Guarnieri et al. Translational Psychiatry (2020) 10:33 https://doi.org/10.1038/s41398-020-0701-5 Translational Psychiatry ARTICLE Open Access Pro-neurogenic effect of fluoxetine in the olfactory bulb is concomitant to improvements in social memory and depressive-like behavior of socially isolated mice Leonardo O. Guarnieri1,2, Ana Raquel Pereira-Caixeta1, Daniel C. Medeiros2,NayaraS.S.Aquino3, Raphael E. Szawka3, EduardoM.A.M.Mendes 2,MárcioF.D.Moraes1,2 and Grace S. Pereira 1 Abstract Although loneliness is a human experience, it can be estimated in laboratory animals deprived from physical contact with conspecifics. Rodents under social isolation (SI) tend to develop emotional distress and cognitive impairment. However, it is still to be determined whether those conditions present a common neural mechanism. Here, we conducted a series of behavioral, morphological, and neurochemical analyses in adult mice that underwent to 1 week of SI. We observed that SI mice display a depressive-like state that can be prevented by enriched environment, and the antidepressants fluoxetine (FLX) and desipramine (DES). Interestingly, chronic administration of FLX, but not DES, was able to counteract the deleterious effect of SI on social memory. We also analyzed cell proliferation, neurogenesis, and astrogenesis after the treatment with antidepressants. Our results showed that the olfactory bulb (OB) was the neurogenic niche with the highest increase in neurogenesis after the treatment with FLX. Considering that after FLX 1234567890():,; 1234567890():,; 1234567890():,; 1234567890():,; treatment social memory was rescued and depressive-like behavior decreased, we propose neurogenesis in the OB as a possible mechanism to unify the FLX ability to counteract the deleterious effect of SI. Introduction and weight changes are common, though inconstant Even though humans are naturally social animals, physiological markers for depression8. Anatomically, choosing to be alone can provide a positive state of hippocampus and olfactory bulb (OB) are reduced in satisfaction. Conversely, loneliness, characterized by being some cases of depression9. Furthermore, depression is a non-temporary condition, is a subjective state with usually associated with disruption of episodic mem- – negative impacts on physical and mental health1 3. In fact, ory10,11, such as impaired recollection, exacerbated the consistent unpleasant experience of being alone can memory for negative content12, and the weakening of – even predict depressive symptomatology4 7. positive memories13,14. Nevertheless, the mechanisms of Depression is a mood disorder with heterogeneous memory disruption in depression are still poorly symptoms and etiology (American Psychiatric Associa- understood15. – tion, 1994). Despite the behavioral symptoms6 8, appetite The pharmacological treatment for depression includes fluoxetine, a selective serotonin (5-HT) reuptake inhi- bitor, and desipramine, a tricyclic antidepressant which Correspondence: Grace S. Pereira ([email protected]) 1Núcleo de Neurociências, Universidade Federal de Minas Gerais, Belo inhibits the reuptake of norepinephrine (NE), but also 5- Horizonte, Brazil HT16,17. Lower concentrations of NE and 5-HT in the 2 – Centro de Tecnologia e Pesquisa em Magneto Ressonância, Programa de Pós- brain18 20 along with the effectiveness of antidepressant Graduação em Engenharia Elétrica, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil Full list of author information is available at the end of the article. © The Author(s) 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a linktotheCreativeCommons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. Guarnieri et al. Translational Psychiatry (2020) 10:33 Page 2 of 14 drugs21,22 support the monoaminergic hypothesis of (CONTROL); group housed in enriched environment depression. (CONTROL + ENRICHMENT); individually housed in Alternatively, neurogenesis has arisen as a pivotal standard environment (SOCIAL ISOLATION), and mechanism in the pathogenesis of depression23. Unme- individually housed in enriched environment (SOCIAL dicated depressed adults presented less granule neurons ISOLATION + ENRICHMENT). Animals were main- in the anterior dentate gyrus (DG) when compared to tained in polypropylene cages (28 cm × 17 cm × 12 cm), healthy controls24, which agrees with reduced hippo- except the control + enrichment group that was main- campal