Environmental enrichment: dissociated effects between physical activity and changing environmental complexity on anxiety and neurogenesis in adult male Balb/C mice.

Several factors, including environmental modifications, stimulate neuroplasticity. One type of neuroplasticity consists in the generation of new neurons in the dentate gyrus of the hippocampus. Neurogenesis is modulated by environmental enrichment (ENR, tunnels plus running wheel) and affected by the time of exposure to ENR. Despite the wide use of ENR to stimulate neuroplasticity, the degree to which ENR variations modeled by temporally changing the level of environmental complexity affect hippocampal neurogenesis and anxiety is still unclear. Thus, we investigated the effects of five housing conditions on young adult male Balb/C mice exposed for 42 days. The groups were as follows: standard conditions without ENR, constant ENR complexity, gradual increase of ENR complexity followed by a gradual decrease of ENR complexity, gradual increase of ENR complexity followed by constant ENR complexity, and constant ENR complexity followed by a gradual decrease of ENR complexity. On day 44, mice were exposed to the elevated plus-maze to evaluate anxiety. Further, we analyzed neurogenesis and quantified corticosterone levels. In an additional experiment, we explored the effect of voluntary physical activity on anxiety, neurogenesis, and corticosterone during the variations in ENR complexity. Our results showed that any change in ENR complexity over time reduced anxiety. Also, voluntary physical activity alone or in the context of a complex environment increased doublecortin cell maturation in the granular cell layer of the hippocampus. Finally, our study supports that physical activity acts proneurogenic, whereas any change in environmental complexity decreases anxiety-like behavior. However, the decrease in corticosterone levels elicited by physical activity was lower than the decrease produced by the decrement in environmental complexity.

Environmental Enrichment Differentially Activates Neural Circuits in FVB/N Mice, Inducing Social Interaction in Females but Agonistic Behavior in Males.

Environmental enrichment induces behavioral and structural modifications in rodents and influences the capability of mice to cope with stress. However, little is understood about hippocampal neurogenesis and the appearance of social/agonistic (aggressive) behavior upon activation of different neuronal circuits in FVB/N mice. Thus, in this study we hypothesized that environmental enrichment differentially regulates neurogenesis, neural circuit activation and social/agonistic behavior in male and female FVB/N mice. We explored the (1) neurogenic process as an indicative of neuroplasticity, (2) neuronal activation in the limbic system, and (3) social behavior using the resident-intruder test. On postnatal day 23 (PD23), mice were assigned to one of two groups: Standard Housing or Environmental Enrichment. At PD53, rodents underwent the resident-intruder test to evaluate social behaviors. Results revealed that environmental enrichment increased neurogenesis and social interaction in females. In males, environmental enrichment increased neurogenesis and agonistic behavior. Enriched male mice expressed higher levels of agonistic-related behavior than female mice housed under the same conditions. Neural circuit analysis showed lower activation in the amygdala of enriched males and higher activation in enriched females than their respective controls. Enriched females also showed higher activation in the frontal cortex without differences in male groups. Moreover, the insular cortex was less activated in females than in males. Thus, our results indicate that environmental enrichment has different effects on neuroplasticity and social/agonistic behavior in FVB/N mice, suggesting the relevance of sexual dimorphism in response to environmental stimuli.

Short Daily Exposure to Environmental Enrichment, Fluoxetine, or Their Combination Reverses Deterioration of the Coat and Anhedonia Behaviors with Differential Effects on Hippocampal Neurogenesis in Chronically Stressed Mice.

