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Frontotemporal dementia (FTD) causes progressive personality, behavior and/or language disturbances and represents the second most common form of dementia under the age of 65. Over half of all FTD cases are classified pathologically as frontotemporal lobar degeneration (FTLD) with TAR DNA-binding protein of 43 kDa (TDP-43) pathology (FTLD-TDP). In FTLD-TDP brains, TDP-43 is phosphorylated, C-terminally cleaved, lost from the nucleus and accumulates in the cytoplasm and processes of neurons and glia. However, the contribution of TDP-43 C-terminal fragments (CTFs) to pathogenesis remains poorly understood. Here, we developed transgenic (Tg) mice with forebrain Camk2a-controlled doxycycline-suppressible expression of a TDP-43 CTF (amino acids 208-414, designated 208 TDP-43 CTF), previously identified in FTLD-TDP brains. In these 208 TDP-43 Tg mice, detergent-insoluble 208 TDP-43 CTF was present in a diffuse punctate pattern in neuronal cytoplasm and dendrites without forming large cytoplasmic inclusions. Remarkably, the hippocampus showed progressive neuron loss and astrogliosis in the dentate gyrus (DG). This was accompanied by phosphorylated TDP-43 in the CA1 subfield, and ubiquitin and mitochondria accumulations in the stratum lacunosum moleculare (SLM) layer, without loss of endogenous nuclear TDP-43. Importantly, 208 TDP-43 CTF and phosphorylated TDP-43 were rapidly cleared when CTF expression was suppressed in aged Tg mice, which ameliorated neuron loss in the DG despite persistence of ubiquitin accumulation in the SLM. Our results demonstrate that Camk2a-directed 208 TDP-43 CTF overexpression is sufficient to cause hippocampal pathology and neurodegeneration in vivo, suggesting an active role for TDP-43 CTFs in the pathogenesis of FTLD-TDP and related TDP-43 proteinopathies.

Neuroimmune regulation plays a major role in many facets of human health. In the last 30 years our understanding of this intricate relationship between the brain and immune system has progressed immensely. Pioneering work in the late 1980s and early 1990s began to establish an understanding of the bi-directional communication between these previously considered distinct systems, and its implications in the regulation of mood and cognition (Dantzer and Kelley, 1989; Kelley and Dantzer, 1990; Bluthe et al., 1991). Since then, significant advances in the field of neuroimmunology have been made, and we now know considerably more about the mechanisms responsible for neuroimmune regulation of health and behavior. In this Frontiers research topic Neuroinflammation and Behavior, 10 groups of researchers contributed their expertise to discuss the recent knowledge in the field of neuroimmunology, focusing on the neuroinflammatory mechanisms in affective disorders, early life programming of neuroimmune function, as well as neuroimmune interactions in the aging brain and its associated pathologies.

While chemotherapeutic agents have yielded relative success in the treatment of cancer, patients are often plagued with unwanted and even debilitating side-effects from the treatment which can lead to dose reduction or even cessation of treatment. Common side effects (symptoms) of chemotherapy include (i) cognitive deficiencies such as problems with attention, memory and executive functioning; (ii) fatigue and motivational deficit; and (iii) neuropathy. These symptoms often develop during treatment but can remain even after cessation of chemotherapy, severely impacting long-term quality of life. Little is known about the underlying mechanisms responsible for the development of these behavioral toxicities, however, neuroinflammation is widely considered to be one of the major mechanisms responsible for chemotherapy-induced symptoms. Here, we critically assess what is known in regards to the role of neuroinflammation in chemotherapy-induced symptoms. We also argue that, based on the available evidence, neuroinflammation is unlikely the only mechanism involved in the pathogenesis of chemotherapy-induced behavioral toxicities. We evaluate two other putative candidate mechanisms. To this end we discuss the mediating role of damage-associated molecular patterns (DAMPs) activated in response to chemotherapy-induced cellular damage. We also review the literature with respect to possible alternative mechanisms such as a chemotherapy-induced change in the bioenergetic status of the tissue involving changes in mitochondrial function in relation to chemotherapy-induced behavioral toxicities. Understanding the mechanisms that underlie the emergence of fatigue, neuropathy, and cognitive difficulties is vital to better treatment and long-term survival of cancer patients.

We previously showed that leucine supplementation abrogates lipopolysaccharide (LPS)-induced depression-like behavior by competing with the LAT1-dependent transport of kynurenine into the brain. Since leucine activates mTOR in its target cells the objective of the present series of experiments was to determine whether the antidepressant activity of leucine is mediated by activation of mTOR. Using Western blot we first confirmed that leucine (500 mg/kg IP) activates mTOR in the prefrontal cortex 1 hr after treatment, as measured by phosphorylation of AKT, MAP kinase, and mTOR. We then administered rapamycin (0.75 mg/kg IP) before leucine (500 mg/kg IP) and LPS (0.83 mg/kg IP) to CD-1 mice for behavioral experiments. Leucine abrogated LPS-induced reduction in sucrose preference and increased immobility in the forced swim test. These effects were reversed by rapamycin pretreatment. Because leucine utilization by its target cells requires mTOR-mediated upregulation of LAT1 we examined the possibility that rapamycin blocks the effects of leucine by interfering with this mechanism. Livers and hippocampi of mice treated with rapamycin, leucine and LPS were collected at the end of the experiment and expression of LAT1 was analyzed by qRT-PCR. Administration of LPS and leucine in association increased LAT1 expression in both tissues and this effect was blocked by pretreatment with rapamycin. These findings demonstrate the importance of blood-brain barrier transport mechanisms in inflammation-induced depression. Supported by NIH-NIMH (MH104694) and MDACC.

