Neonatal Immune Activation Research Articles

Age and sex-dependent gut alterations in mice induced by neonatal immune activation with lipopolysaccharide

Neonatal immune activation (NIA) through exposure to lipopolysaccharide (LPS) induces adult behavioral changes in rodents that resemble symptoms of developmental disorders, such as autism spectrum disorder. The neonatal timing of LPS exposure appears to play a crucial role in determining the nature and extent of long-term changes. This study aims to explore whether a 3-day LPS-NIA triggers sex- and age-related changes in gut function, potentially linking LPS-NIA to gastrointestinal dysfunction. Male and female Swiss mice received intraperitoneal injections of LPS or saline on postnatal days (PN) 3, 5, and 7. At PN35 (juvenile) and PN70 (adult), gut inflammation and oxidative stress were evaluated in addition to assessments of working memory, depressive-like symptoms, sociability, and repetitive behavior. Gut examination showed elevated C-X-C motif chemokine receptor 3 (CXCR3) in LPS-NIA mice, while MyD88 and Zonulin expressions were significantly higher only in adult LPS-NIA females. Interleukin (IL)-23 expression increased in juvenile and adult male and juvenile female LPS-NIA mice. Oxidative changes included decreased duodenal reduced glutathione (GSH) in juvenile females and ileal GSH in adult females exposed to LPS-NIA. Regarding behavioral alterations, adult LPS-NIA females exhibited depressive-like behavior. Working memory deficits were observed across all LPS-NIA groups. Only juvenile LPS-NIA females increased grooming, while rearing was higher in adult LPS-NIA mice of both sexes. The findings imply that LPS-NIA impacts intestinal barrier function and causes gut inflammatory alterations that are sex- and age-specific. These findings pave the way for exploring potential mechanisms that could contribute to LPS-induced gastrointestinal disturbances among individuals with ASD.

Effects of different types of induced neonatal inflammation on development and behavior of C57BL/6 and BTBR mice

Neuroinflammation in the early postnatal period can disturb trajectories of the completion of normal brain development and can lead to mental illnesses, such as depression, anxiety disorders, and personality disorders later in life. In our study, we focused on evaluating short- and long-term effects of neonatal inflammation induced by lipopolysaccharide, poly(I:C), or their combination in female and male C57BL/6 and BTBR mice. We chose the BTBR strain as potentially more susceptible to neonatal inflammation because these mice have behavioral, neuroanatomical, and physiological features of autism spectrum disorders, an abnormal immune response, and several structural aberrations in the brain. Our results indicated that BTBR mice are more sensitive to the influence of the neonatal immune activation (NIA) on the formation of neonatal reflexes than C57BL/6 mice are. In these experiments, the injection of lipopolysaccharide had an effect on the formation of the cliff aversion reflex in female BTBR mice. Nonetheless, NIA had no delayed effects on either social behavior or anxiety-like behavior in juvenile and adolescent BTBR and C57BL/6 mice. Altogether, our data show that NIA has mimetic-, age-, and strain-dependent effects on the development of neonatal reflexes and on exploratory activity in BTBR and C57BL/6 mice.

Once induced, it lasts for a long time: the structural and molecular signatures associated with depressive-like behavior after neonatal immune activation.

Adverse factors such as stress or inflammation in the neonatal period can affect the development of certain brain structures and have negative delayed effects throughout the lifespan of an individual, by reducing cognitive abilities and increasing the risk of psychopathologies. One possible reason for these delayed effects is the neuroinflammation caused by neonatal immune activation (NIA). Neuroinflammation can lead to disturbances of neurotransmission and to reprogramming of astroglial and microglial brain cells; when combined, the two problems can cause changes in the cytoarchitecture of individual regions of the brain. In addition, neuroinflammation may affect the hypothalamic-pituitary-adrenal (HPA) axis and processes of oxidative stress, thereby resulting in higher stress reactivity. In our review, we tried to answer the questions of whether depressive-like behavior develops after NIA in rodents and what the molecular mechanisms associated with these disorders are. Most studies indicate that NIA does not induce depressive-like behavior in a steady state. Nonetheless, adult males (but not females or adolescents of both sexes) with experience of NIA exhibit marked depressive-like behavior when exposed to aversive conditions. Analyses of molecular changes have shown that NIA leads to an increase in the amount of activated microglia and astroglia in the frontal cortex and hippocampus, an increase in oxidative-stress parameters, a change in stress reactivity of the HPA axis, and an imbalance of cytokines in various regions of the brain, but not in blood plasma, thus confirming the local nature of the inflammation. Therefore, NIA causes depressive-like behavior in adult males under aversive testing conditions, which are accompanied by local inflammation and have sex- and age-specific effects.

