In vitro modulation of mTOR and mGlur5 influence α-synuclein accumulation.

One of the main hallmarks of Parkinson's disease (PD) is abnormal alpha-synuclein (α-syn) aggregation which forms the main component of intracellular Lewy body inclusions. This short report used preformed α-syn fibrils, as well as an A53T mutant α-syn adenovirus to mimic conditions of pathological protein aggregation in dopaminergic human derived SH-SY5Y neural cells. Since there is evidence that the mTOR pathway and glutamatergic signaling each influence protein aggregation, we also assessed the impact of the mTOR inhibitor, rapamycin and the mGluR5 allosteric modulator, CTEP. We found that both rapamycin and CTEP induced a significant reduction of α-syn fibrils in SH-SY5Y cells and this effect was associated with a reduction in mTOR signaling and enhancement in autophagic pathway factors. These data support the possibility that CTEP (or rapamycin) might be a useful pharmacological approach to target abnormal α-syn accumulation by promoting intracellular degradation or enhanced clearance.

Brain-derived neurotrophic factor (BDNF) has direct anti-inflammatory effects on microglia.

Microglia are the primary immunocompetent cells that protect the brain from environmental stressors, but can also be driven to release pro-inflammatory cytokines and induce a cytotoxic environment. Brain-derived neurotrophic factor (BDNF) is important for the regulation of plasticity, synapse formation, and general neuronal health. Yet, little is known about how BDNF impacts microglial activity. We hypothesized that BDNF would have a direct modulatory effect on primary cortical (Postnatal Day 1-3: P1-3) microglia and (Embryonic Day 16: E16) neuronal cultures in the context of a bacterial endotoxin. To this end, we found that a BDNF treatment following LPS-induced inflammation had a marked anti-inflammatory effect, reversing the release of both IL-6 and TNF-α in cortical primary microglia. This modulatory effect was transferrable to cortical primary neurons, such that LPS-activated microglial media was able produce an inflammatory effect when added to a separate neuronal culture, and again, BDNF priming attenuated this effect. BDNF also reversed the overall cytotoxic impact of LPS exposure in microglia. We speculate that BDNF can directly play a role in regulating microglia state and hence, influence microglia-neuron interactions.

LRRK2 and WAVE2 regulate microglial-transition through distinct morphological phenotypes to induce neurotoxicity in a novel two-hit in vitro model of neurodegeneration.

We report a novel in vitro classification system that tracks microglial activation state and their potential neurotoxicity. Mixed live-cell imaging was used to characterize transition through distinct morphological phenotypes, production of reactive oxygen species (ROS), formation of reactive microglial aggregates, and subsequent cytokine production. Transwell cultures were used to determine microglial migration (control and lipopolysaccharide (LPS) treated) to glutamate pre-stressed or healthy neurons. This two-hit paradigm was developed to model the vast evidence that neurodegenerative conditions, like Parkinson's disease (PD), may stem from the collective impact of multiple environmental stressors. We found that healthy neurons were resistant to microglial-mediated inflammation, whereas glutamate pre-stressed neurons were highly susceptible and in fact, appeared to recruit microglia. The LPS treated microglia progressed through distinct morphological states and expressed high levels of ROS and formed large cellular aggregates. Recent evidence implicates leucine-rich repeat kinase 2 (LRRK2) as an important player in the microglial inflammatory state, as well as in the genesis of PD. We found that inhibition of the LRRK2 signaling pathway using the kinase inhibitor cis-2,6-dimethyl-4-(6-(5-(1-methylcyclopropoxy)-1H-indazol-3-yl)pyrimidin-4-yl)morpholine (MLi2) or inhibition of the actin regulatory protein, Wiskott-Aldrich syndrome family Verprolin-homologous Protein-2 (WAVE2), stunted microglial activation and prevented neurotoxicity. Furthermore, inhibition of LRRK2 kinase activity reduced pro-inflammatory chemokines including MIP-2, CRG-2, and RANTES. These data together support the notion that LRRK2 and WAVE2 are important mediators of cytokine production and cytoskeletal rearrangement necessary for microglial-induced neurotoxicity. Furthermore, our model demonstrated unique microglial phenotypic changes that might be mechanistically important for better understanding neuron-microglial crosstalk.