Chronic pain frequently co-occurs with major depressive disorder but the mechanisms are poorly understood. We investigated the contribution of indoleamine 2,3-dioxygenase-1 (IDO1), a rate-limiting enzyme in the conversion of tryptophan to neurotoxic metabolites, to this comorbidity using the spared nerve injury (SNI) model of neuropathic pain in mice. SNI resulted in unilateral mechanical allodynia, reduced social interaction, and increased immobility in the forced swim test without changes in locomotor activity. These findings indicate SNI-induced pain and comorbid depression-like behavior. These behavioral responses were accompanied by increases in plasma kynurenine/tryptophan ratios and increased expression of Ido1 and Il1b mRNA in the liver. Interestingly, SNI did not induce detectable changes in spinal cord or brain Ido1 mRNA levels. SNI was associated with spinal cord inflammatory activity as evidenced by increased Il1b mRNA expression. The SNI-induced increase of liver Ido1 and Il1b mRNA was abrogated by intrathecal administration of the IL-1 inhibitor IL-1RA. Intrathecal IL-1RA also inhibited both mechanical allodynia and depression-like behavior. We also show that Ido1 is required for the development of depression-like behavior because Ido1 −/− mice do not develop increased immobility in the forced swim test or decreased social exploration in response to SNI. Mechanical allodynia was similar in WT and Ido1 −/− mice. In conclusion, our findings show for the first time that neuropathic pain is associated with an increase of Ido1 in liver, but not brain, downstream of spinal cord IL-1β signaling and that Ido1 mediates comorbid depression. Moreover, comorbidity of neuropathic pain and depression are only partially mediated by a common mechanism because mechanical hyperalgesia develops independently of Ido1 .

Sleep is promoted by numerous pathways in the central nervous system, including factors involved in brain metabolism (e.g., adenosine) and factors involved in immune function (e.g., cytokines). Current mathematical models of sleep regulation do not include any (neuro) immune-related factors and therefore cannot be used to explain or predict effects of inflammation or normal cytokine function on sleep. We have previously developed a receptor-ligand model of the adenosine system, which accurately predicts human sleep patterns, daily changes in extracellular adenosine concentrations in the central nervous system, and cognitive performance under normal or sleep-deprived conditions. Here, we develop an extended mathematical model that includes cytokines that are known to be actively involved in regulating human sleep: IL-1, IL-6, and TNF-alpha. We fit the new model to published data from three conditions: (i) Normal daily variations in cytokine concentrations in healthy individuals. (ii) Changes in cytokine concentrations during and following sleep deprivation in healthy individuals. (iii) Changes in sleep duration and cytokine concentrations during infection. After fitting, the model is used to predict how sleep and cognitive performance would be expected to vary under conditions of sleep restriction and recovery during an active infection. This model provides a novel quantitative framework for studying links between sleep regulation, cognition, and immune function.

Inflammation may contribute to the adverse effects of chemotherapy on cognition and affect, (chemobrain). Here, we tested the hypothesis that the chemoagent paclitaxel induces sustained changes in cognitive and affective-like behavior that correspond with increased inflammation in mice. We treated BALB/c mice with paclitaxel (10 mg/kg i.p.) or vehicle every second day over two weeks. We conducted a battery of tests to examine memory, sickness and affect at 24 h and 14 days after treatment. Paclitaxel significantly reduced memory and increased depression-like behavior as measured by novel object recognition and sucrose preference at both timepoints. These effects were independent of sickness. Analysis of inflammation by ELISA showed no evidence of inflammation at 24 h, suggesting that inflammation may not be responsible. Cytotoxic effects on the brain may be responsible for these immediate behavioral changes. We confirmed that paclitaxel crosses the blood-brain barrier by HPLC, and are performing untargeted metabolomics analyses on brain tissue to examine widespread functional changes in brain metabolism and neurochemistry. Inflammation was apparent at 2 weeks, and we are confirming that inflammation is also elevated in the brain at this timepoint. These findings suggest that inflammation may be responsible for the later paclitaxel-induced cognitive and affective symptoms but cannot be responsible for the immediate effects of paclitaxel on cognition and depression-like behavior.

Phosphorylated and ubiquitinated TAR DNA binding protein 43 (TDP-43) pathology is the neurodegenerative hallmark of the vast majority of amyotrophic lateral sclerosis (ALS) and approximately half of all frontotemporal lobar degeneration (FTLD) cases. An RNA/DNA-binding protein with diverse roles in the nucleus, TDP-43 accumulates in the cytoplasm of affected neurons and glia in disease, indicating that both a loss of normal nuclear function and additional gain of toxicity due to increased cytoplasmic presence likely occur in these TDP-43 proteinopathies. Abundance and splicing of hundreds of transcripts are affected by dysfunction of TDP-43, making identification of the key pathways involved in disease pathogenesis a difficult task. However, endoplasmic reticulum (ER) stress and activation of the unfolded protein response have been described in sporadic ALS patient tissues as well as in cell and animals models of widely-studied superoxide dismutase 1 (SOD1) mutation-linked ALS. These responses to misfolded proteins have more recently been demonstrated in models of TDP-43linked disease, and modulation of ER stress, which is intimately associated with oxidative stress and formation of stress granules in cells, could therefore represent one avenue for potential amelioration of ALS and FTLD. This review summarizes recent findings describing the involvement of ER stress in TDP-43 proteinopathies.

