Brain-resident Cells Research Articles

0124 Brain Resident Mast Cells Regulate Brain Histamine Content and Promote Wakefulness

0124 Brain Resident Mast Cells Regulate Brain Histamine Content and Promote Wakefulness

Glial Cell Expression of PD-L1.

The programmed death (PD)-1/PD-L1 pathway is a well-recognized negative immune checkpoint that results in functional inhibition of T-cells. Microglia, the brain-resident immune cells are vital for pathogen detection and initiation of neuroimmune responses. Moreover, microglial cells and astrocytes govern the activity of brain-infiltrating antiviral T-cells through upregulation of PD-L1 expression. While T-cell suppressive responses within brain are undoubtedly beneficial to the host, preventing cytotoxic damage to this vital organ, establishment of a prolonged anti-inflammatory milieu may simultaneously lead to deficiencies in viral clearance. An immune checkpoint blockade targeting the PD-1: PD-L1 (B7-H1; CD274) axis has revolutionized contemporary treatment for a variety of cancers. However, the therapeutic potential of PD1: PD-L1 blockade therapies targeting viral brain reservoirs remains to be determined. For these reasons, it is key to understand both the detrimental and protective functions of this signaling pathway within the brain. This review highlights how glial cells use PD-L1 expression to modulate T-cell effector function and limit detrimental bystander damage, while still retaining an effective defense of the brain.

Prenatal Allergen Exposure Perturbs Sexual Differentiation and Programs Lifelong Changes in Adult Social and Sexual Behavior

Sexual differentiation is the early life process by which the brain is prepared for male or female typical behaviors, and is directed by sex chromosomes, hormones and early life experiences. We have recently found that innate immune cells residing in the brain, including microglia and mast cells, are more numerous in the male than female rat brain. Neuroimmune cells are also key participants in the sexual differentiation process, specifically organizing the synaptic development of the preoptic area and leading to male-typical sexual behavior in adulthood. Mast cells are known for their roles in allergic responses, thus in this study we sought to determine if exposure to an allergic response of the pregnant female in utero would alter the sexual differentiation of the preoptic area of offspring and resulting sociosexual behavior in later life. Pregnant rats were sensitized to ovalbumin (OVA), bred, and challenged intranasally with OVA on gestational day 15, which produced robust allergic inflammation, as measured by elevated immunoglobulin E. Offspring of these challenged mother rats were assessed relative to control rats in the early neonatal period for mast cell and microglia activation within their brains, downstream dendritic spine patterning on POA neurons, or grown to adulthood to assess behavior and dendritic spines. In utero exposure to allergic inflammation increased mast cell and microglia activation in the neonatal brain, and led to masculinization of dendritic spine density in the female POA. In adulthood, OVA-exposed females showed an increase in male-typical mounting behavior relative to control females. In contrast, OVA-exposed males showed evidence of dysmasculinization, including reduced microglia activation, reduced neonatal dendritic spine density, decreased male-typical copulatory behavior, and decreased olfactory preference for female-typical cues. Together these studies show that early life allergic events may contribute to natural variations in both male and female sexual behavior, potentially via underlying effects on brain-resident mast cells.

The immune suppressive microenvironment of human gliomas depends on the accumulation of bone marrow-derived macrophages in the center of the lesion

BackgroundSystemic and local immune suppression plays a significant role in glioma progression. Glioma microenvironment contains both brain-resident microglial cells (MG) and bone marrow-derived macrophages (BMDM), but the study of their functional and immune regulatory activity has been hampered until now by the lack of markers allowing a proper identification and isolation to collect pure populations.MethodsMyeloid and lymphoid infiltrate were characterized in grade II, III and IV gliomas by multicolor flow cytometry, along with the composition of the cell subsets of circulating myeloid cells. Macrophages were sorted and tested for their immunosuppressive ability. Moreover, following preoperative administration of 5-aminolevulinic acid to patients, distinct areas of tumor lesion were surgically removed and analyzed, based on protoporphyrin IX fluorescence emission.ResultsThe immune microenvironment of grade II to grade IV gliomas contains a large proportion of myeloid cells and a small proportion of lymphocytes expressing markers of dysfunctional activity. BMDM and resident MG cells were characterized through a combination of markers, thus permitting their geographical identification in the lesions, their sorting and subsequent analysis of the functional characteristics. The infiltration by BMDM reached the highest percentages in grade IV gliomas, and it increased from the periphery to the center of the lesion, where it exerted a strong immunosuppression that was, instead, absent in the marginal area. By contrast, MG showed little or no suppression. Functional differences, such as iron metabolism and phagocytosis, characterized resident versus blood-derived macrophages. Significant alterations in circulating monocytes were present in grade IV patients, correlating with accumulation of tumor macrophages.ConclusionsGrade IV gliomas have an alteration in both circulating and tumor-associated myeloid cells and, differently from grade II and III gliomas, show a significant presence of blood-derived, immune suppressive macrophages. BMDM and MG have different functional properties.