Neuroimmune multi-hit perspective of coronaviral infection.

It is well accepted that environmental stressors experienced over a one's life, from microbial infections to chemical toxicants to even psychological stressors, ultimately shape central nervous system (CNS) functioning but can also contribute to its eventual breakdown. The severity, timing and type of such environmental "hits", woven together with genetic factors, likely determine what CNS outcomes become apparent. This focused review assesses the current COVID-19 pandemic through the lens of a multi-hit framework and disuses how the SARS-COV-2 virus (causative agent) might impact the brain and potentially interact with other environmental insults. What the long-term consequences of SAR2 COV-2 upon neuronal processes is yet unclear, but emerging evidence is suggesting the possibility of microglial or other inflammatory factors as potentially contributing to neurodegenerative illnesses. Finally, it is critical to consider the impact of the virus in the context of the substantial psychosocial stress that has been associated with the global pandemic. Indeed, the loneliness, fear to the future and loss of social support alone has exerted a massive impact upon individuals, especially the vulnerable very young and the elderly. The substantial upswing in depression, anxiety and eating disorders is evidence of this and in the years to come, this might be matched by a similar spike in dementia, as well as motor and cognitive neurodegenerative diseases.

Microglia and BDNF at the crossroads of stressor related disorders: Towards a unique trophic phenotype.

Stressors ranging from psychogenic/social to neurogenic/injury to systemic/microbial can impact microglial inflammatory processes, but less is known regarding their effects on trophic properties of microglia. Recent studies do suggest that microglia can modulate neuronal plasticity, possibly through brain derived neurotrophic factor (BDNF). This is particularly important given the link between BDNF and neuropsychiatric and neurodegenerative pathology. We posit that certain activated states of microglia play a role in maintaining the delicate balance of BDNF release onto neuronal synapses. This focused review will address how different "activators" influence the expression and release of microglial BDNF and address the question of tropomyosin receptor kinase B (TrkB) expression on microglia. We will then assess sex-based differences in microglial function and BDNF expression, and how microglia are involved in the stress response and related disorders such as depression. Drawing on research from a variety of other disorders, we will highlight challenges and opportunities for modulators that can shift microglia to a "trophic" phenotype with a view to potential therapeutics relevant for stressor-related disorders.

Characterizing the protracted neurobiological and neuroanatomical effects of paraquat in a murine model of Parkinson's disease.

The primary motor symptoms of Parkinson's disease (PD) result from the degeneration of dopamine-producing neurons of the substantia nigra pars compacta (SNc), and often, the loss is asymmetrical, resulting in unilateral tremor presentation. Notably, age is the primary risk factor for PD, and it is likely that the disease ultimately stems from the impact of environmental factors, which interact with the aging process. Recent research has focused on the role of microglia and pro-oxidative responses in dopaminergic neuronal death. In this study, we sought to examine the neurodegenerative, inflammatory, and stress effects of exposure to the etiologically relevant pesticide, paraquat, over time (up to 6 months after injections). We also were interested in whether a high-resolution, 7-Tesla animal magnetic resonance imaging would be sensitive enough to detect the degenerative impact of paraquat. We found that paraquat induced a loss of dopaminergic SNc neurons and activation of microglia that surprisingly did not change over 6 months after the last injection. A long-lasting reduction was evident for body weight, and alterations in organ (lung and heart) weight were evident, which reflect the peripheral impact of the toxicant. The microglial proinflammatory actin-remodeling factor, WAVE2, along with the inflammatory transcription factor, nuclear factor kappa B were also elevated within the brain. Remarkably, the stress hormone, corticosterone, was still significantly elevated 1 month after paraquat, whereas the inflammasome factor, caspase-1, and antigen presentation factor, MFG-E8, both displayed delayed rises after the 6-month time. Using high-resolution magnetic resonance imaging, we detected no striatal changes but modest hemispheric differences in the SNc and time-dependent volumetric enlargement of the ventricles in paraquat-treated mice. These data suggest that paraquat induces long-term nigrostriatal pathology (possibly asymmetric) and inflammatory changes and stress and trophic/apoptotic effects that appear to either increase with the passage of time or are evident for at least 1 month. In brief, paraquat may be a useful nonspecific means to model widespread stress and inflammatory changes related to PD or age-related disease in general, but not the progressive nature of such diseases.