Delayed effects of neonatal immune activation on brain neurochemistry and hypothalamic-pituitary-adrenal axis functioning.

During the postnatal period, the brain is highly sensitive to stress and inflammation, which are hazardous to normal growth and development. There is increasing evidence that inflammatory processes in the early postnatal period increase the risk of psychopathologies and cognitive impairment later in life. On the other hand, there are few studies on the ability of infectious agents to cause long-term neuroinflammation, leading to changes in the hypothalamic-pituitary-adrenal axis functioning and an imbalance in the neurotransmitter system. In this review, we examine short- and long-term effects of neonatal-induced inflammation in rodents on glutamatergic, GABAergic and monoaminergic systems and on hypothalamic-pituitary-adrenal axis activity.

Neonatal immune challenge influences the microbiota and behaviour in a sexually dimorphic manner

There is comorbidity between anxiety disorders and gastrointestinal disorders, with both linked to adverse early life events. The microbiome gut-brain-axis, a bidirectional communication system, is plastic throughout the neonatal period and is a possible mediator of this relationship. Here, we used a well-established neonatal rodent immune activation model to investigate the long-term effect of neonatal lipopolysaccharide (LPS) exposure on adult behaviour and the relationship to microbiome composition. Wistar rats were injected with LPS (0.05 mg/kg) or saline (equivolume) on postnatal days 3 and 5. In adulthood, behavioural tests were performed to assess anxiety-like behaviour, and microbiota sequencing was performed on stool samples. There were distinctly different behavioural phenotypes for LPS-exposed males and females. LPS-exposed males displayed typical anxiety-like behaviours with significantly decreased social interaction (F(1,22) = 7.576, p = 0.009) and increased defecation relative to saline controls (F(1,23) = 8.623, p = 0.005). LPS-exposed females displayed a different behavioural phenotype with significantly increased social interaction (F(1,22) = 6.094, p = 0.018), and exploration (F(1,24) = 6.359, p = 0.015), compared to saline controls. With respect to microbiota profiling data, Bacteroidota was significantly increased for LPS-exposed females (F(1,14) = 4.931p = 0.035) and Proteobacteria was decreased for LPS-exposed rats of both sexes versus controls (F(1,30) = 4.923p = 0.035). Furthermore, alterations in predicted functional pathways for neurotransmitters in faeces were observed with a decrease in the relative abundance of D-glutamine and D-glutamate metabolism in LPS exposed females compared to control females (p < 0.05). This suggests that neonatal immune activation alters both later life behaviour and adult gut microbiota in sex-specific ways. These findings highlight the importance of sex in determining the impact of neonatal immune activation on social behaviour and the gut microbiota.

Neonatal immune challenge induces female-specific changes in social behavior and somatostatin cell number

Decreases in social behavior are a hallmark aspect of acute “sickness behavior” in response to infection. However, immune insults that occur during the perinatal period may have long-lasting consequences for adult social behavior by impacting the developmental organization of underlying neural circuits. Microglia, the resident immune cells of the central nervous system, are sensitive to immune stimulation and play a critical role in the developmental sculpting of neural circuits, making them likely mediators of this process. Here, we investigated the impact of a postnatal day (PND) 4 lipopolysaccharide (LPS) challenge on social behavior in adult mice. Somewhat surprisingly, neonatal LPS treatment decreased sociability in adult female, but not male mice. LPS-treated females also displayed reduced social interaction and social memory in a social discrimination task as compared to saline-treated females. Somatostatin (SST) interneurons within the anterior cingulate cortex (ACC) have recently been suggested to modulate a variety of social behaviors. Interestingly, the female-specific changes in social behavior observed here were accompanied by an increase in SST interneuron number in the ACC. Finally, these changes in social behavior and SST cell number do not appear to depend on microglial inflammatory signaling, because microglia-specific genetic knock-down of myeloid differentiation response protein 88 (MyD88; the removal of which prevents LPS from increasing proinflammatory cytokines such as TNFα and IL-1β) did not prevent these LPS-induced changes. This study provides novel evidence for enduring effects of neonatal immune activation on social behavior and SST interneurons in females, largely independent of microglial inflammatory signaling.