Alexander disease (AxD) is a rare neurodegenerative disorder characterized pathologically by the presence of eosinophilic inclusions known as Rosenthal fibers (RFs) within astrocytes, and is caused by dominant mutations in the coding region of the gene encoding glial fibrillary acidic protein (GFAP). GFAP is the major astrocytic intermediate filament, and in AxD patient brain tissue GFAP is a major component of RFs. TAR DNA binding protein of 43 kDa (TDP-43) is the major pathological protein in almost all cases of the neurodegenerative disease amyotrophic lateral sclerosis (ALS) and ∼50% of frontotemporal lobar degeneration (FTLD), designated as FTLD-TDP. In ALS and FTLD-TDP, TDP-43 becomes insoluble, ubiquitinated, and pathologically phosphorylated and accumulates in cytoplasmic inclusions in both neurons and glia of affected brain and spinal cord regions. Previously, TDP-43 was detected in RFs of human pilocytic astrocytomas; however, involvement of TDP-43 in AxD has not been determined. Here we show that TDP-43 is present in RFs in AxD patient brains, and that insoluble phosphorylated full-length and high molecular weight TDP-43 accumulates in white matter of such brains. Phosphorylated TDP-43 also accumulates in the detergent-insoluble fraction from affected brain regions ofGfapR236H/+knock-in mice, which harbor a GFAP mutation homologous to one that causes AxD in humans, and TDP-43 colocalizes with astrocytic RF pathology inGfapR236H/+mice and transgenic mice overexpressing human wild-type GFAP. These findings suggest common pathogenic mechanisms in ALS, FTLD, and AxD, and this is the first report of TDP-43 involvement in a neurological disorder primarily affecting astrocytes.

Chronic inflammation in physically ill patients is often associated with the development of symptoms of depression. The mechanisms that are responsible for inflammation-associated depression have been elucidated over the last few years. Kynurenine produced from tryptophan in a reaction catabolized by indoleamine 2,3 dioxygenase is transported into the brain where it is metabolized by microglial enzymes into a number of neurotropic compounds including quinolinic acid, an agonist of N-methyl-D-aspartate receptors. Quinolinic acid can synergize with glutamate released by activated microglia. This chain of events opens the possibility to treat inflammation-induced depression using therapies that target the transport of kynurenine through the blood-brain barrier, the production of quinolinic acid and glutamate by activated microglia, or the efflux of glutamate from the brain to the blood.

We have previously discovered a synergistically therapeutic combination of two Toll-like receptor ligands, an oligodeoxynucleotide (ODN) and Pam2CSK4. Aerosolization of these ligands stimulates innate immunity within the lungs to prevent pneumonia from bacterial and viral pathogens. Here we examined the safety and tolerability of this treatment in mice, and characterized the expression of biomarkers of innate immune activation. We found that neutrophils appeared in lung lavage fluid 4 h after treatment, reached a peak at 48 h, and resolved by 7 days. The peak of neutrophil influx was accompanied by a small increase in lung permeability. Despite the abundance of neutrophils in lung lavage fluid, only rare neutrophils were visible histopathologically in the interstitium surrounding bronchi and veins and none were visible in alveolar airspaces. The cytokines interleukin 6 (IL-6), tumour necrosis factor, and Chemokine (C-X-C motif) ligand 2 rose several hundred-fold in lung lavage fluid 4 h after treatment in a dose-dependent and synergistic manner, providing useful biomarkers of lung activation. IL-6 rose fivefold in serum with delayed kinetics compared to its rise in lavage fluid, and might serve as a systemic biomarker of immune activation of the lungs. The dose–response relationship of lavage fluid cytokines was preserved in mice that underwent myeloablative treatment with cytosine arabinoside to model the treatment of hematologic malignancy. There were no overt signs of distress in mice treated with ODN/Pam2CSK4 in doses up to eightfold the therapeutic dose, and no changes in temperature, respiratory rate, or behavioral signs of sickness including sugar water preference, food disappearance, cage exploration or social interaction, though there was a small degree of transient weight loss. We conclude that treatment with aerosolized ODN/Pam2CSK4 is well tolerated in mice, and that innate immune activation of the lungs can be monitored by the measurement of inflammatory cytokines in lung lavage fluid and serum.

As cancer-related symptoms are similar to those of sickness associated with inflammation, it has been proposed that they are also mediated by increased inflammatory cytokines in the brain. However, there is still limited evidence for this relationship. The current study evaluated the alternative possibility that cancer-related symptoms are associated with changes in gene expression indicative of hypoxia signaling and mitochondrial abnormalities using PCR Arrays. Mice were injected or not with heterotopic tonsil epithelial cells transfected with Ras, E6/E7 of HPV16 and were exposed or not to 3 rounds of cisplatin (5.28 mg/kg) and radiotherapy (8 Gy). Tumor-bearing mice showed increased brain and liver inflammatory cytokine mRNA expression that was associated with decreased burrowing. Chemoradiation in non-tumor-bearing mice decreased burrowing but did not induce liver or brain inflammatory cytokine expression. Chemoradiation in tumor-bearing mice reduced burrowing and attenuated liver and brain inflammation, presumably by reducing tumor volume. Both the tumor and its treatment were associated gene expression changes indicative of mitochondrial dysfunction and HIF-1 α activation in liver and brain, with evidence of an additive effect of tumor and chemoradiation on brain mitochondrial dysfunction. Although correlative at this stage, these findings indicate that targeting mitochondrial dysfunction following cancer and cancer therapy may be a strategy for prevention of cancer-related symptoms. Ongoing work is exploring whether this can be achieved without interfering with the tumor response to chemoradiation.

Cu/Zn-superoxide dismutase is misfolded in familial and sporadic amyotrophic lateral sclerosis, but it is not clear how this triggers endoplasmic reticulum (ER) stress or other pathogenic processes. Here, we demonstrate that mutant SOD1 (mSOD1) is predominantly found in the cytoplasm in neuronal cells. Furthermore, we show that mSOD1 inhibits secretory protein transport from the ER to Golgi apparatus. ER-Golgi transport is linked to ER stress, Golgi fragmentation and axonal transport and we also show that inhibition of ER-Golgi trafficking preceded ER stress, Golgi fragmentation, protein aggregation and apoptosis in cells expressing mSOD1. Restoration of ER-Golgi transport by over-expression of coatomer coat protein II subunit Sar1 protected against inclusion formation and apoptosis, thus linking dysfunction in ER-Golgi transport to cellular pathology. These findings thus link several cellular events in amyotrophic lateral sclerosis into a single mechanism occurring early in mSOD1 expressing cells.