The impact of dextran sodium sulphate and probiotic pre-treatment in a murine model of Parkinson's disease.

BACKGROUND: Recent work has established that Parkinson's disease (PD) patients have an altered gut microbiome, along with signs of intestinal inflammation. This could help explain the high degree of gastric disturbances in PD patients, as well as potentially be linked to the migration of peripheral inflammatory factors into the brain. To our knowledge, this is the first study to examine microbiome alteration prior to the induction of a PD murine model. METHODS: We presently assessed whether pre-treatment with the probiotic, VSL #3, or the inflammatory inducer, dextran sodium sulphate (DSS), would influence the PD-like pathology provoked by a dual hit toxin model using lipopolysaccharide (LPS) and paraquat exposure. RESULTS: While VSL #3 has been reported to have anti-inflammatory effects, DSS is often used as a model of colitis because of the gut inflammation and the breach of the intestinal barrier that it induces. We found that VSL#3 did not have any significant effects (beyond a blunting of LPS paraquat-induced weight loss). However, the DSS treatment caused marked changes in the gut microbiome and was also associated with augmented behavioral and inflammatory outcomes. In fact, DSS markedly increased taxa belonging to the Bacteroidaceae and Porphyromonadaceae families but reduced those from Rikencellaceae and S24-7, as well as provoking colonic pro-inflammatory cytokine expression, consistent with an inflamed gut. The DSS also increased the impact of LPS plus paraquat upon microglial morphology, along with circulating lipocalin-2 (neutrophil marker) and IL-6. Yet, neither DSS nor VSL#3 influenced the loss of substantia nigra dopamine neurons or the astrocytic and cytoskeleton remodeling protein changes that were provoked by the LPS followed by paraquat treatment. CONCLUSIONS: These data suggest that disruption of the intestinal integrity and the associated microbiome can interact with systemic inflammatory events to promote widespread brain-gut changes that could be relevant for PD and at the very least, suggestive of novel neuro-immune communication.

Depression, dementia and immune dysregulation.

Major depression is a prevalent illness that increases the risk of several neurological conditions. These include stroke, cardiovascular disease, and dementia including Alzheimer's disease. In this review we ask whether certain types of depression and associated loneliness may be a harbinger of cognitive decline and possibly even dementia. We propose that chronic stress and inflammation combine to compromise vascular and brain function. The resulting increases in proinflammatory cytokines and microglial activation drive brain pathology leading to depression and mild cognitive impairment, which may progress to dementia. We present evidence that by treating the inflammatory changes, depression can be reversed in many cases. Importantly, there is evidence that anti-inflammatory and antidepressant treatments may reduce or prevent dementia in people with depression. Thus, we propose a model in which chronic stress and inflammation combine to increase brain permeability and cytokine production. This leads to microglial activation, white matter damage, neuronal and glial cell loss. This is first manifest as depression and mild cognitive impairment, but can eventually evolve into dementia. Further research may identify clinical subgroups with inflammatory depression at risk for dementia. It would then be possible to address in clinical trials whether effective treatment of the depression can delay the onset of dementia.

Early life stress facilitates synapse premature unsilencing to enhance AMPA receptor function in the developing hippocampus.