Neonatal Immune Activation May Provoke Long-term Depressive Attributes.

Studies have shown the relationship between neuroinflammation and depressive- like parameters. However, research still has not been carried out to evaluate neuroinflammation in the neonatal period and psychiatric disorders in adulthood. To verify the association between neonatal immune activation and depressive-like parameters in adulthood using an animal model. Two days old C57BL/6 animals were exposed to lipopolysaccharides (LPS) or phosphate- buffered saline (PBS). When the animals were 46 days old, they received PBS or Imipramine at 14 days. At 60 days, the consumption of sucrose; immobility time; adrenal gland and the hippocampus weight; levels of plasma corticosterone and hippocampal Brain-derived neurotrophic factor (BDNF) were evaluated. It was observed that the animals exposed to LPS in the neonatal period and evaluated in adulthood decreased the consumption of sucrose and had reducted hippocampus weight. Also, the exposed animals presented an increase of immobility time, adrenal gland weight and plasma levels of corticosteroids. The use of imipramine did not only modify the decreased hippocampal weight. On the other hand, there were no alterations in the BDNF levels in the hippocampus with or without the use of imipramine. These results suggest that neonatal immune activation may be associated with depressive- like parameters in adulthood. It is believed that endotoxemia may trigger physiological and behavioral alterations, increasing vulnerability for the development of depression in adulthood.

Late Brain Involvement after Neonatal Immune Activation.

The neonatal immune system is still immature, which makes it more susceptible to the infectious agents. Neonatal immune activation is associated with increased permeability of the blood-brain barrier, causing an inflammatory cascade in the CNS and altering behavioral and neurochemical parameters. One of the hypotheses that has been studied is that neuroinflammation may be involved in neurodegenerative processes, such as Alzheimer's disease (AD). We evaluate visuospatial memory, cytokines levels, and the expression of tau and GSK-3β proteins in hippocampus and cortex of animals exposed to neonatal endotoxemia. C57BL/6 mice aging two days received a single injection of subcutaneous lipopolysaccharide (LPS). At 60,120, and 180 days of age, visual-spatial memory was evaluated and the hippocampus and cortex were dissected to evaluate the cytokines levels and expression of tau and GSK-3β proteins. The animals exposed to LPS in the neonatal period present with visuospatial memory impairment at 120 and 180 days of age. Here there was an increase of TNF-α and IL-1β levels in the hippocampus and cortex only at 60 days of age. Here there was an increase in the expression of GSK-3β in hippocampus of the animals at 60, 120, and 180 days of age. In the cortex, this increase occurred in the 120 and 180 days of age. Tau protein expression was high in hippocampus and cortex at 120 days of age and in hippocampus at 180 days of age. The data observed show that neonatal immune activation may be associated with visuospatial memory impairment, neuroinflammation, and increased expression of GSK-3β and Tau proteins in the long term.

Pharmacological and proteomic analyses of neonatal polyI:C-treated adult mice

Perinatal virus infection is an environmental risk factor for neurodevelopmental disorders such as schizophrenia. We previously demonstrated that neonatal treatment with a viral mimetic, polyriboinosinic-polyribocytidilic acid (polyI:C), in mice leads to emotional and cognitive deficits in adolescence. Here, we investigated the effects of antipsychotics on polyI:C-induced behavioral abnormalities. We also performed a proteomic analysis in the hippocampus of polyI:C-treated adult mice using two-dimensional electrophoresis to understand the changes in protein expression following neonatal immune activation. Neonatal mice were subcutaneously injected with polyI:C for 5 days (postnatal day 2–6). At 10 weeks, sensorimotor gating, emotional and cognitive function were analyzed in behavioral tests. Clozapine improved PPI deficit and emotional and cognitive dysfunction in polyI:C-treated mice. However, haloperidol improved only PPI deficit. Proteomic analysis revealed that two candidate proteins were obtained in the hippocampus of polyI:C-treated mice, including aldehyde dehydrogenase family 1 member L1 (ALDH1L1) and collapsin response mediator protein 5 (CRMP5). These data suggest that the neonatal polyI:C-treated mouse model may be useful for evaluating antipsychotic activity of compounds. Moreover, changes in the protein expression of ALDH1L1 and CRMP5 support our previous findings that astrocyte-neuron interaction plays a role in the pathophysiology of neurodevelopmental disorders induced by neonatal immune activation.

