, medial habenula, and infralimbic cortex was increased selectively for the stress-susceptible rats.</p><p>Conclusions</p><p>The study was initiated to characterize neuronal substrates associated with stress-coping mechanisms. Six areas, all of which represents limbic structures, were found to be sensitive to stress exposure. The effects within these areas associate to the hedonic status of the rats. Hence, these areas might be associated to stress-coping mechanisms underlying the chronic mild stress induced segregation into stress-susceptible and stress-resilient endophenotypes.</p></div

Stereological estimates of total c-Fos-ir in the habenular complexes.

<p>The estimates of total c-Fos-ir in the lateral (A) and medial (B) habenular complexes analysed with stereology. In plot A and B number of cells counted in the LHb as the sum of dark and medial c-Fos-ir and the sum of dark, medial and light c-Fos-ir, respectively. In plot C and D number of cells counted in the MHb as the sum of dark and medial c-Fos-ir and the sum of dark, medial and light c-Fos-ir, respectively. One-way ANOVA. LHb, lateral habenula; MHb, medial habenula.</p

Results summary.

<p>Results summary.</p

Animal groups investigated in the present CMS study.

<p>n – number of rats in each group resulting in 3 subgroups per CMS group.</p

Relationships between groups built up for PANTHER analysis.

<p>Statistical enrichment of biological pathways was assessed by binomial statistics within the PANTHER system. (A and B) Relationships for classification of pathways implicated in depression and recovery; (C and D) Relationships for classification of pathways implicated in treatment response. Dashed lines indicate significant differences in pathways between groups (<i>p</i><0.05), regular lines indicate no differences in pathways between groups. U-V, unchallenged control rats; CMS-V, CMS vehicle; Esc-R, CMS escitalopram responders; Esc-NR, CMS escitalopram non-responders.</p

Sucrose consumption following four weeks of the CMS regime.

<p>Four weeks exposure to chronic mild stressors resulted in a significant decrease of sucrose consumption of the anhedonic animals (n = 10) when compared to unchallenged controls (n = 10). The graph shows sucrose consumption of the control (n = 10), resilient (n = 10) and anhedonic-like (n = 10) animals at 4 different time points (week 1; week 2; week 3 and week 4). Significant differences on sucrose consumptions were detected among goups (p<0,0001) at different time points (p = 0,0084). The sucrose intake of anhedonic-like animals was significantly diminished when compared to control and resilient animals. Two-way ANOVA followed by post hoc Tukey´s multiple comparison test. * p<0,05; **p<0,01; ***p<0,001. Data is presented as mean (±SEM) sucrose intake, indexed to baseline values.</p

Study design showing the experimental outline and time course.

<p>(modified from [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0179434#pone.0179434.ref070" target="_blank">70</a>]).</p

Pathway categories implicated in recovery.

<p>U-V, unchallenged control rats; CMS-V, CMS vehicle; Esc-R, CMS escitalopram responders; Esc-NR, CMS escitalopram non-responders.</p