2025 Volume 48 Issue 1 Pages 6-10
Enhanced inflammatory and immune responses have been observed in patients with major depressive disorder, pointing to anti-inflammatory substances as potential seeds for developing novel antidepressants. Omega-3 polyunsaturated fatty acid metabolites, such as resolvin D and E series, maresins, and protectins (collectively known as specialized pro-resolving mediators) demonstrate anti-inflammatory effects. This study examined the antidepressant-like effects of maresin-1 (MaR1) on lipopolysaccharide (LPS)-induced depression-like behaviors in mice. Using the tail suspension test (TST) and the forced swim test (FST), we assessed depression-like behaviors 26 and 28 h after intraperitoneal injection of LPS (0.8 mg/kg), respectively. An open field test (OFT) was also conducted to evaluate locomotor activity 24 h after LPS injection. Intracerebroventricular (i.c.v.) injection of MaR1 (10 ng/mouse) immediately after the LPS challenge mitigated the increased immobility time in the TST and FST, without affecting locomotor activity in the OFT, indicating the preventive effects of MaR1 on LPS-induced depression-like behaviors. Furthermore, i.c.v. injection of MaR1 23 h after the LPS challenge reduced the immobility time in both tests, underscoring its therapeutic potential. These findings suggest that MaR1 could be a promising seed for developing novel antidepressants.
Major depressive disorder (MDD), one of the most prevalent psychiatric disorders, incurs significant personal and socioeconomic burdens.1) Current monoamine-based antidepressants face notable drawbacks, including limited efficacy and delayed effects,2) highlighting the need for new therapeutic mechanisms. Elevated inflammatory and immune responses in MDD patients, characterized by increased levels of pro-inflammatory cytokines3) and microglial activation,4) suggest that anti-inflammatory agents could be promising seeds for developing novel antidepressants. Omega-3 polyunsaturated fatty acid metabolites, such as resolvin D and E series, maresins and protectins, are noted for their anti-inflammatory actions and collectively named specialized pro-resolving mediators (SPMs).5,6) Our prior research demonstrated that intracerebroventricular (i.c.v.) injections of resolvin D1 (RvD1) or D2 (RvD2) improve lipopolysaccharide (LPS)-induced7) and chronic unpredictable stress (CUS)-induced8) depression-like behaviors in mice. Additionally, we have shown that resolvin E1 (RvE1) and E2 (RvE2) through i.c.v. injections significantly counteract LPS-induced depression-like behaviors.9) These findings suggest that SPMs target brain sites to elicit their antidepressant-like effects.
This study investigates whether i.c.v. injections of maresin-1 (MaR1), an SPM derived from docosahexaenoic acid, have antidepressant-like effects on LPS-induced depression-like behaviors in mice.
Male C57BL/6J mice (7 weeks, n = 69) were acquired from Japan SLC (Hamamatsu, Japan), and group-housed (4 per cage) at a consistent ambient temperature (22 ± 2°C) with a 12-h light/dark cycle, and had ad libitum access to food and water. All experiments received approval of the Institutional Animal Care and Use Committee at Kanazawa University.
DrugsMaR1, in a 100% ethanol solution, was purchased from Cayman Chemical (Ann Arbor, MI, U.S.A.) and kept at −80°C. Prior to use, this solution was diluted with phosphate-buffered saline (PBS) to achieve a final ethanol concentration of 2%. LPS (serotype 0127:B8; Sigma, St. Louis, MO, U.S.A.) was dissolved in sterile saline.
Stereotaxic SurgeryI.c.v. infusion was performed with slight modifications from previously described methods.10) Under chloral hydrate anesthesia (400 mg/kg, intraperitoneal [i.p.]), a 26-gauge guide cannula (Plastics One, Roanoke, VA, U.S.A.) was implanted above the lateral ventricle (0.3 mm posterior, 1.0 mm lateral, and 2.2 mm ventral to bregma).11) After surgery, each mouse received an i.p. injection of carprofen (5 mg/kg; Zoetis, Parsippany, NJ, U.S.A.), was housed individually, and allowed a recovery period of at least 7 d.
LPS Challenge and MaR1 TreatmentEach mouse was given an i.p. injection of either LPS (0.8 mg/kg) or saline, as previously described.7,9,12,13) To assess the preventive effects of MaR1, immediately after the LPS challenge, MaR1 (10 ng/5 µL) or vehicle (2% ethanol/PBS, 5 µL) was unilaterally injected into the right lateral ventricle at a rate of 2.5 µL/min using a 33-gauge injector (Plastics One) that protruded 0.6 mm beyond the tip of the guide cannula. The injector was left in place for an additional 1 min to prevent drug backflow. To test the therapeutic effects of MaR1, i.c.v. infusion of MaR1 or vehicle was performed 23 h after the LPS challenge.