Advances and Challenges in Understanding Cerebral Toxoplasmosis.

Toxoplasma gondii is a widespread parasitic pathogen that infects over one third of the global human population. The parasite invades and chronically persists in the central nervous system (CNS) of the infected host. Parasite spread and persistence is intimately linked to an ensuing immune response, which does not only limit parasite-induced damage but also may facilitate dissemination and induce parasite-associated immunopathology. Here, we discuss various aspects of toxoplasmosis where knowledge is scarce or controversial and, the recent advances in the understanding of the delicate interplay of T. gondii with the immune system in experimental and clinical settings. This includes mechanisms for parasite passage from the circulation into the brain parenchyma across the blood-brain barrier during primary acute infection. Later, as chronic latent infection sets in with control of the parasite in the brain parenchyma, the roles of the inflammatory response and of immune cell responses in this phase of the disease are discussed. Additionally, the function of brain resident cell populations is delineated, i.e., how neurons, astrocytes and microglia serve both as target cells for the parasite but also actively contribute to the immune response. As the infection can reactivate in the CNS of immune-compromised individuals, we bring up the immunopathogenesis of reactivated toxoplasmosis, including the special case of congenital CNS manifestations. The relevance, advantages and limitations of rodent infection models for the understanding of human cerebral toxoplasmosis are discussed. Finally, this review pinpoints questions that may represent challenges to experimental and clinical science with respect to improved diagnostics, pharmacological treatments and immunotherapies.

Glial TLR2-driven innate immune responses and CD8+ T cell activation against brain tumor.

Growing interest has been focused on the roles of microglia as sentinels and effector cells that guard diverse pathological milieu in the brain. Here, it has been reported that microglial TLR2 is a crucial molecule that confers innate and adaptive immunity against brain tumor. TLR2 is preferentially expressed on microglia, brain-resident immune cells, in the tumor-bearing cerebral hemisphere of mouse and rat intracranial tumor models. Microglial TLR2 rapidly responds to brain tumor and modulates the inflammation-associated immune responses including phagocytosis, which are markedly decreased in TLR2-deficient mice. We further reveal that TLR2, but not TLR4, is essential for the tumor-triggered increase of MHC I in microglia. in vitro co-culture and in vivo experiments show that the glial TLR2-MHC I axis contributes to the proliferation and activation of CD8+ T cells by brain tumor. In addition, brain tumor-bearing β2m-/- , Tlr2-/- , or Rag2 -/- γc -/- mice exhibit higher tumor volumes compared with WT mice with tumor. Survival analysis of GL26-bearing MHC I-defective mice also support the contribution of glial TLR2-MHC I axis to brain tumor immunity. Moreover, using publicly available data sets of human brain tumor patients, we find that glioblastoma (GBM) tissues with high TLR2 level have similar co-occurrence patterns with MHC I molecules, and the amounts and activity of infiltrating CD8+ T cells are correlated with TLR2 level in tissues from GBM patients. Collectively, our findings provide the importance of glial TLR2-driven innate and adaptive immune responses in the brain tumor microenvironment.

Leukemia-derived exosomes and cytokines pave the way for entry into the brain.

Infiltration of acute lymphoblastic leukemia (ALL) blasts into the CNS remains as a major clinical problem, with high risk for chemotherapy-resistant relapse and treatment-related morbidity. Despite the common inclusion of CNS prophylaxis treatments in therapy regimens, there are significant gaps in understanding the mechanisms that mediate leukemia cell entry into the CNS as well as roles for resident cells in the brain. In this study, we employ a xenograft model of human B cell precursor (BCP)-ALL in immunocompromised mice. This model system recapitulates key pathological characteristics of leptomeningeal involvement seen in patients and provides insights into rare cases that involve parenchymal invasion. We examine the infiltration of engrafted leukemia blasts into brains of recipient mice and provide evidence that the interaction between blasts and brain resident cells causes aberrant activation of host cells in the brain microenvironment. BCP-ALL blasts also release multiple cytokines and exosomes containing IL-15 that bind and are internalized by astrocytes and brain vessel endothelial cells. Leukemic invasion is linked to production of VEGF-AA by astrocytes and disruption of the blood-brain-barrier (BBB) integrity. Knockdown of either IL-15 or IL-15Rαin the NALM6 cell line decreases CNS infiltration in engrafted mice. These results provide important insights into the multiple mechanisms by which lymphoblasts modulate the brain microenvironment to breach the BBB for metastatic invasion.