Amyotrophic lateral sclerosis (ALS) is a fatal and rapidly progressing neurodegenerative disorder and the majority of ALS is sporadic, where misfolding and aggregation of Cu/Zn-superoxide dismutase (SOD1) is a feature shared with familial mutant-SOD1 cases. ALS is characterized by progressive neurospatial spread of pathology among motor neurons, and recently the transfer of extracellular, aggregated mutant SOD1 between cells was demonstrated in culture. However, there is currently no evidence that uptake of SOD1 into cells initiates neurodegenerative pathways reminiscent of ALS pathology. Similarly, whilst dysfunction to the ER–Golgi compartments is increasingly implicated in the pathogenesis of both sporadic and familial ALS, it remains unclear whether misfolded, wildtype SOD1 triggers ER–Golgi dysfunction. In this study we show that both extracellular, native wildtype and mutant SOD1 are taken up by macropinocytosis into neuronal cells. Hence uptake does not depend on SOD1 mutation or misfolding. We also demonstrate that purified mutant SOD1 added exogenously to neuronal cells inhibits protein transport between the ER–Golgi apparatus, leading to Golgi fragmentation, induction of ER stress and apoptotic cell death. Furthermore, we show that extracellular, aggregated, wildtype SOD1 also induces ER–Golgi pathology similar to mutant SOD1, leading to apoptotic cell death. Hence extracellular misfolded wildtype or mutant SOD1 induce dysfunction to ER–Golgi compartments characteristic of ALS in neuronal cells, implicating extracellular SOD1 in the spread of pathology among motor neurons in both sporadic and familial ALS.

In amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration, TAR DNA binding protein 43 (TDP-43) accumulates in the cytoplasm of affected neurons and glia, where it associates with stress granules (SGs) and forms large inclusions. SGs form in response to cellular stress, including endoplasmic reticulum (ER) stress, which is induced in both familial and sporadic forms of ALS. Here we demonstrate that pharmacological induction of ER stress causes TDP-43 to accumulate in the cytoplasm, where TDP-43 also associates with SGs. Furthermore, treatment with salubrinal, an inhibitor of dephosphorylation of eukaryotic initiation factor 2-α, a key modulator of ER stress, potentiates ER stress-mediated SG formation. Inclusions of C-terminal fragment TDP-43, reminiscent of disease-pathology, form in close association with ER and Golgi compartments, further indicating the involvement of ER dysfunction in TDP-43-associated disease. Consistent with this notion, over-expression of ALS-linked mutant TDP-43, and to a lesser extent wildtype TDP-43, triggers several ER stress pathways in neuroblastoma cells. Similarly, we found an interaction between the ER chaperone protein disulphide isomerase and TDP-43 in transfected cell lysates and in the spinal cords of mutant A315T TDP-43 transgenic mice. This study provides evidence for ER stress as a pathogenic pathway in TDP-43-mediated disease.

Brain inflammation in early life has been proposed to play important roles in the development of anxiety and psychosis-related behaviors in adulthood, behaviors that rely on the integrity of dopamine and/or serotonin systems. Moreover recent behavioral and anatomical evidence suggests involvement of CB1 receptors in the control of emotion and mood. In this study, we determined the effects of neonatal LPS treatment on dopamine, serotonin, and cannabinoid receptor binding in adulthood. Rats were treated with the bacterial endotoxin lipopolysaccharide (LPS) on postnatal day (PND) 3 and 5. Dopamine D1, D2, serotonin 5HT1A, 5HT2A, and serotonin transporter and cannabinoid CB1 receptor binding across several brain regions were measured autoradiographically in adulthood (PND 85). Neonatal LPS treatment caused a significant increase in dopamine D2 in the nucleus accumbens and olfactory tubercle, a decrease in 5HT1A receptor binding in the hippocampus CA1 and ventromedial hypothalamus. A decrease in CB1 receptor binding after neonatal LPS was observed in the amygdala. Neonatal LPS had no significant impact on dopamine D1, serotonin 5HT2A or serotonin transporter binding in any of the brain regions examined. Our results suggest long lasting, region specific effects and differential impact on dopamine, serotonin and cannabinoid receptor systems following neonatal inflammation, that may form the basis for compromised anxiety and psychosis related behaviors.

Depression is a common comorbid condition in substance abuse disorders. In particular, comorbidity between alcoholism and depression occurs with extremely high prevalence in the general population. Early studies investigating this dual diagnosis issue reported alcohol abuse to occur in almost a quarter of patients with affective disorders [1, 2]. Prevalence rises as high as 32 % if other substance abuse disorders are included [2]. These findings made it clear that enormous challenges lie ahead in tracing the individual etiology of substance abuse and depression, let alone their interaction. Clearly, this dilemma was to produce further difficulties in providing adequate and appropriate treatment for sufferers.