Behavioral TestsThe open field test (OFT), tail suspension test (TST), and forced swim test (FST) were conducted 24, 26, and 28 h after the LPS challenge, respectively, as previously described.12,13) Briefly, for the OFT, each mouse was placed in a test box (L38 × W26 × H24 cm) for 10 min, and the total distance traveled was automatically measured using Smart 3.0 software (Panlab Harvard Apparatus, Holliston, MA, U.S.A.). In the TST, each mouse was suspended by its tail, which was attached to a hook with adhesive tape, and immobility duration was measured for 6 min in a blinded manner. Data from mice that climbed their tail (n = 2 mice) or fell off the hook (n = 1 mouse) during the test were excluded from the analysis. In the FST, each mouse was placed in a 4-L glass beaker (16 cm diameter, 24.5 cm height) filled with water (24 ± 1°C, 15 cm depth), and the immobility duration was recorded between 2 and 6 min by a blinded experimenter.
HistologyAfter behavioral assessments, histological analyses were conducted as previously described.10) Briefly, mice were decapitated, and their brains were rapidly removed and frozen in powdered dry ice. Coronal sections (50 µm) were prepared using a cryostat (CM3050S, Leica Biosystems, Wetzlar, Germany), mounted on slides, and stained with thionin to confirm infusion sites. Data from mice with incorrect i.c.v. injections were excluded from the analysis (n = 5 mice).
Statistical AnalysesData are presented as means ± standard error of the mean (S.E.M.). Analyses were performed by 2-way ANOVA followed by Tukey’s post hoc test using GraphPad Prism 9 software (GraphPad Software, La Jolla, CA, U.S.A.).
As previously reported,7,9) in vehicle-treated (i.c.v.) mice, an LPS challenge induced depression-like behaviors in the TST and FST, but did not affect locomotor activity in the OFT (Figs. 1B–1E). To evaluate the preventive effects of MaR1 on LPS-induced depression-like behaviors, i.c.v. infusion of MaR1 was performed immediately after the LPS challenge (Fig. 1A). MaR1 significantly reduced the LPS-induced increase in immobility time in both the TST (Fig. 1D) and FST (Fig. 1E), without affecting locomotor activity (Figs. 1B, 1C).
(A) Experimental timeline for LPS challenge (0.8 mg/kg, i.p.), i.c.v. infusion (MaR1 10 ng or vehicle), and behavioral testing. (B) Representative movement trajectories of mice in the OFT. (C) Locomotor activity in the OFT (interaction, F1, 25 = 2.00, p = 0.170; LPS, F1, 25 = 5.75, p = 0.0242; MaR1, F1, 25 = 0.115, p = 0.738). (D) Immobility time in the TST (interaction, F1, 23 = 26.3, p < 0.0001). The data from 2 LPS + vehicle-treated mice were excluded due to tail-climbing or falling off. E. Immobility time in the FST (interaction, F1, 25 = 17.3, p = 0.0003). In panels (C, D), and (E), open circles show individual data points, and bars and lines show means ± S.E.M. *** p < 0.001 (Tukey’s post hoc test). OFT: open field test; S.E.M.: standard error of the mean; FST: forced swim test; MaR1: maresin-1; TST: tail suspension test; LPS: lipopolysaccharide.
To assess the therapeutic effects of MaR1 on LPS-induced depression-like behaviors, i.c.v. infusion of MaR1 was performed 23 h after the LPS challenge (Fig. 2A). MaR1 significantly decreased the LPS-induced increase in immobility time in both the TST (Fig. 2D) and FST (Fig. 2E), without affecting locomotor activity (Figs. 2B, 2C).
(A) Experimental timeline for LPS challenge (0.8 mg/kg, i.p.), i.c.v. infusion (MaR1 10 ng or vehicle), and behavioral testing. (B) Representative movement trajectories of mice in the OFT. (C) Locomotor activity in the OFT (interaction, F1, 31 = 0.335, p = 0.567; LPS, F1, 31 = 5.30, p = 0.0282; MaR1, F1, 31 = 5.99, p = 0.0202). (D) Immobility time in the TST (interaction, F1, 30 = 9.77, p = 0.0039). The data from 1 LPS + MaR1-treated mouse was excluded due to tail-climbing. (E) Immobility time in the FST (interaction, F1, 31 = 3.11, p = 0.0876; LPS, F1, 31 = 4.66, p = 0.0388; MaR1, F1, 31 = 14.0, p = 0.0008). In panels (C, D), and (E), open circles show individual data points, and bars and lines show means ± S.E.M. *p < 0.05, **p < 0.01, ***p < 0.001 (Tukey’s post hoc test). S.E.M.: standard error of the mean; LPS: lipopolysaccharide; TST: tail suspension test; FST: forced swim test; MaR1 maresin-1; OFT: open field test.
LPS challenge activates pro-inflammatory cytokine signaling, leading to sickness behaviors that peak between 2 and 6 h post-injection. These behaviors, especially reduced locomotor activity, can complicate the assessment of depression-like behaviors, such as increased immobility time in the TST and FST. However, 24 h after LPS injection, sickness behaviors subside, while depression-like behaviors persist.14) Consistent with our previous reports,7,9) this study observed no significant difference in locomotor activity between the LPS-challenged and non-LPS-challenged groups 24 h after LPS injection, whereas immobility time in the TST and FST significantly increased at 26 and 28 h after LPS injection, respectively, in the absence of MaR1 treatment.