Healthy Brain Aging Modifies Microglial Calcium Signaling In Vivo.

Brain aging is characterized by a chronic, low-grade inflammatory state, promoting deficits in cognition and the development of age-related neurodegenerative diseases. Malfunction of microglia, the brain-resident immune cells, was suggested to play a critical role in neuroinflammation, but the mechanisms underlying this malfunctional phenotype remain unclear. Specifically, the age-related changes in microglial Ca2+ signaling, known to be linked to its executive functions, are not well understood. Here, using in vivo two-photon imaging, we characterize intracellular Ca2+ signaling and process extension of cortical microglia in young adult (2–4-month-old), middle-aged (9–11-month-old), and old (18–21-month-old) mice. Our data revealed a complex and nonlinear dependency of the properties of intracellular Ca2+ signals on an animal’s age. While the fraction of cells displaying spontaneous Ca2+ transients progressively increased with age, the frequencies and durations of the spontaneous Ca2+ transients followed a bell-shaped relationship, with the most frequent and largest Ca2+ transients seen in middle-aged mice. Moreover, in old mice microglial processes extending toward an ATP source moved faster but in a more disorganized manner, compared to young adult mice. Altogether, these findings identify two distinct phenotypes of aging microglia: a reactive phenotype, abundantly present in middle-aged animals, and a dysfunctional/senescent phenotype ubiquitous in old mice.

Fumaric Acids Directly Influence Gene Expression of Neuroprotective Factors in Rodent Microglia.

Dimethylfumarate (DMF) has been approved the for treatment of relapsing-remitting multiple sclerosis. The mode of action of DMF and its assumed active primary metabolite monomethylfumarate (MMF) is still not fully understood, notably for brain resident cells. Therefore we investigated potential direct effects of DMF and MMF on microglia and indirect effects on oligodendrocytes. Primary rat microglia were differentiated into M1-like, M2-like and M0 phenotypes and treated in vitro with DMF or MMF. The gene expression of pro-inflammatory and anti-inflammatory factors such as growth factors (IGF-1), interleukins (IL-10, IL-1β), chemokines (CCl3, CXCL-10) as well as cytokines (TGF-1β, TNFα), iNOS, and the mannose receptor (MRC1) was examined by determining their transcription level with qPCR, and on the protein level by ELISA and FACS analysis. Furthermore, microglia function was determined by phagocytosis assays and indirect effects on oligodendroglial proliferation and differentiation. DMF treatment of M0 and M1-like polarized microglia demonstrated an upregulation of gene expression for IGF-1 and MRC1, but not on the protein level. While the phagocytic activity remained unchanged, DMF and MMF treated microglia supernatants led to an enhanced proliferation of oligodendrocyte precursor cells (OPC). These results suggest that DMF has anti-inflammatory effects on microglia which may result in enhanced proliferation of OPC.

Conventional DCs sample and present myelin antigens in the healthy CNS and allow parenchymal T cell entry to initiate neuroinflammation.

The central nervous system (CNS) is under close surveillance by immune cells, which mediate tissue homeostasis, protection, and repair. Conversely, in neuroinflammation, dysregulated leukocyte invasion into the CNS leads to immunopathology and neurological disability. To invade the brain parenchyma, autoimmune encephalitogenic T helper (TH) cells must encounter their cognate antigens (Ags) presented via local Ag-presenting cells (APCs). The precise identity of the APC that samples, processes, and presents CNS-derived Ags to autoaggressive T cells is unknown. Here, we used a combination of high-dimensional single-cell mapping and conditional MHC class II ablation across all CNS APCs to systematically interrogate each population for its ability to reactivate encephalitogenic TH cells in vivo. We found a population of conventional dendritic cells, but not border-associated macrophages or microglia, to be essential for licensing T cells to initiate neuroinflammation.

From Systemic Inflammation to Neuroinflammation: The Case of Neurolupus.

