Fish-oil supplementation decreases Indoleamine-2,3-Dioxygenase expression and increases hippocampal serotonin levels in the LPS depression model
Introduction
Depression is a common mental disorder with more than 320 million people affected, and it is the leading cause of disability worldwide [1]. The pathophysiology of this disorder is multifactorial, and a growing body of evidence shows a bidirectional relationship between depression and inflammation. Evidence for this relationship comes from findings that depressed patients display increased rate of autoimmune disorders, whereas patients with inflammatory diseases, like cancer, HIV and rheumatoid arthritis have higher rates of depression [2]. Furthermore, increased levels of pro-inflammatory cytokines, such as interleukin 1β (IL-1β), IL-6 and tumor necrosis factor α (TNF-α) have been found in depressed patients [3,4]. Systemic injection of bacterial endotoxin lipopolysaccharide (LPS), which is the major component of the outer membrane of Gram-negative bacteria, produces an acute inflammatory response in rodents and has been used as a model of depressive-like behavior [[5], [6], [7]]. One possible pathway linking inflammation and depression involves pro-inflammatory cytokines-induced low serotonin (5-HT) levels, by activation of the enzyme Indoleamine-2,3-Dioxygenase (IDO). This enzyme catabolizes the 5-HT precursor, tryptophan, into kynurenines, resulting in a depressive state [8,9].
Omega-3 Polyunsaturated Fatty Acids (ω-3 PUFAs) are critical components of biological membranes and play key roles in cell integrity, homeostasis, development and function [10]. Some studies show lower ω-3 PUFAs and higher ω-6 to ω-3 PUFAs ratio in depressive patients’ blood and brains [[11], [12], [13], [14], [15], [16]]. Fish oil (FO) supplementation (rich in ω-3 PUFAs) during pregnancy and lactation has a preventive antidepressant effect in the adult offspring by increasing hippocampal 5-HT levels [[17], [18], [19]]. Interestingly, ω-3 PUFAs also exhibit anti-inflammatory effects, especially regulating cytokines and chemokines, and decreasing inflammatory prostaglandins and eicosanoids [20].
Given the evidence of the involvement of IDO pathway in depression, we hypothesized that ω-3 PUFAs antidepressant activity in the LPS model would be mediated by an inhibitory effect on this enzyme. In order to test this hypothesis, animals were supplemented with FO or received water (Control group – C). Both groups were subjected to an immune challenge with systemic LPS injection 24 h before being tested in the Modified Forced Swim (MFST) and Open Field Tests. This study was performed in two independent experiments, using two different IDO inhibitors, the direct inhibitor 1-methyl-DL-tryptophan (1-MT) and Minocycline (MINO) – that is considered an indirect IDO inhibitor since it has anti-inflammatory properties [7,21] – both administered 23 h, 5 h and 1 h before the behavioral tests. After the tests, the hippocampi were removed for quantification of 5-HT and its metabolite 5-HIAA by HPLC, and IDO expression was determined by Western blot.
Section snippets
Animals
Two hundred and forty-three, 60 day-old, male Wistar rats were kept under a 12 h light/12 h dark cycle (lights on at 07:00 am), in a room under controlled humidity, temperature (21 ± 2 °C), and air exchange, with food (rat chow, Nuvital Nuvilab CR1- Nuvital Nutrientes S/A, Colombo, Paraná, Brazil) and water ad libitum. All animals (four rats/cage) were housed in plastic cages (41 × 32 × 16.5 cm), filled with sawdust bedding, which was changed 3 times a week. All rats were tested when they were
Experiment 1
For BW gain, three-way ANOVA revealed an effect of group [F(1,107) = 21.8; p ≤ 0.00009], immune challenge [F(1,107) = 138.94; p ≤ 0.0000001] and an interaction between group and immune challenge [F(1,107) = 25.6; p ≤ 0.000002]. Tukey’s post hoc test showed that LPS led to a pronounced weight loss (p ≤ 0.0001) and FO attenuated this effect (p ≤ 0.0002). There was no effect of IDO blocker [F(1,107) = 0.16; n.s.] nor interactions between group and IDO blocker [F(1,107) = 0.3; n.s.], immune
Discussion
The major finding of this study was the pronounced antidepressant-like effect of FO related to increased hippocampal levels of 5-HT and decreased IDO expression. These effects were observed in LPS-injected animals, highlighting the important protective effect of ω-3 PUFAs in inflammation-associated depression.
A single systemic injection of LPS is a model widely employed to investigate depressive-like behavior in rodents, which induces physiological and behavioral changes known as sickness
Declaration of competing interest
The authors declare they have no conflicts of interest to disclose.
CRediT authorship contribution statement
Bruno Carabelli: Investigation, Conceptualization, Methodology, Visualization, Formal analysis, Writing - original draft, Writing - review & editing. Ana Márcia Delattre: Investigation. Ana Paula Farias Waltrick: Investigation. Giulia Araújo: Investigation. Deborah Suchecki: Investigation, Resources, Writing - original draft, Writing - review & editing. Ricardo Borges Machado: Investigation. Luiz Eduardo Rizzo de Souza: Investigation. Silvio M. Zanata: Resources. Janaína Menezes Zanoveli:
Acknowledgments
This study was supported by grants from CNPq (Casadinho/PROCAD Grant No.552226/2011-4) and CAPES. The authors are indebted to Laboratório Herbarium Botânico S/A, which kindly donated the fish oil capsules rich in DHA and EPA. Deborah Suchecki is the recipient of a Research Fellowship from CNPq (Grant No. 303449/2015-2).
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