I.c.v. injection of MaR1 at 10 ng/mouse immediately after the LPS challenge inhibited the LPS-induced prolongation of immobility time in both the TST and FST, demonstrating the preventive effects of MaR1 on LPS-induced depression-like behaviors. Moreover, i.c.v. injection of MaR1 23 h after the LPS challenge reversed the LPS-induced prolongation of immobility time in both tests, indicating its therapeutic effects on LPS-induced depression-like behaviors. Recently, Shi et al. reported that simultaneous i.p. injection of MaR1 at 5 µg/kg (approx. 100 ng/mouse) improved LPS-induced depression-like behavior in the TST.15) However, it remains unclear whether the site of action of MaR1 in their study was the central nervous system or peripheral tissues, including blood cells. Our study confirms that MaR1 exerts its antidepressant-like effects by acting on the central nervous system, given that i.c.v. administration of MaR1 at approximately one-tenth the dose used in their study improved LPS-induced depression-like behaviors.
Increasing evidence shows that inflammatory responses, including elevated circulating pro-inflammatory cytokines, microglial activation, and leukocyte infiltration into the brain, play roles in the pathophysiology of depression and that anti-inflammatory agents have antidepressant-like effects.3,4,16) MaR1 is included in the category of SPMs, which are metabolites of omega-3 polyunsaturated fatty acids known for their anti-inflammatory and pro-resolving actions in various animal models of peripheral inflammation.5,6) Shi et al. also reported that simultaneous i.p. injection of MaR1 suppressed LPS-induced microglial activation and interleukin (IL)-1β mRNA expression 1 day after the LPS injection.15) Xian et al. demonstrated that i.c.v. injection of MaR1 at 1 ng/mouse reduced cerebral infarct volume and neurological impairments in a mouse model of cerebral ischemia/reperfusion injury.17) They also found that MaR1 administered within 10 min after reperfusion decreased cytokine production, NFκB activation, microglia activation, and neutrophil infiltration at 24 h after reperfusion. These findings indicate that peripherally or centrally administered MaR1 exerts an anti-inflammatory effect in the brains of pathological animal models, suggesting that the antidepressant-like effects of MaR1 administered immediately after LPS injection could be mediated by its anti-inflammatory effect. In a human study, Qiu et al. reported that adolescent patients with MDD exhibited lower serum levels of MaR1 and higher levels of IL-6 compared to a healthy control group. Treatment with the antidepressant drug fluoxetine increased the serum levels of MaR1 and IL-4 while decreasing levels of IL-6 and IL-1β.18) Furthermore, they observed that MaR1 levels negatively correlated with the severity of depression. These findings suggest that a reduced level of MaR1 may elevate pro-inflammatory cytokine levels and exacerbate the pathology of MDD.
Clinical studies have shown that ketamine produces rapid antidepressant effects.19) Preclinical studies have suggested that the rapid antidepressant-like effects of this drug are mediated by the brain-derived neurotrophic factor (BDNF)–TrkB–phosphoinositide 3-kinase (PI3K)–protein kinase B (Akt)–mammalian target of rapamycin complex 1 (mTORC1) signaling pathway.20) Ma et al. reported the rapid antidepressant-like effect of the combination treatment with fluoxetine and brexpiprazole, but not fluoxetine alone, on LPS-induced depression-like behaviors in the TST and FST.21) They also demonstrated that the combination treatment with these drugs ameliorated LPS-induced alterations in BDNF–TrkB signaling, whereas fluoxetine alone showed no such effect. These findings suggest that compounds that activate the BDNF–TrkB–PI3K–Akt-mTORC1 signaling pathway may have rapid antidepressant effects. We previously demonstrated that the rapid antidepressant-like effects of RvD1, RvD2, and RvE1 on LPS-induced depression-like behaviors were blocked by rapamycin, an inhibitor of mTORC1, suggesting the involvement of mTORC1 signaling in their rapid antidepressant-like effects.7,9) Since Wei et al. reported that MaR1 activated the PI3K–Akt–mTORC1 signaling pathway in dorsal root ganglion neurons,22) the rapid therapeutic effects of MaR1 on LPS-induced depression-like behaviors observed in this study may be mediated through this pathway.
In the current study, MaR1 showed both preventive and rapid therapeutic effects in a mouse model of depression. Although further research is necessary to clarify the molecular and cellular mechanisms underlying these effects, MaR1 may be a promising target for developing novel antidepressants.
This work was supported by JSPS KAKENHI Grant Number: JP19K07120 (S.D.) and Japan Agency for Medical Research and Development (AMED) Grant Numbers: 22gm0910012s0106 and JP24zf0127004 (M.M.).
The authors declare no conflict of interest.