Chronic early life stress (ELS) increases vulnerability to psychopathologies and cognitive deficits in adulthood by disrupting the function of related neural circuits. However, whether this disruption emerges early in the developing brain remains largely unexplored. In the current study, using an established limited-bedding and nesting model of ELS in postnatal day (P)2-10 mice, we provide direct evidence that ELS caused early modification of hippocampal glutamatergic synapses in the developing brain. We demonstrated that ELS induced rapid enhancement of AMPA receptor function in hippocampal CA1 pyramidal neurons through a postsynaptic mechanism, and importantly, this was associated with premature unsilencing of NMDA receptor-only silent hippocampal synapses. These results suggest that potentiation of AMPAR function may represent an early mediator of ELS-induced alterations of neural networks in the developing brain and may potentially contribute to subsequent cognitive impairments later in life.NEW & NOTEWORTHY Early life stress (ELS) is known to increase the risk of later life cognitive deficits by disrupting neural circuit function. However, whether this disruption emerges early in the developing brain remains largely unexplored. The current study presents direct evidence that ELS prematurely unsilences hippocampal synapses to enhance AMPA receptor functions in a limited-bedding and nesting model, revealing an early mediator of ELS-induced neural circuit reorganizations.

Alleviating toxic α-Synuclein accumulation by membrane depolarization: evidence from an in vitro model of Parkinson's disease.

Parkinson's disease (PD) is characterized by the formation of toxic, fibrillar form alpha-synuclein (α-Syn) protein aggregates in dopaminergic neurons. Accumulating evidence has shown a multifactorial interplay between the intracellular calcium elevation and α-Syn dynamics. However, whether membrane depolarization regulates toxic α-Syn aggregates remains unclear. To understand this better, we used an in vitro α-Syn preformed fibrils (PFF) model of PD in human neural cells. We demonstrated functional membrane depolarization in differentiated SH-SY5Y cells induced by two independent treatments: high extracellular K+ and the GABAA receptor blocker picrotoxin. We then observed that these treatments significantly alleviated toxic α-Syn aggregation in PFF-treated SH-SY5Y cells. Moreover, clinically relevant direct current stimulation (DCS) also remarkably decreased toxic α-Syn aggregation in PFF-treated SH-SY5Y cells. Taken together, our findings suggest that membrane depolarization plays an important role in alleviating PFF-induced toxic α-Syn aggregates, and that it may represent a novel therapeutic mechanism for PD.

Leucine-rich repeat kinase-2 (LRRK2) modulates microglial phenotype and dopaminergic neurodegeneration.

Leucine-rich repeat kinase 2 (LRRK2) is a common gene implicated in Parkinson's disease and many inflammatory processes. Thus, we assessed the role of LRRK2 in the context of endotoxin (lipopolysaccharide, LPS)-induced inflammation of the substantia nigra together with the environmental toxicant, paraquat, that has been implicated in PD. Here we found that LRRK2 ablation prevented the loss of dopaminergic neurons and behavioral deficits (motor) induced by LPS priming followed by paraquat exposure. The LRRK2 ablation also provoked a phenotypic shift in LPS-primed microglia cells. The LRRK2 deficiency reduced their "activated" morphology and upregulation of the inflammatory phagocytic regulator, WAVE2 (critical for actin remodeling), while the chemokine receptor, CX3CR1, was elevated in isolated CD11b+ myeloid cells. Furthermore, LRRK2 knockout attenuated the signs of oxidative stress and morphological changes induced in primary microglia by LPS treatment. However, induced WAVE2 expression together with LPS exposure in microglia overcame the inhibitory effects of LRRK2 knockout, suggesting WAVE2 may be acting downstream of LRRK2. Neither WAVE2 nor did LRRK2 knockout influence LPS-induced cytokine elevations in the microglia. We are the first to show the importance of LRRK2 in neurodegenerative and inflammatory processes in this multi-hit toxin model of PD. These data are consistent with the proposition that LRRK2 and WAVE2 are useful therapeutic targets for PD or other conditions with a prominent neuroinflammatory component.