Depression is a neuropsychiatric disorder with a high impact on the worldwide population. To overcome depression, antidepressant drugs are the first line of treatment. However, pre-clinical studies have pointed out that antidepressants are not entirely efficacious and that the quality of the living environment after stress cessation may play a relevant role in increasing their efficacy. As it is unknown whether a short daily exposure to environmental enrichment during chronic stress and antidepressant treatment will be more effective than just the pharmacological treatment, this study analyzed the effects of fluoxetine, environmental enrichment, and their combination on depressive-associated behavior. Additionally, we investigated hippocampal neurogenesis in mice exposed to chronic mild stress. Our results indicate that fluoxetine reversed anhedonia. Besides, fluoxetine reversed the decrement of some events of the hippocampal neurogenic process caused by chronic mild stress. Conversely, short daily exposure to environmental enrichment changed the deterioration of the coat and anhedonia. Although, this environmental intervention did not produce significant changes in the neurogenic process affected by chronic mild stress, fluoxetine plus environmental enrichment showed similar effects to those caused by environmental enrichment to reverse depressive-like behaviors. Like fluoxetine, the combination reversed the declining number of Ki67, doublecortin, calretinin cells and mature newborn neurons. Finally, this study suggests that short daily exposure to environmental enrichment improves the effects of fluoxetine to reverse the deterioration of the coat and anhedonia in chronically stressed mice. In addition, the combination of fluoxetine with environmental enrichment produces more significant effects than those caused by fluoxetine alone on some events of the neurogenic process. Thus, environmental enrichment improves the benefits of pharmacological treatment by mechanisms that need to be clarified.

Apple Peel and Flesh Contain Pro-neurogenic Compounds.

As mammals evolved with exposure to particular diets, naturally abundant compounds may have become part of the set of environmental co-determinants that shaped brain structure and function. Here we investigated whether bioactive factors found in apples directly affect hippocampal neurogenesis in the adult mouse. We found that quercetin, the most abundant flavanol in apple peel, was anti-proliferative at high concentrations but pro-neurogenic at low concentrations. This was confirmed in vivo, with intraperitoneally delivered quercetin promoting survival and neuronal differentiation, without affecting proliferation. Using a bioassay-guided fractionation approach we also identified additional pro-neurogenic compounds in apple flesh that were not related to flavonoids. We found that 3,5-dihydroxybenzoic acid significantly increased neural precursor cell proliferation and neurogenesis. This work shows that both flavonoids and 3,5-dihydroxybenzoic acid are pro-neurogenic, not only by activating precursor cell proliferation but also by promoting cell-cycle exit, cellular survival, and neuronal differentiation.

Melatonin Reverses the Depression-associated Behaviour and Regulates Microglia, Fractalkine Expression and Neurogenesis in Adult Mice Exposed to Chronic Mild Stress.

Depression may be precipitated by the negative impact of chronic stress, which is considered to play a key role in this neuropsychiatric disorder. Interestingly, depressed patients show decreased levels of melatonin. This hormone acts pro-neurogenic and exhibits anti-depressant effects in rodent models of predictive antidepressant-like effects. However, the benefits of melatonin in reversing the deleterious effects of chronic mild stress on the alterations in behaviour and in the neurogenic niche of the hippocampus in male BALB/c mice are unknown. In this study, we compared the effects of melatonin (2.5 mg/kg) and citalopram (5 mg/kg), an antidepressant drug belonging to the selective serotonin reuptake inhibitors, in male BALB/c mice exposed to chronic mild stress (CMS). We also investigated the potential effects of melatonin and citalopram on microglial cells, hippocampal neurogenesis and peripheral cytokine profiles. Melatonin and citalopram induced similar antidepressant-like activities that occurred with some of the the following findings: (1) reversal of the morphological alterations in microglia; (2) reversal of the decreased immunoreactivity to CX3CL1 and CX3CR1 in the dentate gyrus; (3) positive regulation of cell proliferation, survival and complexity of the dendritic trees of doublecortin-cells; and (4) modifications of peripheral CX3CL1 expression. This outcome is consistent with the hypothesis about the antidepressant-like effect of melatonin and supports its relevance as a modulator of the niche in the dentate gyrus.

Melatonin Modulates Dendrite Maturation and Complexity in the Dorsal- and Ventral- Dentate Gyrus Concomitantly with Its Antidepressant-Like Effect in Male Balb/C Mice.