volume findings observed in patients with major tained in bigger polypropylene cage (40 cm × 33 cm × depression25. Accordingly, neurogenesis has been pro- 16 cm). The enriched environment was provided by the posed by several independent researchers as one of the addition of ribbons, pieces of plastic, cardboard rolls and mechanisms for the antidepressant effect of toys into the cage. Animals were maintained in each fluoxetine26,27. condition during at least 7 days. Evidence suggest that loneliness is not a solely human All experiments were performed in compliance with the experience28. Objective loneliness can be induced in guidelines from the National Council for Animal Experi- laboratory animals by depriving them from physical mentation Control (CONCEA-BRAZIL). All protocols contact with conspecifics. Rearing mice on social isolation were approved by the Institutional Ethics Committee on (SI) may induce depressive, anxious and aggressive the Use of Animals at the Universidade Federal de Minas behaviors later on life29,30 and it is usually used as an Gerais (CEUA/UFMG) (no. 177/2014). animal model for stress31,32. SI during adulthood, how- The investigator was blinded to the group allocation ever, is less studied, although their behavioral effects has during the experiment and during assessing the out- been shown to depend on SI duration33. coming results. One emerging field that has been the focus of our research is to understand the effects of SI on a specific Metabolic evaluation type of episodic memory named social recognition Animals were weighted daily during 7 days. During the memory23,34. One week of SI is deleterious for the long- same period, the consumption of food and water was – term maintenance of social memory in mice35 37, measured. All these measurements were taken at 9:00 a.m. although did not affect other hippocampus-dependent memories36. Interestingly, enriched environment blunted Behavioral analysis the effect of SI on memory36 in a neurogenesis-dependent Forced-swimming test manner38. In fact, the formation of social recognition Animals were placed individually in a glass cylinder memory seems reliant on neurogenesis39,40. (35 cm high, 24 cm in diameter) filled with water (28 °C) Here we raised the hypothesis that SI is a condition that to the height of 14 cm. During 6 min, time of immobility induces episodic memory impairment and depressive-like was quantified41,42. behaviors, allowing to investigate the mechanisms of memory disruption in depression. This work examines the behavioral, pharmacological, morphological, and Tail suspension test neurochemical features of depression in isolated mice. We Mice were suspended by the tail and attached to a metal also tested the antidepressant potential of fluoxetine, rod (50 cm height) for a period of 6 min. After 1 min desipramine, and the enriched environment in the SI habituation, total immobility time was scored. Immobility model. Our study proposes neurogenesis as a core was considered as the absence of movements in the hin- mechanism involved in the antidepressant and promnesic dlimbs and forelimbs43,44. effects of fluoxetine and enriched environment. Material and methods Sucrose preference test Subjects Animals were individually placed in cages containing We used 164 adults (8–12 weeks of age) (30 females and two bottles, one filled with water and the other one with 134 males) and 36 juveniles (21–35 days of age, only used 3% sucrose solution. During the first 3 h, the bottles were as a social stimulus) Swiss mice. All animals were main- weighed every 1 h. After, bottles were weighed 12, 24, and tained in a climate-controlled environment (22 ± 2 °C, 48 h. Between each measure we changed the position of humidity at 55 ± 10%) under a 12 h light/dark cycle. All the bottle to avoid spatial cues. The preference for behavioral experiments were performed during the light sucrose over water was calculated by the following for- phase. Both food and water were available ad libitum. The mula: [sucrose consumed/(sucrose + water consumed)]. animals were randomly placed in one of the following Results were expressed as the percentage of sucrose groups: group housed in standard environment preference45. Guarnieri et al. Translational Psychiatry (2020) 10:33 Page 3 of 14 Social recognition for 20 min at 12,000g. Protein content was determined Habituation phase consisted in introducing the adult from the pellet