Microglia depletion prior to lipopolysaccharide and paraquat treatment differentially modulates behavioral and neuronal outcomes in wild type and G2019S LRRK2 knock-in mice.

BACKGROUND: Substantial data have implicated microglial-driven neuroinflammation in Parkinson's disease (PD) and environmental toxicants have been long expected as triggers of such inflammatory processes. Of course, these environmental insults act in the context of genetic vulnerability factors and in this regard, leucine rich repeat kinase 2 (LRRK2), may play a prominent role. METHODS: We used a double hit, lipopolysaccharide (LPS; endotoxin) followed by paraquat (pesticide toxicant) model of PD in mice with the most common LRRK2 mutation G2019S, knockin mice and wild type littermates. In order to assess the contribution of microglia, we depleted these cells (through 14 days of the CSF-1 antagonist, PLX-3397) prior to LPS and paraquat exposure. RESULTS: We found that the G2019S mice displayed the greatest signs of behavioral pathology, but that the PLX-3397 induced microglial depletion at the time of LPS exposure diminished toxicity and weight loss and blunted the reduction in home-cage activity with subsequent paraquat exposure. However, neither the PLX-3397 pre-treatment nor the G2019S mutation affected the LPS â€‹+ â€‹paraquat induced loss of substantia nigra pars compacta (SNc) dopamine neurons or elevation of circulating immune (IL-6) or stress (corticosterone) factors. Intriguingly, microglial morphological ratings were basally enhanced in G2019S mice and the PLX-3397 pre-treatment reversed this effect. Moreover, PLX-3397 pre-treatment selectively elevated soluble a-synuclein and SIRT3 levels, while reducing SNc caspase-1 and 3, along with CX3CR1. Hence, the re-populated "new" microglia following cessation of PLX-3397 clearly had an altered phenotype or were immature at the time of sacrifice (i.e. after 11 days). CONCLUSIONS: Collectively, these findings suggest that G2019S knock-in and PLX-3397 microglial depletion at the time of LPS exposure affects behavioral, but not neurodegenerative responses to subsequent environmental toxin exposure.

mGluR5 Allosteric Modulation Promotes Neurorecovery in a 6-OHDA-Toxicant Model of Parkinson's Disease.

Parkinson's disease is a neurodegenerative disease characterized by a loss of dopaminergic substantia nigra neurons and depletion of dopamine. To date, current therapeutic approaches focus on managing motor symptoms and trying to slow neurodegeneration, with minimal capacity to promote neurorecovery. mGluR5 plays a key role in neuroplasticity, and altered mGluR5 signaling contributes to synucleinopathy and dyskinesia in patients with Parkinson's disease. Here, we tested whether the mGluR5-negative allosteric modulator, (2-chloro-4-[2[2,5-dimethyl-1-[4-(trifluoromethoxy) phenyl] imidazol-4-yl] ethynyl] pyridine (CTEP), would be effective in improving motor deficits and promoting neural recovery in a 6-hydroxydopamine (6-OHDA) mouse model. Lesions were induced by 6-ODHA striatal infusion, and 30 days later treatment with CTEP (2 mg/kg) or vehicle commenced for either 1 or 12 weeks. Animals were subjected to behavioral, pathological, and molecular analyses. We also assessed how long the effects of CTEP persisted, and finally, using rapamycin, determined the role of the mTOR pathway. CTEP treatment induced a duration-dependent improvement in apomorphine-induced rotation and performance on rotarod in lesioned mice. Moreover, CTEP promoted a recovery of striatal tyrosine hydroxylase-positive fibers and normalized FosB levels in lesioned mice. The beneficial effects of CTEP were paralleled by an activation of mammalian target of rapamycin (mTOR) pathway and elevated brain-derived neurotrophic factor levels in the striatum of lesioned mice. The mTOR inhibitor, rapamycin (sirolimus), abolished CTEP-induced neurorecovery and rescue of motor deficits. Our findings indicate that mTOR pathway is a useful target to promote recovery and that mGluR5 allosteric regulators may potentially be repurposed to selectively target this pathway to enhance neuroplasticity in patients with Parkinson's disease.