Neonatal immune activation by lipopolysaccharide causes inadequate emotional responses to novel situations but no changes in anxiety or cognitive behavior in Wistar rats

Infection during the prenatal or neonatal stages of life is considered one of the major risk factors for the development of mental diseases such as schizophrenia or autism. However, the impacts of such an immune challenge on adult behavior are still not clear. In our study, we used a model of early postnatal immune activation by the application of bacterial endotoxin lipopolysaccharide (LPS) to rat pups at a dose of 2 mg/kg from postnatal day (PD) 5 to PD 9. In adulthood, the rats were tested in a battery of tasks probing various aspects of behavior: spontaneous activity (open field test), social behavior (social interactions and female bedding exploration), anxiety (elevated plus maze), cognition (active place avoidance in Carousel) and emotional response (ultrasonic vocalization recording). Moreover, we tested sensitivity to acute challenge with MK-801, a psychotomimetic drug. Our results show that the application of LPS led to increased self-grooming in the female bedding exploration test and inadequate emotional reactions in Carousel maze displayed by ultrasonic vocalizations. However, it did not have serious consequences on exploration, locomotion, social behavior or cognition. Furthermore, exposition to MK-801 did not trigger social or cognitive deficits in the LPS-treated rats. We conclude that the emotional domain is the most sensitive to the changes induced by neonatal immune activation in rats, including a disrupted response to novel and stressful situations in early adulthood (similar to that observed in human patients suffering from schizophrenia or autism), while other aspects of tested behavior remain unaffected.

Neonatal immune activation and a ‘second hit’ of adulthood psychological stress alters central inflammatory mediators: Implications for female fertility

The fundamentals of female reproductive health are established in early life, where immune pathways contribute to reproductive development. Inflammatory disturbances during this critical period permanently modify reproductive functioning. Moreover, early life stress is known to create a predisposition for subsequent stress vulnerability. Using a rat model, we have previously demonstrated that neonatal immune activation (NIA) with bacterial mimetic lipopolysaccharide (LPS) is associated with; altered immune milieu, advanced female puberty, depleted ovarian reserve, mating impairments, and stress vulnerability. What has not been established are the central inflammatory changes associated with this phenotype and the impact of psychological stress. In the current study, female rats were exposed to intraperitoneal injection of saline (control) or LPS (0.05 mg/kg) to induce NIA on postnatal days 3 and 5. In adulthood, half of the animals were exposed to restraint stress. NIA advanced puberty onset and reduced adulthood ovarian follicle populations (p

Neonatal infection produces sex-specific changes in neuroimmune function with no associated deficits in spatial learning in juvenile rats

Immune activation during early development can have profound effects on later immune function, cognition, and behavior. Epidemiological data indicate a strong correlation between early-life immune activation and later diagnosis of disorders such as autism, schizophrenia, and depression. However, there has been little investigation into the impact of sex or early-life immune activation on the developing immune system or the ontogeny of cognitive processes, such as learning. Microglia are the resident immune cells of the brain and affect neural function and behavior. Male rats have more microglia than female rats in the hippocampus (HP) on postnatal day four (P4), suggesting that males may be more vulnerable to the long- term consequences of early-life immune activation than females. We examined the effect of neonatal immune activation (E.coli; 1 × 106 CFU/0.1 mL/kg) alone, or following a second immune challenge of LPS (Lipopolysaccharide; 25 μg/mL/kg), during late juvenile development on behavior and microglia function in males and females. Neither neonatal infection nor a second immune activation produced learning deficits. However, neonatal infection produced sex-specific alterations in CD11b, IL-β, and IL-6 in the HP, prefrontal cortex (PFC), and cerebellum. Additionally, we found sex-specific alterations in microglial density in CA3, dentate gyrus, and PFC. These data suggest that neonatal infection produces sex-specific changes in cytokine expression and microglia activation in juveniles, but the consequences of these changes may have a delayed effect on behavioral outcomes.