It took decades to arrive at the general consensus dismissing the notion that the immune system is independent of the central nervous system. In the case of uncontrolled systemic inflammation, the relationship between the two systems is thrown off balance and results in cognitive and emotional impairment. It is specifically true for autoimmune pathologies where the central nervous system is affected as a result of systemic inflammation. Along with boosting circulating cytokine levels, systemic inflammation can lead to aberrant brain-resident immune cell activation, leakage of the blood–brain barrier, and the production of circulating antibodies that cross-react with brain antigens. One of the most disabling autoimmune pathologies known to have an effect on the central nervous system secondary to the systemic disease is systemic lupus erythematosus. Its neuropsychiatric expression has been extensively studied in lupus-like disease murine models that develop an autoimmunity-associated behavioral syndrome. These models are very useful for studying how the peripheral immune system and systemic inflammation can influence brain functions. In this review, we summarize the experimental data reported on murine models developing autoimmune diseases and systemic inflammation, and we explore the underlying mechanisms explaining how systemic inflammation can result in behavioral deficits, with a special focus on in vivo neuroimaging techniques.

The Multifarious Role of Microglia in Brain Metastasis.

The immune landscape in brain metastasis is a very heterogeneous framework. Amongst a broad plethora of cells within the tumor microenvironment, the presence of activated microglia has been perfectly described. The innate role of microglial cells is to detect and eliminate any insults that may disturb the regular behavior of the brain. As part of its defensive role, it releases pro- and anti-inflammatory cytokines that aim to modulate the inflammatory scenario at the metastatic foci. However, the long term effects that these cells may exert on the metastatic progression is not clear. One of the biggest challenges in the field is to distinguish between brain resident microglial cells and infiltrated bone-marrow derived macrophages. Part of this issue is the fact that both cell types share similar phenotypes. Current studies are based on the modulation of the immune response against cancer cells (immunotherapy). However, most of current clinical trials and newly developed drugs focus on the adaptive immune response (e.g., immune blockade check-points). Additionally, the unique structure of the central nervous system with the presence of the blood-brain barrier have hindered a significant advance in novel therapies against brain metastasis. In this manuscript, we describe current advances in characterization of tumor-associated microglia and macrophages, the importance of microglia during the anti-cancerous response, and the future direction for the development of new strategies against this complex disease.

The metastatic microenvironment: Melanoma-microglia cross-talk promotes the malignant phenotype of melanoma cells.

Melanoma has the highest propensity to metastasize to the brain compared to other cancers, as brain metastases are found frequently high in patients who have prolonged survival with visceral metastasis. Once disseminated in the brain, melanoma cells communicate with brain resident cells that include astrocytes and microglia. Microglia cells are the resident macrophages of the brain and are the main immunological cells in the CNS involved in neuroinflammation. Data on the interactions between brain metastatic melanoma cells and microglia and on the role of microglia-mediated neuroinflammation in facilitating melanoma brain metastasis are lacking. To elucidate the role of microglia in melanoma brain metastasis progression, we examined the bidirectional interactions between microglia and melanoma cells in the tumor microenvironment. We identified the molecular and functional modifications occurring in brain-metastasizing melanoma cells and microglia cells after the treatment of each cell type with supernatants of the counter cell type. Both cells induced alteration in gene expression programs, cell signaling, and cytokine secretion in the counter cell type. Moreover, melanoma cells exerted significant morphological changes on microglia cells, enhanced proliferation, induced matrix metalloproteinase-2 (MMP-2) activation, and cell migration. Microglia cells induced phenotypic changes in melanoma cells increasing their malignant phenotype: increased melanoma proliferation, MMP-2 activity, cell migration, brain endothelial penetration, and tumor cells ability to grow as spheroids in 3D cultures. Our work provides a novel insight into the bidirectional interactions between melanoma and micoglia cells, suggesting the contribution of microglia to melanoma brain metastasis formation.

Outside in: Unraveling the Role of Neuroinflammation in the Progression of Parkinson's Disease.

Neuroinflammation is one of the most important processes involved in the pathogenesis of Parkinson's disease (PD). The current concept of neuroinflammation comprises an inflammation process, which occurs in the central nervous system due to molecules released from brain-resident and/or blood-derived immune cells. Furthermore, the evidence of the contribution of systemic delivered molecules to the disease pathogenesis, such as the gut microbiota composition, has been increasing during the last years. Under physiological conditions, microglia and astrocytes support the well-being and well-function of the brain through diverse functions, including neurotrophic factor secretion in both intact and injured brain. On the other hand, genes that cause PD are expressed in astrocytes and microglia, shifting their neuroprotective role to a pathogenic one, contributing to disease onset and progression. In addition, growth factors are a subset of molecules that promote cellular survival, differentiation and maturation, which are critical signaling factors promoting the communication between cells, including neurons and blood-derived immune cells. We summarize the potential targeting of astrocytes and microglia and the systemic contribution of the gut microbiota in neuroinflammation process archived in PD.