Adult neurogenesis occurs in the dentate gyrus (DG) of the hippocampus. New neurons help to counteract the effects of stress and several interventions including antidepressant drugs, environmental modifications and internal factors act pro-neurogenic with consequences in the dorsal and ventral DG. Melatonin, the main product synthesized by the pineal gland, induces antidepressant-like effects and modulates several events of the neurogenic process. However, the information related to the capability of melatonin to modulate dendrite maturation and complexity in the dorsal and ventral regions of the DG and their correlation with its antidepressant-like effect is absent. Thus, in this study, we analyzed the impact of melatonin (0, 0.5, 1, 2.5, 5 or 10 mg/kg) administered daily for fourteen days on the number, dendrite complexity and distribution of doublecortin (DCX)-cells in the dorsal-ventral regions of the DG in male Balb/C mice. Doublecortin is a microtubule-associated protein that is expressed during the course of dendritic maturation of newborn neurons. Also, we analyzed the impact of melatonin on despair-like behavior in the forced swim test. We first found a significant increase in the number and higher dendrite complexity, mainly with the doses of 2.5, 5 and 10 mg/kg of melatonin (81%, 122%, 78%). These cells showed more complex dendritic trees in the ventral- and the dorsal- DG. Concomitantly, the doses of 5 and 10 mg/kg of melatonin decreased depressant-like behavior (76%, 82%). Finally, the data corroborate the antidepressant-like effect of melatonin and the increasing number of doublecortin-associated cells. Besides, the data indicate that melatonin favors the number and dendrite complexity of DCX-cells in the dorsal- and ventral- region of the DG, which may explain part of the antidepressant-like effect of melatonin.

Melatonin Influences Structural Plasticity in the Axons of Granule Cells in the Dentate Gyrus of Balb/C Mice.

Melatonin, the main product synthesized by the pineal gland, acts as a regulator of the generation of new neurons in the dentate gyrus (DG). Newborn neurons buffer the deleterious effects of stress and are involved in learning and memory processes. Furthermore, melatonin, through the regulation of the cytoskeleton, favors dendrite maturation of newborn neurons. Moreover, newborn neurons send their axons via the mossy fiber tract to Cornu Ammonis 3 (CA3) region to form synapses with pyramidal neurons. Thus, axons of newborn cells contribute to the mossy fiber projection and their plasticity correlates with better performance in several behavioral tasks. Thus, in this study, we analyzed the impact of exogenous melatonin (8 mg/kg) administered daily for one- or six-months on the structural plasticity of infrapyramidal- and suprapyramidal mossy fiber projection of granule cells in the DG in male Balb/C mice. We analyzed the mossy fiber projection through the staining of calbindin, that is a calcium-binding protein localized in dendrites and axons. We first found an increase in the number of calbindin-positive cells in the granular cell layer in the DG (11%, 33%) after treatment. Futhermore, we found an increase in the volume of suprapyramidal (>135%, 59%) and infrapyramidal (>128%, 36%) mossy fiber projection of granule neurons in the DG after treatment. We also found an increase in the volume of CA3 region (>146%, 33%) after treatment, suggesting that melatonin modulates the structural plasticity of the mossy fiber projection to establish functional synapses in the hippocampus. Together, the data suggest that, in addition to the previously reported effects of melatonin on the generation of new neurons and its antidepressant like effects, melatonin also modulates the structural plasticity of axons in granule cells in the DG.

Newly Generated Cells in the Dentate Gyrus Differentially Respond to Brief Spatial Exploration and Forced Swim in Adult Female Balb/C Mice.