article i nfo This article is part of a Special Issue "Neuroendocrine-Immune Axis in Health and Disease." Neonatal lipopolysaccharide (LPS) exposure alters neuroendocrine, immune and behavioural responses in adult rats. Recent findings indicate that neonatal LPS treatment may have a more pronounced effect on the mating behaviours of females compared to males. The current study further explored the impact of neonatal inflammation on reproductive development in the female rat. Wistar rats were administered LPS (0.05 mg/kg, i.p.) or saline (equivolume) on postnatal days (PNDs) 3 and 5. The immediate effect of treat- ment was assessed on plasma corticosterone and tyrosine hydroxylase (TH) phosphorylation in the adrenal medulla. Weight gain and vaginal opening were recorded, and oestrous cyclicity was monitored post-puberty and in late adulthood. Blood and ovaries were collected throughout development to assess HPA and HPG hor- mones and to examine ovarian morphology. Reproductive success in the first (F1) generation and reproduc- tive development in the second (F2) generation were also assessed. Neonatal LPS exposure resulted in increased TH phosphorylation in the neonatal adrenals. LPS treatment increased the corticosterone concen- trations of females as juveniles, adolescents and adults, and reduced FSH in adolescence. Increased catch-up growth was evident in LPS-treated females, prompting earlier onset of puberty. Diminished follicu- lar reserve was observed in neonatally LPS-treated females along with the advanced reproductive senes- cence. While fertility rates were not compromised, higher mortality and morbidity were observed in litters born to LPS-treated mothers. Female offspring of LPS-treated mothers displayed increased corticosterone on PND 14, increased catch-up growth and delayed emergence of the first oestrous cycle. No differences in any of the parameters assessed were observed in F2 males. These data suggest that neonatal immunological challenge has a profound impact on the female reproductive development, via the alteration of metabolic and neuroendocrine factors which regulate sexual maturation. Evidence of altered development in the female, but not male offspring of LPS-treated dams suggests increased susceptibility of females to the deleterious effects of neonatal immunological stress and its possible transferability to a subsequent generation.

Infections during pregnancy and adolescent cannabis use have both been identified as environmental risk factors for schizophrenia. We combined these factors in an animal model and looked at their effects, alone and in combination, on serotonin 5HT1A receptor binding (5HT1AR) binding longitudinally from late adolescence to adulthood. Pregnant rats were exposed to the viral mimic poly I:C on embryonic day 15. Adolescent offspring received daily injections of the cannabinoid HU210 for 14 days starting on postnatal day (PND) 35. Hippocampal and cortical 5HT1AR binding was quantified autoradiographically using [3H]8-OH-DPAT, in late adolescent (PND 55), young adult (PND 65) and adult (PND 90) rats. Descendants of poly I:C treated rats showed significant increases of 15–18% in 5HT1AR in the hippocampus (CA1) compared to controls at all developmental ages. Offspring of poly I:C treated rats exposed to HU210 during adolescence exhibited even greater elevations in 5HT1AR (with increases of 44, 29, and 39% at PNDs 55, 65, and 90). No effect of HU210 alone was observed. Our results suggest a synergistic effect of prenatal infection and adolescent cannabinoid exposure on the integrity of the serotoninergic system in the hippocampus that may provide the neurochemical substrate for abnormal hippocampal-related functions relevant to schizophrenia.

Endoplasmic reticulum (ER) stress is an important pathway to cell death in amyotrophic lateral sclerosis (ALS). We previously demonstrated that ER stress is linked to neurotoxicity associated with formation of inclusions of mutant Cu,Zn-superoxide dismutase 1 (SOD1). Cells bearing mutant inclusions undergo mitochondrial apoptotic signalling. Here, we demonstrate that the BH3-only protein, Bim, is a direct link between ER stress and mitochondrial apoptosis. In the murine neuroblastoma cell line, Neuro2a, bearing mutant SOD1 inclusions, indicators of both ER stress and apoptosis are expressed. Bim knockdown by siRNA significantly reduced nuclear apoptotic features in these inclusion-bearing cells (but did not affect the proportion of cells overall that bear inclusions). Further, both Bax recruitment to mitochondria and cytochrome c redistribution were also decreased under Bim-depletion conditions. However, upregulation of CHOP, a marker of ER stress, was not reduced by Bim knockdown. Significantly, knockdown of CHOP by siRNA reduced the extent of apoptosis in cells bearing mutant SOD1 inclusions. These sequential links between ER stress, CHOP upregulation, and Bim activation of mitochondrial apoptotic signalling indicate a clear pathway to cell death mediated by mutant SOD1.

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease characterised by the progressive loss of motor neurons, leading to paralysis and death within several years of onset. Although protein misfolding is a key feature of ALS, the upstream triggers of disease remain elusive. Recently, endoplasmic reticulum (ER) stress was identified as an early and central feature in ALS disease models as well as in human patient tissues, indicating that ER stress could be an important process in disease pathogenesis. One important chaperone induced by ER stress is protein disulphide isomerase (PDI), which is both upregulated and posttranslationally inhibited by S-nitrosylation in ALS. In this paper, we present evidence from studies of genetics, model organisms, and patient tissues which indicate an active role for PDI and ER stress in ALS disease processes.

Mutations in the gene encoding fused in sarcoma (FUS) are linked to amyotrophic lateral sclerosis (ALS), but the mechanisms by which these mutants trigger neurodegeneration remain unknown. Endoplasmic reticulum (ER) stress is increasingly recognized as an important and early pathway to motor neuron death in ALS. FUS is normally located in the nucleus but in ALS, FUS redistributes to the cytoplasm and forms inclusions. In this study, we investigated whether FUS induces ER stress in a motor neuron like cell line (NSC-34). We demonstrate that ER stress is triggered in cells expressing mutant FUS, and this is closely associated with redistribution of mutant FUS to the cytoplasm. Mutant FUS also colocalized with protein disulfide-isomerase (PDI), an important ER chaperone, in NSC-34 cells and PDI was colocalized with FUS inclusions in human ALS lumbar spinal cords, in both sporadic ALS and mutant FUS-linked familial ALS tissues. These findings implicate ER stress in the pathophysiology of FUS, and provide evidence for common pathogenic pathways in ALS linked to the ER.