Regulated Necrosis Orchestrates Microglial Cell Death in Manganese-Induced Toxicity

Microglia, the brain resident immune cells, play prominent roles in immune surveillance, tissue repair and neural regeneration. Despite these pro-survival actions, the relevance of these cells in the progression of several neuropathologies has been established. In the context of manganese (Mn) overexposure, it has been proposed that microglial activation contributes to enhance the neurotoxicity. However, the occurrence of a direct cytotoxic effect of Mn on microglial cells remains controversial. In the present work, we investigated the potential vulnerability of immortalized mouse microglial cells (BV-2) toward Mn2+, focusing on the signaling pathways involved in cell death. Evidence obtained showed that Mn2+ induces a decrease in cell viability which is associated with reactive oxygen species (ROS) generation. In this report we demonstrated, for the first time, that Mn2+ triggers regulated necrosis (RN) in BV-2 cells involving two central mechanisms: parthanatos and lysosomal disruption. The occurrence of parthanatos is supported by several cellular and molecular events: (i) DNA damage; (ii) AIF translocation from mitochondria to the nucleus; (iii) mitochondrial membrane permeabilization; and (iv) PARP1-dependent cell death. On the other hand, Mn2+ induces lysosomal membrane permeabilization (LMP) and cathepsin D (CatD) release into the cytosol supporting the lysosomal disruption. Pre-incubation with CatB and D inhibitors partially prevented the Mn2+-induced cell viability decrease. Altogether these events point to lysosomes as players in the execution of RN. In summary, our results suggest that microglial cells could be direct targets of Mn2+ damage. In this scenario, Mn2+ triggers cell death involving RN pathways.

Microglia prevent peripheral immune cell invasion and promote an anti-inflammatory environment in the brain of APP-PS1 transgenic mice

BackgroundUndoubtedly, neuroinflammation is a major contributor to Alzheimer’s disease (AD) progression. Neuroinflammation is characterized by the activity of brain resident glial cells, in particular microglia, but also by peripheral immune cells, which infiltrate the brain at certain stages of disease progression. The specific role of microglia in shaping AD pathology is still controversially discussed. Moreover, a possible role of microglia in the interaction and recruitment of peripheral immune cells has so far been completely ignored.MethodsWe ablated microglia cells in 12-month-old WT and APP-PS1 transgenic mice for 4 weeks using the CSF1R inhibitor PLX5622 and analyzed its consequences to AD pathology and in particular to peripheral immune cell infiltration.ResultsPLX5622 treatment successfully reduced microglia numbers. Interestingly, it uncovered a treatment-resistant macrophage population (Iba1+/TMEM119−). These cells strongly expressed the phagocytosis marker CD68 and the lymphocyte activation, homing, and adhesion molecule CD44, specifically at sites of amyloid-beta plaques in the brains of APP-PS1 mice. In consequence, ablation of microglia significantly raised the number of CD3+/CD8+ T-cells and reduced the expression of anti-inflammatory genes in the brains of APP-PS1 mice.ConclusionWe conclude that in neurodegenerative conditions, chronically activated microglia might limit CD3+/CD8+ T-cell recruitment to the brain and that local macrophages connect innate with adaptive immune responses. Investigating the role of peripheral immune cells, their interaction with microglia, and understanding the link between innate and adaptive immune responses in the brain might be a future directive in treating AD pathology.

Robust Control of a Neurotropic Parasite Through MHC I Presentation by Infected Neurons

Inadequate immune control of Central Nervous System (CNS) pathogens may result in fatal neuroinflammation. CD8 T cells are instrumental for CNS pathogen control but the functions of brain-resident antigen-presenting cells remain poorly understood. Here, we analyzed the modalities of MHC I presentation of Toxoplasma gondii, a parasite chronically residing in the CNS. Using transgenic parasites with stage-restricted expression of a protective antigen, we show that MHC I presentation of the rapidly dividing tachyzoites prevents encephalitis and that improper parasite control during encephalitis correlates with sub-optimal MHC I presentation of T. gondii by neurons. Thanks to conditional MHC I-deficient mice, we demonstrate that neuronal MHC I presentation is dispensable for brain CD8 T cell accumulation but is required for durable pathogen control. Thus, MHC I presentation of parasite antigens by CNS neurons is a pivotal checkpoint that determines the clinical outcome of a prevalent and chronic brain infection.