Neurogenesis in the hippocampus is influenced by several factors including external stimuli. In addition to their involvement in learning and memory processes, newborn neurons of the dentate gyrus (DG) buffer against the effects of stress. Although the response of these cells to environmental stimuli has been shown, the age of the cells that respond to a brief spatial exploration or a stressful situation produced by forced-swim stress in adult female Balb/C mice is still unknown. Here, we investigated the activation of newborn neurons after three (IdU) or six weeks (CldU) postlabelling with the expression of Arc in the same mice but exposed to different environmental stimuli. Mice housed in standard conditions showed an increase in the activation of CldU-labelled cells after two exposures to a brief spatial exploration but no increase in the activation of IdU-labelled cells compared with the control group. Additionally, we analysed neuronal activation in the DG of mice housed in standard conditions and further exposed to forced-swim stress. We found a decreased activation of IdU-labelled cells in mice exposed to forced-swim stress with increase number of CldU-labelled cells. Our results suggest that based on their time postlabelling, newly generated hippocampal neurons show a different response to several environmental stimuli.

Soluble Factors from Human Olfactory Neural Stem/Progenitor Cells Influence the Fate Decisions of Hippocampal Neural Precursor Cells.

Neurogenesis plays a significant role during adulthood, and the observation that neural stem cells reside in the central nervous system and the olfactory epithelium has attracted attention due to their importance in neuronal regeneration. In addition, soluble factors (SFs) release by neural stem cells may modulate the neurogenic process. Thus, in this study, we identified the SFs released by olfactory human neural stem/progenitor cells (hNS/PCs-OE). These cells express Ki67, nestin, and ßIII-tubulin, indicating their neural lineage. The hNS/PCs-OE also express PSD95 and tau proteins during proliferation, but increased levels are observed after differentiation. Thus, we evaluated the effects of SFs from hNS/PCs-OE on the viability, proliferation, and differentiation potential of adult murine hippocampal neural precursor cells (AHPCs). SFs from hNS/PCs-OE maintain cells in the precursor and proliferative stages and mainly promote the astrocytic differentiation of AHPCs. These effects involved the activation, as measured by phosphorylation, of several proteins (Erk1/2; Akt/PRAS40/GSK3ß and JAK/STAT) involved in key events of the neurogenic process. Moreover, according to the results from the antibody-based microarray approach, among the soluble factors, hNS/PCs-OE produce interleukin-6 (IL-6) and neurotrophin 4 (NT4). However, residual epidermal growth factor (EGF) was also detected. These proteins partially reproduced the effects of SFs from hNS/PCs-OE on AHPCs, and the mechanism underlying these effects is mediated by Src proteins, which have been implicated in EGF-induced transactivation of TrkB receptor. The results of the present study suggest the potential use of SFs from hNS/PCs-OE in controlling the differentiation potential of AHPCs. Thus, the potential clinical relevance of hNS/PCs-OE is worth pursuing.

Exposure to Patterned Auditory Stimuli during Acute Stress Prevents Despair-Like Behavior in Adult Mice That Were Previously Housed in an Enriched Environment in Combination with Auditory Stimuli.

Several interventions have been shown to counteract the effects of stress that may be related to improved neuroplasticity and neuronal activation. In this sense, environmental enrichment (ENR) protects against acute stress and increases neuroplasticity. It has been suggested that the use of patterned auditory stimuli (PAS) may be beneficial in increasing the effectiveness of ENR on disorders related to stress, such as depression and anxiety. Examples of PAS are classical music compositions that have interesting effects at both clinical and preclinical levels. Thus, we analyzed the effects of the exposure to PAS, represented in this study by Mozart's compositions, during ENR housing for 35 days in adult male Balb/C mice to evaluate depression-associated behavior using the forced-swim test (FST) paradigm with an additional short exposure to PAS. We found that the ENR mice that were exposed to PAS during both housing and behavioral task (ENR + PAS/FST + PAS) show decreased immobility and the number of despair episodes within a higher latency to show the first bout of immobility. Additionally, we found increased neuronal activation evaluated by the identification of activity-regulated cytoskeleton-associated protein- (Arc-) labeled cells in the prefrontal cortex (PFC) in mice exposed to PAS during housing and in the absence or presence of PAS during FST. Moreover, we found increased neuronal activation in the auditory cortex (AuCx) of mice exposed to PAS during FST. Our study suggests that the exposure to PAS during an emotional challenge decreases despair-like behavior in rodents that were previously housed in an enriched environment in combination with auditory stimuli. Thus, our data indicate that the role of the exposure to PAS as an intervention or in combination with positive environment to aid in treating neuropsychiatric disorders is worth pursuing.