Inflammation is associated with activation of the tryptophan-kynurenine metabolism pathway. Circulating kynurenine is transported into the brain at the vascular blood–brain barrier (BBB) by the large neutral amino-acid transporter LAT-1. LAT-1 transports tryptophan and kynurenine. To determine how LAT-1 responds to inflammation, we used mouse microvascular endothelial cells that form tight junctions in vitro and constitutively express the tryptophan degrading enzyme indoleamine 2,3-dioxygenase (IDO) and LAT-1. We first showed that addition of lipopolysaccharide and interferon-γ to the culture significantly increased endothelial cell IDO mRNA and the transport capacity of LAT-1 as measured with tryptophan from the luminal to the abluminal side of an in vitro model of the BBB. Before using leucine as a competitive inhibitor of LAT-1-mediated transport of kynurenine, we determined whether leucine can block the transport of tryptophan across BBB endothelial cells. Whereas leucine had no effect on tryptophan in the untreated BBB, it inhibited a portion of the enhanced transport of tryptophan conferred by pretreatment with lipopolysaccharide and interferon-γ. Consistent with this ability to block LPS-mediated brain uptake of tryptophan, leucine (administered 500 mg/kg IP before and 6 h after lipopolysaccharide) was also effective in abrogating lipopolysaccharide-induced decreases in locomotor activity and sucrose preference. Experiments are under way to assess the effect of leucine on kynurenine transport into the brain both in vivo and in vitro .

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease primarily affecting motor neurons. No previous family history exists in the majority of cases, but mutations in the gene encoding superoxide dismutase 1 (SOD1) cause ∼20% of familial disease. Although the mechanisms

Amyotrophic lateral sclerosis is a rapidly progressing fatal neurodegenerative disease characterized by the presence of protein inclusions within affected motor neurons. Endoplasmic reticulum stress leading to apoptosis was recently recognized to be an important process in the pathogenesis of sporadic human amyotrophic lateral sclerosis as well as in transgenic models of mutant superoxide dismutase 1-linked familial amyotrophic lateral sclerosis. Endoplasmic reticulum stress occurs early in disease, indicating a critical role in pathogenesis, and involves upregulation of an important endoplasmic reticulum chaperone, protein disulphide isomerase. We aimed to investigate the involvement of protein disulphide isomerase in endoplasmic reticulum stress induction, protein aggregation, inclusion formation and toxicity in amyotrophic lateral sclerosis. Motor neuron-like NSC-34 cell lines were transfected with superoxide dismutase 1 and protein disulphide isomerase encoding vectors and small interfering RNA, and examined by immunocytochemistry and immunoblotting. Expression of mutant superoxide dismutase 1 induced endoplasmic reticulum stress, predominantly in cells bearing mutant superoxide dismutase 1 inclusions but also in a proportion of cells expressing mutant superoxide dismutase 1 without visible inclusions. Over-expression of protein disulphide isomerase decreased mutant superoxide dismutase 1 aggregation, inclusion formation, endoplasmic reticulum stress induction and toxicity, whereas small interfering RNA targeting protein disulphide isomerase increased mutant superoxide dismutase 1 inclusion formation, indicating a protective role for protein disulphide isomerase against superoxide dismutase 1 misfolding. Aberrant modification of protein disulphide isomerase by S-nitrosylation of active site cysteine residues has previously been shown as an important process in neurodegeneration in Parkinson's and Alzheimer's disease brain tissue, but has not been described in amyotrophic lateral sclerosis. Using a biotin switch assay, we detected increased levels of S-nitrosylated protein disulphide isomerase in transgenic mutant superoxide dismutase 1 mouse and human sporadic amyotrophic lateral sclerosis spinal cord tissues. Hence, despite upregulation, protein disulphide isomerase is also functionally inactivated in amyotrophic lateral sclerosis, which may prevent its normal protective function and contribute to disease. We also found that a small molecule mimic of the protein disulphide isomerase active site, (+/-)-trans-1,2-bis(mercaptoacetamido)cyclohexane, protected against mutant superoxide dismutase 1 inclusion formation. These studies reveal that endoplasmic reticulum stress is important in the formation of mutant superoxide dismutase 1 inclusions, and protein disulphide isomerase has an important function in ameliorating mutant superoxide dismutase 1 aggregation and toxicity. Functional inhibition of protein disulphide isomerase by S-nitrosylation may contribute to pathophysiology in both mutant superoxide dismutase 1-linked disease and sporadic amyotrophic lateral sclerosis. Protein disulphide isomerase is therefore a novel potential therapeutic target in amyotrophic lateral sclerosis and (+/-)-trans-1,2-bis(mercaptoacetamido)cyclohexane and other molecular mimics of protein disulphide isomerase could be of benefit in amyotrophic lateral sclerosis and other neurodegenerative diseases related to protein misfolding.

The accumulation of ubiquitinated protein inclusions is a hallmark of amyotrophic laterals sclerosis (ALS), a rapidly progressing fatal neurodegenerative disease primarily affecting motor neurons. However, the exact cause of motor neuron death in ALS remains unclear. The unfolded protein response (UPR) is a homeostatic mechanism, which is activated in response to endoplasmic reticulum (ER) stress caused by unfolded or misfolded proteins within the ER lumen. The UPR activates three signalling pathways that lead to an up-regulation of protein chaperones and a block in general protein synthesis. However, chronic UPR activation promotes cell death via apoptosis. Here we review evidence from human patients and experimental models implicating ER-stress induced cell death in ALS. Recently, activation of all three UPR transduction pathways was shown in spinal cords of patients affected by ALS, including up-regulation of protein disulfide isomerase (PDI), an important ER chaperone, and activation of the ER stress-specific apoptotic factor caspase-4. Similarly, expression of mutant superoxide dismutase 1 (SOD1) proteins, which cause 20% of familial ALS cases, leads to activation of the UPR in neuronal cell culture and transgenic mutant SOD1 mice, which are the most widely accepted animal model of disease. Microsomal localisation and SNARE complex-regulated secretion of SOD1 implies entry into the ER-Golgi secretory pathway, and we discuss the disruptive effects of mutant SOD1 on the Golgi apparatus and general protein export. Importantly, the links between ER stress and other pathways implicated in ALS, including autophagy, oxidative stress and the ubiquitin-proteasome system, are becoming increasingly recognised, suggesting that ER stress is a central mechanism in disease. These observations suggest novel potential therapeutic targets for ALS.