Quantum dot conjugated saporin activates microglia and induces selective substantia nigra degeneration.

Parkinson's disease (PD) is characterized by profound microglial driven inflammatory processes and the loss of dopamine neurons of the substantia nigra (SNc). Both microglia and dopamine neurons that are affected in the SNc are particularly vulnerable to environmental toxicants and finding more selective ways of targeting these cell types is of importance. Quantum dots (QDs) might be a useful vehicle for selectively delivering toxicants to microglia and owing to their fluorescent capability, they can be microscopically tracked within the cell. Accordingly, we assessed the impact of QDs alone and QDs conjugated to the ribosomal toxin, saporin, upon SNc microglia and dopamine neurons. We found that intra-SNc infused QDs selectively entered microglia and induced morphological changes consistent with an activated state. QDs conjugated to saporin also caused a significant loss of dopamine neurons and motor coordination (on a rotarod test) deficits, along with an increase in the inflammatory microglial actin regulatory factors, WAVE2. These data suggest that QDs might be a viable route for toxicant delivery and also has an added advantage of being fluorescently visible. Ultimately, we found SNc neurons to be exceptionally vulnerable to QD-saporin and suggest that this could be a novel targeted approach to model PD-like inflammatory pathology.

Lrrk2 alleles modulate inflammation during microbial infection of mice in a sex-dependent manner.

Variants in the leucine-rich repeat kinase-2 (LRRK2) gene are associated with Parkinson's disease, leprosy, and Crohn's disease, three disorders with inflammation as an important component. Because of its high expression in granulocytes and CD68-positive cells, LRRK2 may have a function in innate immunity. We tested this hypothesis in two ways. First, adult mice were intravenously inoculated with Salmonella typhimurium, resulting in sepsis. Second, newborn mouse pups were intranasally infected with reovirus (serotype 3 Dearing), which induced encephalitis. In both mouse models, wild-type Lrrk2 expression was protective and showed a sex effect, with female Lrrk2-deficient animals not controlling infection as well as males. Mice expressing Lrrk2 carrying the Parkinson's disease-linked p.G2019S mutation controlled infection better, with reduced bacterial growth and longer animal survival during sepsis. This gain-of-function effect conferred by the p.G2019S mutation was mediated by myeloid cells and was abolished in animals expressing a kinase-dead Lrrk2 variant, p.D1994S. Mouse pups with reovirus-induced encephalitis that expressed the p.G2019S Lrrk2 mutation showed increased mortality despite lower viral titers. The p.G2019S mutant Lrrk2 augmented immune cell chemotaxis and generated more reactive oxygen species during virulent infection. Reovirus-infected brains from mice expressing the p.G2019S mutant Lrrk2 contained higher concentrations of α-synuclein. Animals expressing one or two p.D1994S Lrrk2 alleles showed lower mortality from reovirus-induced encephalitis. Thus, Lrrk2 alleles may alter the course of microbial infections by modulating inflammation, and this may be dependent on the sex and genotype of the host as well as the type of pathogen.

Chronic unpredictable stress influenced the behavioral but not the neurodegenerative impact of paraquat.