Exfoliated Human Olfactory Neuroepithelium: A Source of Neural Progenitor Cells.

Neural progenitor cells (NPC) contained in the human adult olfactory neuroepithelium (ONE) possess an undifferentiated state, the capability of self-renewal, the ability to generate neural and glial cells as well as being kept as neurospheres in cell culture conditions. Recently, NPC have been isolated from human or animal models using high-risk surgical methods. Therefore, it was necessary to improve methodologies to obtain and maintain human NPC as well as to achieve better knowledge of brain disorders. In this study, we propose the establishment and characterization of NPC cultures derived from the human olfactory neuroepithelium, using non-invasive procedures. Twenty-two healthy individuals (29.7 ± 4.5 years of age) were subjected to nasal exfoliation. Cells were recovered and kept as neurospheres under serum-free conditions. The neural progenitor origin of these neurospheres was determined by immunocytochemistry and qPCR. Their ability for self-renewal and multipotency was analyzed by clonogenic and differentiation assays, respectively. In the cultures, the ONE cells preserved the phenotype of the neurospheres. The expression levels of Nestin, Musashi, Sox2, and ßIII-tubulin demonstrated the neural origin of the neurospheres; 48% of the cells separated could generate neurospheres, determining that they retained their self-renewal capacity. Neurospheres were differentiated in the absence of growth factors (EGF and FGF), and their multipotency ability was maintained as well. We were also able to isolate and grow human neural progenitor cells (neurospheres) through nasal exfoliates (non-invasive method) of the ONE from healthy adults, which is an extremely important contribution for the study of brain disorders and for the development of new therapies.

Vascular endothelial growth factor influences migration and focal adhesions, but not proliferation or viability, of human neural stem/progenitor cells derived from olfactory epithelium.

In humans, new neurons are continuously added in the olfactory epithelium even in the adulthood. The resident neural stem/progenitor cells (hNS/PCs-OE) in the olfactory epithelium are influenced by several growth factors and neurotrophins. Among these modulators the vascular endothelial growth factor (VEGF) has attracted attention due its implicated in cell proliferation, survival and migration of other type of neural/stem progenitor cells. Interestingly, VEGFr2 receptor expression in olfactory epithelium has been described in amphibians but not in humans. Here we show that VEGFr is expressed in the hNS/PCs-OE. We also investigated the effect of VEGF on the hNS/PCs-OE proliferation, viability and migration in vitro. Additionally, pharmacological approaches showed that VEGF (0.5 ng/ml)-stimulated migration of hNS/PCs-OE was blocked with the compound DMH4, which prevents the activation of VEGFr2. Similar effects were found with the inhibitors for Rac (EHT1864) and p38MAPK (SB203850) proteins, respectively. These observations occurred with changes in focal adhesion contacts. However, no effects of VEGF on proliferation or viability were found in hNS/PCs-OE. Our results suggest that hNS/PCs-OE respond to VEGF involving VEGFr2, Rac and p38MAPK.

Human neural stem/progenitor cells derived from the olfactory epithelium express the TrkB receptor and migrate in response to BDNF.

Neurogenesis constitutively occurs in the olfactory epithelium of mammals, including humans. The fact that new neurons in the adult olfactory epithelium derive from resident neural stem/progenitor cells suggests a potential use for these cells in studies of neural diseases, as well as in neuronal cell replacement therapies. In this regard, some studies have proposed that the human olfactory epithelium is a source of neural stem/progenitor cells for autologous transplantation. Although these potential applications are interesting, it is important to understand the cell biology and/or whether human neural stem/progenitor cells in the olfactory epithelium sense external signals, such as brain-derived neurotrophic factor (BDNF), that is also found in other pro-neurogenic microenvironments. BDNF plays a key role in several biological processes, including cell migration. Thus, we characterized human neural stem/progenitor cells derived from the olfactory epithelium (hNS/PCs-OE) and studied their in vitro migratory response to BDNF. In the present study, we determined that hNS/PCs-OE express the protein markers Nestin, Sox2, Ki67 and ßIII-tubulin. Moreover, the doubling time of hNS/PCs-OE was approximately 38h. Additionally, we found that hNS/PCs-OE express the BDNF receptor TrkB, and pharmacological approaches showed that the BDNF-induced (40ng/ml) migration of differentiated hNS/PCs-OE was affected by the compound K252a, which prevents TrkB activation. This observation was accompanied by changes in the number of vinculin adhesion contacts. Our results suggest that hNS/PCs-OE exhibit a migratory response to BDNF, accompanied by the turnover of adhesion contacts.