The double-hit hypothesis posits that an early life genetic or environmental insult sets up a neural predisposition to psychopathology, which may emerge in the presence of a subsequent insult, or 'second hit' in later life. The current study assessed the effect of neonatal lipopolysaccharide (LPS) exposure on anxiety-like behaviours in the adult Wistar rat. Rats were administered either LPS (Salmonella enterica, serotype enteritidis, 0.05 mg/kg, i.p.) or saline (equivolume) on days 3 and 5 of life (birth=day 1). In adulthood (85 days), subjects were allocated to either "stress" or "no stress" treatment groups. For the "stress" group, subjects were exposed to a three-day stress protocol consisting of a 30 min period of restraint and isolation. The "no stress" group was left unperturbed but were handled during this period to control for handling effects between adult "stress" and "no stress" conditions. All animals then underwent behavioural testing using standardised tests of anxiety-like behaviour, including either the Hide Box/Open Field, Elevated Plus Maze (EPM) or Acoustic Startle Response (ASR). Time and event measures for restraint and isolation, the Hide Box/Open Field and EPM were recorded using automated tracking software. Startle amplitude and habituation across time was measured in the ASR test. Prior to and following behavioural test sessions, peripheral blood was collected to assess serum corticosterone and ACTH levels. Data analysis indicated that LPS-treated animals exposed to stress in adulthood exhibited increased anxiety-like behaviour across all behavioural tests compared to controls. Sexually dimorphic effects were observed with males exhibiting increased anxiety-related behaviours compared to females (p

A number of intracellular proteins that are protective after brain injury are classically thought to exert their effect within the expressing cell. The astrocytic metallothioneins (MT) are one example and are thought to act via intracellular free radical scavenging and heavy metal regulation, and in particular zinc. Indeed, we have previously established that astrocytic MTs are required for successful brain healing. Here we provide evidence for a fundamentally different mode of action relying upon intercellular transfer from astrocytes to neurons, which in turn leads to uptake-dependent axonal regeneration. First, we show that MT can be detected within the extracellular fluid of the injured brain, and that cultured astrocytes are capable of actively secreting MT in a regulatable manner. Second, we identify a receptor, megalin, that mediates MT transport into neurons. Third, we directly demonstrate for the first time the transfer of MT from astrocytes to neurons over a specific time course in vitro. Finally, we show that MT is rapidly internalized via the cell bodies of retinal ganglion cells in vivo and is a powerful promoter of axonal regeneration through the inhibitory environment of the completely severed mature optic nerve. Our work suggests that the protective functions of MT in the central nervous system should be widened from a purely astrocytic focus to include extracellular and intra-neuronal roles. This unsuspected action of MT represents a novel paradigm of astrocyte-neuronal interaction after injury and may have implications for the development of MT-based therapeutic agents.

Metallothionein (MT) expression is rapidly up-regulated following CNS injury, and there is a strong correlation between the presence or absence of MTand improved or impaired (respectively) recovery from such trauma.We now report that a distinct subset of NG2-positive, GFAP-negative glial cells bordering the injury tract express MT following focal injury to the adult rat neocortex. To confirm the ability of these NG2 glial cells to express MT, we have isolated and cultured them and identified that they can express MT following stimulation with zinc. To investigate the functional importance of MT expression by NG2 glial cells, we plated cortical neurons onto these cells and found that expression of MT enhanced the permissivity of NG2 glial cells to neurite outgrowth. Our data suggest that expression of MT by NG2 glial cells may contribute to the overall permissiveness of these cells to axon regeneration.

The health status of an individual in adulthood is proposed to be determined by events occurring in the prenatal and early postnatal period. A common early life event proven to have long lasting effects on the developing fetus is stress, including pain. Exposure of fetal and neonatal infants to repetitive psychological (e.g., maternal stress) or physiological (e.g., pain, infection, and noise) stress during this period is proposed to alter mechanisms involved in the regulation of stress, immunological maturation, pain perception, and cognition. Such changes, which persist into adulthood, may occur via alterations in the development of the hypothalamic-pituitary-adrenal (HPA) axis. This process is typically referred to as ‘perinatal programming’. Ontogenic alterations in the development of the HPA-axis have been related to a number of adult pathologies such as cardiovascular disease, type 2 diabetes, asthma, as well as psychopathologies such as anxiety and depression. Objective: In this review, the effectiveness of complementary and alternative medicine (CAM), such as music, dietary supplements, massage and aromatherapy, in reducing perinatal stress in mothers and infants is examined. An emphasis is placed on these therapies as preventative measures which may be of value to individuals at risk of developing disease profiles associated with the consequences of adverse perinatal programming. The widening interest in perinatal programming and CAM suggests the potential for CAM to become a valuable tool in offsetting negative adult health outcomes resulting from perinatal programming associated with adverse gestational early life environments.

Inflammation activates the tryptophan-kynurenine metabolism pathway. Circulating kynurenine is transported into the brain at the vascular blood–brain barrier (BBB) by the large amino-acid transporter LAT-1, where it is further metabolized into neurotoxic metabolites responsible for depressive symptoms. LAT-1 transports tryptophan, kynurenine and the amino acid leucine. We sought to competitively inhibit kynurenine blood-to-brain transport with leucine in order to attenuate lipopolysaccharide-induced depression-like behavior in CD-1 mice. Given that the concentration of circulating kynurenine is 100 fold less than kynurenine, we hypothesized that leucine should inhibit kynurenine more than tryptophan transport. We had already demonstrated that inflammation increases luminal-to-abluminal transport of tryptophan using an in vitro model of BBB transport. Here we show that lipopolysaccharide-treated mice (0.83 mg/kg, ip) develop depression-like behavior at 24 h (reduced sucrose preference and increased forced swim test immobility; p p p p