The impact of psychological stressors on the progression of motor and non-motor disturbances observed in Parkinson's disease (PD) has received little attention. Given that PD likely results from many different environmental "hits", we were interested in whether a chronic unpredictable stressor regimen would act additively or possibly even synergistically to augment the impact of the toxicant, paraquat, which has previously been linked to PD. Our findings support the contention that paraquat itself acted as a systemic stressor, with the pesticide increasing plasma corticosterone, as well as altering glucocorticoid receptor (GR) expression in the hippocampus. Furthermore, stressed mice that also received paraquat displayed synergistic motor coordination impairment on a rotarod test and augmented signs of anhedonia (sucrose preference test). The individual stressor and paraquat treatments also caused a range of non-motor (e.g. open field, Y and plus mazes) deficits, but there were no signs of an interaction (neither additive nor synergistic) between the insults. Similarly, paraquat caused the expected loss of substantia nigra dopamine neurons and microglial activation, but this effect was not further influenced by the chronic stressor. Taken together, these results indicate that paraquat has many effects comparable to that of a more traditional stressor and that at least some behavioral measures (i.e. sucrose preference and rotarod) are augmented by the combined pesticide and stress treatments. Thus, although psychological stressors might not necessarily increase the neurodegenerative effects of the toxicant exposure, they may promote co-morbid behaviors pathology.

Leucine-rich repeat kinase-2 (LRRK2) modulates paraquat-induced inflammatory sickness and stress phenotype.

BACKGROUND: Leucine-rich repeat kinase 2 (LRRK2) is a common gene implicated in Parkinson's disease (PD) and is also thought to be fundamentally involved in numerous immune functions. Thus, we assessed the role of LRRK2 in the context of the effects of the environmental toxicant, paraquat, that has been implicated in PD and is known to affect inflammatory processes. METHODS: Male LRRK2 knockout (KO) and transgenic mice bearing the G2019S LRRK2 mutation (aged 6-8 months) or their littermate controls were exposed to paraquat (two times per week for 3 weeks), and sickness measures, motivational scores, and total home-cage activity levels were assessed. Following sacrifice, western blot and ELISA assays were performed to see whether or not LRRK2 expression would alter processes related to plasticity, immune response processes, or the stress response. RESULTS: Paraquat-induced signs of sickness, inflammation (elevated IL-6), and peripheral toxicity (e.g., organ weight) were completely prevented by LRRK2 knockout. In fact, LRRK2 knockout dramatically reduced not only signs of illness, but also the motivational (nest building) and home-cage activity deficits induced by paraquat. Although LRRK2 deficiency did not affect the striatal BDNF reduction that was provoked by paraquat, it did blunt the corticosterone elevation induced by paraquat, raising the possibility that LRRK2 may modulate aspects of the HPA stress axis. Accordingly, we found that transgenic mice bearing the G2019S LRRK2 mutation had elevated basal corticosterone, along with diminished hippocampal 5-HT1A levels. CONCLUSION: We are the first to show the importance of LRRK2 in the peripheral neurotoxic and stressor-like effects of paraquat. These data are consistent with LRRK2 playing a role in the general inflammatory tone and stressor effects induced by environmental toxicant exposure.

Paraquat Exposure Increases Oxidative Stress Within the Dorsal Striatum of Male Mice With a Genetic Deficiency in One-carbon Metabolism.

Paraquat is an herbicide that is commonly used worldwide. Exposure to paraquat results in Parkinson's disease (PD)-like symptoms including dopaminergic cell loss. Nutrition has also been linked in the pathogenesis of PD, such as reduced levels of folic acid, a B-vitamin, and component of one-carbon metabolism. Within one-carbon metabolism, methylenetetrahydrofolate reductase (MTHFR) catalyzes the irreversible conversion of 5, 10-methylenetetrahydrofolate to 5-methyltetrahydrofolate. A polymorphism in MTHFR (677 C&→T) has been reported in 5%-15% of North American and European human populations. The MTHFR polymorphism is also prevalent in PD patients. The goal of this study was to investigate the impact of paraquat-induced PD-like pathology in the context of reduced levels of MTHFR. Three-month-old male Mthfr+/- mice, which model the MTHFR polymorphism observed in humans, were administered intraperitoneal injections of paraquat (10 mg/kg) or saline 6 times over 3 weeks. At the end of paraquat treatment, motor and memory function were assessed followed by collection of brain tissue for biochemical analysis. Mthfr+/- mice treated with paraquat showed impaired motor function. There was increased microglial activation within the substantia nigra (SN) of Mthfr+/- mice treated with paraquat. Additionally, all Mthfr+/- mice that were treated with paraquat showed increased oxidative stress within the dorsal striatum, but not the SN. The present results show that paraquat exposure increases PD-like pathology in mice deficient in one-carbon metabolism.