Brain-Derived Neurotrophic Factor Induces Cell Survival and the Migration of Murine Adult Hippocampal Precursor Cells During Differentiation In Vitro.

The generation of new neurons during adulthood involves local precursor cell migration and terminal differentiation in the dentate gyrus. These events are influenced by the hippocampal microenvironment. Brain-derived neurotrophic factor (BDNF) is relevant for hippocampal neuronal development and behavior. Interestingly, studies that have been performed in controlled in vitro systems that involve isolated precursor cells that were derived from the dentate gyrus (AHPCs) have shown that BDNF induces the activation of the TrkB receptor and, consequentially, might activate signaling pathways that favor survival and neuronal differentiation. Based on the fact that the cellular events of AHPCs that are induced by single factors can be studied in this controlled in vitro system, we investigated the ability of BDNF and the involvement of protein kinase C (PKC), as one of the TrkB-downstream activated signaling proteins, in the regulation of migration, here reflected by motility, of AHPCs. Precursor cells were cultured following a concentration-response curve (1-640 ng/ml) for 24 or 96 h. We found that BDNF favored cell survival without altering the viability under culture proliferative conditions of the AHPCs. Concomitantly, glial- and neuronal-differentiated precursor cells increased as a consequence of survival promoted by BDNF. Additionally, pharmacological approaches showed that BDNF (40 ng/ml)-induced migration of AHPCs was blocked with the compounds K252a and GF109203x, which prevent the activation of TrkB and PKC, respectively. The results indicate that in the in vitro migration of differentiated AHPCs it is involved the BDNF and TrkB cascade. Our results provide additional information about the mechanism by which BDNF impacts adult neurogenesis in the hippocampus.

Acute deep brain stimulation in the thalamic reticular nucleus protects against acute stress and modulates initial events of adult hippocampal neurogenesis.

Deep brain stimulation (DBS) is used as an alternative therapeutic procedure for pharmacoresistant psychiatric disorders. Recently the thalamic reticular nucleus (TRN) gained attention due to the description of a novel pathway from the amygdala to this nucleus suggesting that may be differentially disrupted in mood disorders. The limbic system is implicated in the regulation of these disorders that are accompanied by neuroplastic changes. The hippocampus is highly plastic and shows the generation of new neurons, process affected by stress but positively regulated by antidepressant drugs. We explored the impact of applying acute DBS to the TRN (DBS-TRN) in male Wistar rats exposed to acute stress caused by the forced-swim Porsolt's test (FST) and on initial events of hippocampal neurogenesis. After the first session of forced-swim, rats were randomly subdivided in a DBS-TRN and a Sham group. Stimulated rats received 10min of DBS, thus the depressant-like behavior reflected as immobility was evaluated in the second session of forced-swim. Locomotricity was evaluated in the open field test. Cell proliferation and doublecortin-associated cells were quantified in the hippocampus of other cohorts of rats. No effects of electrode implantation were found in locomotricity. Acute DBS-TRN reduced immobility in comparison to the Sham group (p<0.001). DBS-TRN increased cell proliferation (Ki67 or BrdU-positive cells; p=0.02, p=0.02) and the number of doublecortin-cells compared to the Sham group (p<0.02). Similar effects were found in rats previously exposed to the first session of forced-swim. Our data could suggest that TRN brain region may be a promising target for DBS to treat intractable depression.