Microglial translational profiling reveals a convergent APOE pathway from aging, amyloid, and tau.

Alzheimer's disease (AD) is an age-associated neurodegenerative disease characterized by amyloidosis, tauopathy, and activation of microglia, the brain resident innate immune cells. We show that a RiboTag translational profiling approach can bypass biases due to cellular enrichment/cell sorting. Using this approach in models of amyloidosis, tauopathy, and aging, we revealed a common set of alterations and identified a central APOE-driven network that converged on CCL3 and CCL4 across all conditions. Notably, aged females demonstrated a significant exacerbation of many of these shared transcripts in this APOE network, revealing a potential mechanism for increased AD susceptibility in females. This study has broad implications for microglial transcriptomic approaches and provides new insights into microglial pathways associated with different pathological aspects of aging and AD.

Pharmacologic Depletion of Microglia Increases Viral Load in the Brain and Enhances Mortality in Murine Models of Flavivirus-Induced Encephalitis.

Flaviviruses account for most arthropod-borne cases of human encephalitis in the world. However, the exact mechanisms of injury to the central nervous system (CNS) during flavivirus infections remain poorly understood. Microglia are the resident immune cells of the CNS and are important for multiple functions, including control of viral pathogenesis. Utilizing a pharmacologic method of microglia depletion (PLX5622 [Plexxikon Inc.], an inhibitor of colony-stimulating factor 1 receptor), we sought to determine the role of microglia in flaviviral pathogenesis. Depletion of microglia resulted in increased mortality and viral titer in the brain following infection with either West Nile virus (WNV) or Japanese encephalitis virus (JEV). Interestingly, microglial depletion did not prevent virus-induced increases in the expression of relevant cytokines and chemokines at the mRNA level. In fact, the expression of several proinflammatory genes was increased in virus-infected, microglia-depleted mice compared to virus-infected, untreated controls. In contrast, and as expected, expression of the macrophage marker triggering receptor expressed on myeloid cells 2 (TREM2) was decreased in virus-infected, PLX5622-treated mice compared to virus-infected controls.IMPORTANCE As CNS invasion by flaviviruses is a rare but life-threatening event, it is critical to understand how brain-resident immune cells elicit protection or injury during disease progression. Microglia have been shown to be important in viral clearance but may also contribute to CNS injury as part of the neuroinflammatory process. By utilizing a microglial depletion model, we can begin to parse out the exact roles of microglia during flaviviral pathogenesis with hopes of understanding specific mechanisms as potential targets for therapeutics.

Early Maternal Deprivation Induces Microglial Activation, Alters Glial Fibrillary Acidic Protein Immunoreactivity and Indoleamine 2,3-Dioxygenase during the Development of Offspring Rats.

Maternal deprivation (MD) induces behavioral changes and impacts brain circuits that could be associated with the pathophysiology of depression. This study investigated the markers of microglia and astrocyte activation as well as indoleamine 2,3-dioxygenase (IDO) expression in developmental programming after early life MD (on postnatal days (PNDs) 20, 30, 40, and 60). On PND 60, the rats that were subjected to MD displayed depressive-like behavior. On PND 10, it was found that there was a decrease in the level of glial fibrillary acidic protein (GFAP) immunopositive cells, a decrease in the level of IDO expression, and an increase in the level of Iba-1 (microglial marker) in the hippocampus of rats that were subjected to MD. On PND 20, levels of GFAP were also found to have decreased in the hippocampus, and there was an increase in the level of Iba-1 in the hippocampus. AIF-1 (microglial marker) expression was observed in the PFC following MD. On PND 30, the levels of Iba-1 remained elevated. On PND 40, the levels of GFAP were found to have increased in the hippocampus of rats that were subjected to MD. On PND 60, the levels of GFAP and AIF-1 remained elevated following MD. These results suggest that early life stress induces negative developmental programming in rats, as demonstrated by depressive-like behavior in adult life. Moreover, MD increases microglial activation in both early and late developmental phases. The levels of GFAP and IDO decreased in the early stages but were found to be higher in later developmental periods. These findings suggest that MD could differentially affect the expression of the IDO enzyme, astrocytes, and microglial activation depending on the neurodevelopmental period. The onset of an inflammatory state from resident brain cells could be associated with the activation of the kynurenine pathway and the development of depressive behavior in adulthood.