Perinatal antibiotic exposure alters composition of murine gut microbiota and may influence later responses to peanut antigen.

BACKGROUND: Accumulating evidence suggests that the gut microbiota shapes developmental processes within the immune system. Early life antibiotic use is one factor which may contribute to immune dysfunction and the recent surge in allergies by virtue of its effects on gut microbiota. OBJECTIVE AND METHODS: As a first step towards determining whether a relationship exists between perinatal antibiotic induced changes in the gut microbiota and the later development of a peanut allergy, we exposed newborn mice to either the broad-spectrum antibiotic vancomycin or to a vehicle for 6 weeks and then used a novel murine model of peanut allergy. RESULTS: Early-life treatment with vancomycin resulted in a significant shift in the gut microbiota community characterized by a reduction in the abundance of firmicutes and preponderance of inflammatory proteobacteria. Mice with an antibiotic-altered microbiota, showed a localized allergic-like response characterized by ear swelling and scratching following intra-dermal peanut antigen challenge. Likewise, circulating IgE levels were increased in antibiotic-treated mice, but no evidence of a systemic allergic or anaphylactic-like response was observed. Importantly, we utilized the naturally occurring pro-inflammatory cytokine, tumor necrosis factor-α (TNF-α), rather than the more commonly used cholera toxin, as an adjuvant together with the peanut antigen. CONCLUSION: Our data suggest that early antibiotic exposure promotes a shift in the gut microbiota community that may in turn, influence how mice later respond to a TNF-α + antigen challenge. However, further studies verifying the capacity of microbiota restoration to protect against allergic responses will be needed to confirm a causal role of antibiotic-induced microbiota variations in promoting allergic disease phenotypes.

Profiling changes in cortical astroglial cells following chronic stress.

Recent studies have suggested that cortical astroglia play an important role in depressive-like behaviors. Potential astroglial contributions have been proposed based on their known neuroplastic functions, such as glutamate recycling and synaptic plasticity. However, the specific mechanisms by which astroglial cells may contribute or protect against a depressive phenotype remain unknown. To delineate astroglial changes that accompany depressive-like behavior, we used astroglial-specific bacTRAP mice exposed to chronic variable stress (CVS) and profiled the astroglial translatome using translating ribosome affinity purification (TRAP) in conjunction with RNAseq. As expected, CVS significantly increased anxiety- and depressive-like behaviors and corticosterone levels and decreased GFAP expression in astroglia, although this did not reflect a change in the total number of astroglial cells. TRAPseq results showed that CVS decreased genes associated with astroglial plasticity: RhoGTPases, growth factor signaling, and transcription regulation, and increased genes associated with the formation of extracellular matrices such as perineuronal nets (PNNs). PNNs inhibit neuroplasticity and astroglia contribute to the formation, organization, and maintenance of PNNs. To validate our TRAPseq findings, we showed an increase in PNNs following CVS. Degradation of PNNs in the prefrontal cortex of mice exposed to CVS reversed the CVS-induced behavioral phenotype in the forced swim test. These data lend further support to the neuroplasticity hypothesis of depressive behaviors and, in particular, extend this hypothesis beyond neuronal plasticity to include an overall decrease in genes associated with cortical astroglial plasticity following CVS. Further studies will be needed to assess the antidepressant potential of directly targeting astroglial cell function in models of depression.