Neonatal immune activation depletes the ovarian follicle reserve and alters ovarian acute inflammatory mediators in neonatal rats†

Normal ovarian development is crucial for female reproductive success and longevity. Interruptions to the delicate process of initial folliculogenesis may lead to ovarian dysfunction. We have previously demonstrated that an early life immune challenge in the rat, induced by administration of lipopolysaccharide (LPS) on postnatal day (PND) 3 and 5, depletes ovarian follicle reserve long term. Here, we hypothesized that this neonatal immune challenge leads to an increase in peripheral and ovarian inflammatory signaling, contributing to an acute depletion of ovarian follicles. Morphological analysis of neonatal ovaries indicated that LPS administration significantly depleted PND 5 primordial follicle populations and accelerated follicle maturation. LPS exposure upregulated circulating interleukin 6, tumor necrosis factor alpha (TNFa), and C-reactive protein on PND 5, and upregulated ovarian mRNA expression of Tnfa, mitogen-activated protein kinase 8 (Mapk8/Jnk1), and growth differentiation factor 9 (Gdf9) (P < 0.05). Mass spectrometry and cell signaling pathway analysis indicated upregulation of cellular pathways associated with acute phase signaling, and cellular survival and assembly. Apoptosis assessed by terminal deoxynucleotidyl transferase dUTP nick end labeling indicated significantly increased positive staining in the ovaries of LPS-treated neonates. These findings suggest that increased proinflammatory signaling within the neonatal ovary may be responsible for the LPS-induced depletion of the primordial follicle pool. These findings also have implications for female reproductive health, as the ovarian reserve is a major determinate of female reproductive longevity.

Two-hit model of schizophrenia induced by neonatal immune activation and peripubertal stress in rats: Study of sex differences and brain oxidative alterations

Schizophrenia is considered to be a developmental disorder with distinctive sex differences. Aiming to simulate the vulnerability of the third trimester of human pregnancy to the developmental course of schizophrenia, an animal model was developed, using neonatal poly(I:C) as a first-hit, and peripubertal stress as a second-hit, i.e. a two-hit model. Since, to date, there have been no references to sex differences in the two-hit model, our study sought to determine sex influences on the development of behavior and brain oxidative change in adult rats submitted to neonatal exposure to poly(I:C) on postnatal days 5–7 as well as peripubertal unpredictable stress (PUS). Our results showed that adult two-hit rats present sex-specific behavioral alterations, with females showing more pronounced deficits in prepulse inhibition of the startle reflex and hyperlocomotion, while males showing more deficits in social interaction. Male and female animals exhibited similar working memory deficits. The levels of the endogenous antioxidant, reduced glutathione, were decreased in the prefrontal cortex (PFC) of both male and female animals exposed to both poly(I:C) and poly(I:C) + PUS. Only females presented decrements in GSH levels in the striatum. Nitrite levels were increased in the PFC of male and in the striatum of female poly(I:C) + PUS rats. Increased lipid peroxidation was observed in the PFC of females and in the striatum of males and females exposed to poly(I:C) and poly(I:C) + PUS. Thus, the present study presents evidence for sex differences in behavior and oxidative brain change induced by a two-hit model of schizophrenia.

Immune responses in perinatal brain injury

The perinatal period has often been described as immune deficient. However, it has become clear that immune responses in the neonate following exposure to microbes or as a result of tissue injury may be substantial and play a role in perinatal brain injury. In this article we will review the immune cell composition under normal physiological conditions in the perinatal period, both in the human and rodent. We will summarize evidence of the inflammatory responses to stimuli and discuss how neonatal immune activation, both in the central nervous system and in the periphery, may contribute to perinatal hypoxic-ischemic brain injury.

P.1.h.047 - Pubertal stress and neonatal immune activation with lipopolysaccharide promotes anxious- and negative-like schizophrenia symptom in adulthood rats: effect of gender

P.1.h.047 - Pubertal stress and neonatal immune activation with lipopolysaccharide promotes anxious- and negative-like schizophrenia symptom in adulthood rats: effect of gender

The expression and function of human CD300 receptors on blood circulating mononuclear cells are distinct in neonates and adults.