The RNA/DNA-binding protein, TDP-43, is the key component of ubiquitinated inclusions characteristic of amyotrophic lateral sclerosis (ALS) and the majority of frontotemporal lobar degeneration (FTLD-TDP) referred to collectively as TDP-43 proteinopathies. To further elucidate mechanisms of pathological TDP-43 processing and identify TDP-43 epitopes that could be useful as potential biomarkers of TDP-43 proteinopathies, we developed a panel of novel monoclonal antibodies (MAbs) directed at regions extending across the length of TDP-43. Here, we confirm previous observations that there is no or minimal accumulation of TDP-43 N-terminal domains in neocortical inclusions in human TDP-43 proteinopathy tissues and we identify a subset of these MAbs that are specific for human versus mouse TDP-43. Notably, one of these MAbs recognized an epitope that preferentially detected pathological TDP-43 inclusions with negligible reactivity for normal nuclear TDP-43 resembling anti-phospho-TDP-43 specific antibodies that only bind pathological TDP-43. Hence, we infer that this new MAb recognizes a phosphorylation independent but disease-specific pathologic conformation in abnormal TDP-43. These data suggest that the novel MAbs reported here will be useful for patient-oriented research as well as for studies of animal and cell-based models of TDP-43 proteinopathies including ALS and FTLD-TDP.

We have demonstrated that lipopolysaccharide (LPS) induces depressive-like behavior by activating indoleamine 2,3 dioxygenase (IDO), which degrades tryptophan along the kynurenine pathway. Using LC-MS analysis of kynurenine metabolites in the brain and plasma of mice injected with LPS (1 mg/kg, intraperitoneally), we demonstrate that LPS selectively activates the kynunrenine monooxygenase pathway that results in the production of the N-methyl- d -aspartate (NMDA) receptor agonist quinolinic acid. We also show that administration of the NMDA receptor antagonist ketamine (6 mg/kg, intraperitoneally) immediately prior to LPS (0.83 mg/kg, intraperitoneally) in C57Bl/6J mice abrogates the onset of LPS-induced depressive-like behavior in the forced swim and sucrose preference tests but fails to influence LPS-induced sickness. Ketamine did not affect hepatic and brain LPS-induced cytokine expression, IDO activation, and brain-derived neurotrophic factor (BDNF) transcripts. The antidepressant-like effect of ketamine independent of any effect on LPS-induced sickness was confirmed when ketamine was administered 10 h after LPS instead of immediately beforehand. Ketamine had no effect when administered 24 h prior to LPS. Confirmation of ketamine’s ability to abrogate LPS-induced depressive-like behavior by antagonizing the NMDA receptor and enhancing AMPA-mediated glutamatergic neurotransmission was obtained by pretreating mice with the AMPA receptor antagonist NBQX (10 mg/kg intraperitoneally). NBQX pretreatment restored LPS-induced decreased sucrose preference. These findings suggest that LPS-induced depressive-like behavior is mediated by NMDA receptor activation as a consequence of formation of quinolinic acid.

Ketamine, an NMDA receptor antagonist, has been shown to rapidly treat depressive-like behavior in clinical and pre-clinical studies. To date, its efficacy in treating inflammation-induced depression has not been reported. Here we demonstrate that peripheral administration of ketamine (6 mg/kg, i.p.) immediately prior to peripheral administration of lipopolysaccharide (LPS) (0.83 mg/kg, i.p.) in C57Bl/6J mice abrogates LPS-induced depressive-like behavior. LPS induced a reduction in sucrose preference and increase in immobility in the forced swim test (p NIH R01 MH079829 to R.D. and NIH R01 AG-029573 to K.W.K.

Background : Inflammation associated with cancer and induced by cancer therapy is associated with clinical signs of sickness behavior that can culminate in the development of symptoms of depression. Intraperitoneal administration of lipopolysaccharide to mice induces depressive-like behavior. Lipopolysaccharide-induced depressive-like behavior is mediated by activation of indoleamine 2,3-dioxygenase (IDO) that degrades tryptophan along the kynurenine pathway and produces kynurenine metabolites such as quinolinic acid that acts as a N-Methyl-D-aspartate (NMDA) receptor agonist. The present study was carried out to determine whether the NMDA receptor antagonist ketamine alleviates lipopolysaccharide-induced depressive-like behavior. Methods : Mice were injected intraperitoneally with ketamine or saline immediately before administration of the cytokine inducer lipopolysaccharide or saline via the same route. Their behavior and bodyweight were monitored up to 28 h post-injection. Depressive-like behavior was measured by increased immobility in the forced swim test and decreased sucrose preference for a sucrose solution. Brain, liver and plasma were collected 6 h and 28 h after treatment to measure biomarkers of inflammation. Results : Lipopolysaccharide induced the expression of cytokine, IDO, tryptophan 2,3-dioxygenase (TDO) and heme oxygenase-1 at the periphery and in the brain. This effect was not altered by ketamine pretreatment. Ketamine blocked the development of depressive-like behavior but had no effect on sickness behavior measured by body weight loss, reduced food intake and decreased motor activity. Conclusions : These data indicate that ketamine is able to abrogate inflammation-induced depressive-like behavior by antagonising the activating effects of kynurenine metabolites on NMDA receptors.

11. Postnatal exposure to a bacterial mimetic increases microglial activation and histone H3 acetylation in rats L. Sominsky , A.K. Walker , L.K. Ong , F.R. Walker , D.M. Hodgson a a Laboratory of Neuroimmunology, School of Psychology, Faculty of Science and IT, The University of Newcastle, Callaghan, NSW 2308, Australia b School of Biomedical Sciences & Pharmacy, The University of Newcastle, Australia c Priority Research Centre for Brain and Mental Health Research, University of Newcastle, Australia Hunter Medical Research Institute, Newcastle, Australia