The neurogenic effects of an enriched environment and its protection against the behavioral consequences of chronic mild stress persistent after enrichment cessation in six-month-old female Balb/C mice.

Because stress may underlie the presence of depressive episodes, strategies to produce protection against or to reverse the effects of stress on neuroplasticity and behavior are relevant. Preclinical studies showed that exposure to stimuli, such as physical activity and environmental enrichment (ENR), produce beneficial effects against stress causing antidepressant-like effects in rodents. Additionally, ENR induces positive effects on neuroplasticity, neurochemistry and behavior at any age of rodents tested. Here, we analyzed whether ENR exposure prevents the development of depressive-like behavior produced by unpredictable, chronic mild stress (CMS) exposure as well as changes in hippocampal neurogenesis in a six-month-old female Balb/C mice, strain that shows low baseline levels of hippocampal neurogenesis. Mice were assigned to one of four groups: (1) normal housing-normal housing (NH-NH), (2) NH-CMS, (3) ENR-NH, or (4) ENR-CMS. The animals were exposed over 46 days to ENR or NH and subsequently to NH or CMS for 4 weeks. ENR induces long-term effects protecting against CMS induction of anhedonia and hopelessness behaviors. Independent of housing conditions, ENR increased the number of proliferative cells (Ki67), and CMS decreased the number of proliferative cells. ENR increased the newborn cells (BrdU) and mature phenotypes of neurons; these effects were not changed by CMS exposure. Similarly, the number of doublecortin-positive cells was not affected by CMS in ENR mice, which showed more cells with complex dendrite arborizations. Our study suggests that ENR induces protection against the effects of CMS on behavior and neuroplasticity in six-month-old Balb/C mice.

Chronic administration of a melatonin membrane receptor antagonist, luzindole, affects hippocampal neurogenesis without changes in hopelessness-like behavior in adult mice.

Melatonin is involved in the regulation of hippocampal neuronal development during adulthood. Emerging evidence indicates that exogenous melatonin acts during different events of the neurogenic process and exerts antidepressant-like behavior in rodents. Thus, melatonin might act through different mechanism, including acting as an antioxidant, interacting with intracellular proteins and/or activating membrane receptors. The melatonin membrane receptors (MMRs; Mt1/Mt2) are distributed throughout the hippocampus with an interesting localization in the hippocampal neurogenic microenvironment (niche), suggesting the involvement of these receptors in the beneficial effects of melatonin on hippocampal neurogenesis and behavior. In this study, we analyzed the participation of MMRs in the baseline neurogenesis in C57BL/6 mice. To this end, we used a pharmacological approach, administering luzindole (10 mg/kg) for 14 days. We observed a decrease in the absolute number of doublecortin-positive cells (49%) without changes in either the dendrite complexity of mature doublecortin-cells or the number of apoptotic cells (TUNEL). However, after the chronic administration of luzindole, cell proliferation (Ki67) significantly decreased (36%) with increasing (>100%) number of neural stem cells (NSCs; GFAP(+)/Sox2(+)) in the subgranular zone of the dentate gyrus of the hippocampus. In addition, luzindole did not affect hopelessness-like behavior in the forced swim test (FST) or changes in the novelty suppressed feeding test (NST) after 14 days of treatment either neuronal activation in the dentate gyrus after FST. These results suggest that the MMRs are involved in the effects of endogenous melatonin to mediate the transition from NSCs and proliferative cells to the following developmental stages implicated in the hippocampal neurogenic process of adult female C57BL/6 mice.

Resveratrol Enhances Neuroplastic Changes, Including Hippocampal Neurogenesis, and Memory in Balb/C Mice at Six Months of Age.