Neonates are more susceptible to infections than adults. This susceptibility is thought to reflect neonates’ qualitative and quantitative defects in the adaptive and innate immune responses. Differential expression of cell surface receptors may result in altered thresholds of neonatal immune cell activation. We determined whether the expression and function of the lipid-binding CD300 family of receptors are different on neonatal immune cells compared to adult immune cells. A multiparametric flow cytometry analysis was performed to determine the expression of CD300 receptors on adult peripheral blood mononuclear cells and neonatal cord blood mononuclear cells. The expression of the CD300a inhibitory receptor was significantly reduced on cells from the newborn adaptive immune system, and neonatal antigen presenting cells exhibited a different CD300 receptors expression pattern. We also found differential LPS-mediated regulation of CD300 receptors expression on adult monocytes compared to cord blood monocytes, and that CD300c and CD300e-mediated activation was quantitatively different in neonatal monocytes. This is the first complete study examining the expression of CD300 receptors on human neonatal immune cells compared with adult immune cells. Significant differences in the expression and function of CD300 receptors may help to explain the peculiarities and distinctness of the neonatal immune responses.

Abstract # 1761 Neonatal immune activation with lipopolysaccharide and a ‘second hit’ of adulthood stress alters ovarian inflammatory mediators: Implications for female subfertility

Agnogentic subfertility is a clinical concern for many women of a healthy reproductive age. Accumulating evidence suggests developmental factors, such as immune status, are involved in the aetiology of female subfertility. Normal immune function is crucial for the initial quality and quantity of the ovarian follicular pool, and continued reproductive success. Our laboratory, in parallel with others, has established that early-life immune stress leads to sustained alterations in immune and neuroendocrine function, and perinatally programs vulnerability to subsequent stressors. However, investigation into the effects of early-life immune stress on female reproductive development is limited. This study investigates the immediate and long-term effects of immune activation during early-life on ovarian development and continued female reproductive health. Using a rat model, our findings suggest that neonatal immune activation (NIA) with lipopolysaccharide on post-natal days 3 and 5 leads to significant depletion of the ovarian follicular pool, acute upregulation of ovarian proinflammatory mediators, and precocious onset of puberty ( p p

Developmental minocycline treatment reverses the effects of neonatal immune activation on anxiety- and depression-like behaviors, hippocampal inflammation, and HPA axis activity in adult mice.

Neonatal infection is associated with increased lifetime risk for neuropsychiatric disorders including anxiety and depression, with evidence showing that dysregulation of the hypothalamic-pituitary-adrenal-(HPA)-axis system may be partly responsible. Preclinical and clinical studies demonstrate that minocycline exhibits antidepressant effects through inhibition of microglial activation and anti-inflammatory actions, and of interest is that recent studies suggest that minocycline alleviates the behavioral abnormalities induced by early-life insults. The current study was designed to determine if developmental minocycline treatment attenuates the neonatal immune activation-induced anxiety- and depression-like symptoms and HPA-axis-dysregulation later in life. To this end, neonatal mice were treated to either lipopolysaccharide or saline on postnatal days (PND) 3-5, then dams during lactation (PND 6-20) and male offspring during adolescence (PND 21-40) received oral administration of minocycline or water via regular drinking bottles. Anxiety- and depression-like behaviors, HPA-axis-reactivity (corticosterone), and hippocampal inflammation (TNF-α and IL-1β) after exposure to stress were evaluated. The results indicated that neonatal immune activation resulted in increased anxiety and depression-like symptoms, HPA-axis-hyperactivity, and elevated the levels of TNF-α and IL-1β in the hippocampus in response to stress in adulthood. Interestingly, developmental minocycline treatment significantly reduced the abnormalities induced by neonatal inflammation in adult mice. In addition, minocycline, regardless of postnatal inflammation, did not have any detrimental effects on the above measured parameters. Considering that minocycline is currently under exploration as an alternative or adjunctive therapy for reducing the symptoms of neurological disorders, our findings suggest that minocycline during development can decrease the behavioral abnormalities induced by early life inflammation in adulthood.

Neonatal immune activation during early and late postnatal brain development differently influences depression-related behaviors in adolescent and adult C57BL/6 mice

Neuroimmunology and Neuroinflammation is an open access journal, with focuses on neuroimmunology and neuroinflammation research, and coverage extending to other basic and clinical studies related to neuroscience.