Resveratrol (RVTL) is a flavonoid found in red wine and has been publicized heavily as an anti-aging compound. Indeed, basic research confirms that although there is much hype in the promotion of RVTL, flavonoids such as RVTL have a wide range of biological effects. We here investigated the effects of RVTL treatment on hippocampal plasticity and memory performance in female Balb/C mice, a strain with low baseline levels of adult neurogenesis. Two weeks of treatment with RVTL (40 mg/kg) induced the production of new neurons in vivo by increasing cell survival and possibly precursor cell proliferation. In addition, RVTL decreased the number of apoptotic cells. The number of doublecortin (DCX)-expressing intermediate cells was increased. RVTL stimulated neuronal differentiation in vitro without effects on proliferation. In the dentate gyrus, RVTL promoted the formation and maturation of spines on granule cell dendrites. RVTL also improved performance in the step down passive avoidance test. The RVTL-treated mice showed increase in the levels of two key signaling proteins, phospho-Akt and phospho-PKC, suggesting the involvement of these signaling pathways. Our results support the vision that flavonoids such as resveratrol deserve further examination as plasticity-inducing compounds in the context of successful cognitive aging.

Melatonin maintains calcium-binding calretinin-positive neurons in the dentate gyrus during aging of Balb/C mice.

Melatonin, the main product synthesized by the pineal gland, modulates several brain functions through different mechanisms, some of them involving the activation or participation of calcium binding intracellular proteins, such as the alpha calcium dependent protein kinase C and calmodulin. Another calcium-binding protein is calretinin, which exerts an essential role for adult hippocampal neurogenesis. Melatonin favors calretinin-positive neurons in the dentate gyrus (DG) of young mice but hippocampal neurogenesis and plasma levels of melatonin decrease during aging. Thus, in this study, we analyzed the impact of exogenous supplementation with melatonin in calretinin-neurons and their distribution along the dorsal-ventral DG in the hippocampus at three different time points (1, 3, or 6 months) after daily treatment with melatonin (8 mg/kg) in male Balb/C mice. We found an increase in the number of calretinin-positive neurons in the DG after treatment (>66%). Although a significant decline in the number of calretinin-neurons was found in both treated (~60.46-69.56%) and untreated mice (~68.81-70.34%) with respect to the youngest mice analyzed, melatonin still maintained higher number of cells in the DG. Also, the distribution of calretinin-neurons along the dorsal-ventral DG significantly showed more cells in the ventral-DG of mice treated with melatonin. Together, the data suggest that melatonin also acts on calretinin in the DG, supporting it as a molecule connecting calcium signaling and neuronal development.

Melatonin synergizes with citalopram to induce antidepressant-like behavior and to promote hippocampal neurogenesis in adult mice.

Adult hippocampal neurogenesis is affected in some neuropsychiatric disorders such as depression. Numerous evidence indicates that plasma levels of melatonin are decreased in depressed patients. Also, melatonin exerts positive effects on the hippocampal neurogenic process and on depressive-like behavior. In addition, antidepressants revert alterations of hippocampal neurogenesis present in models of depression following a similar time course to the improvement of behavior. In this study, we analyzed the effects of both, citalopram, a widely used antidepressant, and melatonin in the Porsolt forced swim test. In addition, we investigated the potential antidepressant role of the combination of melatonin and citalopram (MLTCITAL), its type of pharmacological interaction on depressive behavior, and its effect on hippocampal neurogenesis. Here, we found decreased immobility behavior in mice treated with melatonin (<14-33%) and citalopram (<17-30%). Additionally, the MLTCITAL combination also decreased immobility (<22-35%) in comparison with control mice, reflecting an antidepressant-like effect after 14 days of treatment. Moreover, MLTCITAL decreased plasma corticosterone levels (≤13%) and increased cell proliferation (>29%), survival (>39%), and the absolute number of -associated new neurons (>53%) in the dentate gyrus of the hippocampus. These results indicate that the MLTCITAL combination exerts synergism to induce an antidepressant-like action that could be related to the modulation of adult hippocampal neurogenesis. This outcome opens the opportunity of using melatonin to promote behavioral benefits and hippocampal neurogenesis in depression and also supports the use of the MLTCITAL combination as an alternative to treat depression.