Broad activation of the Parkin pathway induces synaptic mitochondrial deficits in early tauopathy.

Mitochondrial defects are a hallmark of early pathophysiology in Alzheimer's disease, with pathologically phosphorylated tau reported to induce mitochondrial toxicity. Mitophagy constitutes a key pathway in mitochondrial quality control by which damaged mitochondria are targeted for autophagy. However, few details are known regarding the intersection of mitophagy and pathologies in tauopathy. Here, by applying biochemical and cell biological approaches including time-lapse confocal imaging in live tauopathy neurons, combined with gene rescue experiments via stereotactic injections of adeno-associated virus particles into tauopathy mouse brains, electrophysiological recordings and behavioural tests, we demonstrate for the first time that mitochondrial distribution deficits at presynaptic terminals are an early pathological feature in tauopathy brains. Furthermore, Parkin-mediated mitophagy is extensively activated in tauopathy neurons, which accelerates mitochondrial Rho GTPase 1 (Miro1) turnover and consequently halts Miro1-mediated mitochondrial anterograde movement towards synaptic terminals. As a result, mitochondrial supply at tauopathy synapses is disrupted, impairing synaptic function. Strikingly, increasing Miro1 levels restores the synaptic mitochondrial population by enhancing mitochondrial anterograde movement and thus reverses tauopathy-associated synaptic failure. In tauopathy mouse brains, overexpression of Miro1 markedly elevates synaptic distribution of mitochondria and protects against synaptic damage and neurodegeneration, thereby counteracting impairments in learning and memory as well as synaptic plasticity. Taken together, our study reveals that activation of the Parkin pathway triggers an unexpected effect-depletion of mitochondria from synaptic terminals, a characteristic feature of early tauopathy. We further provide new mechanistic insights into how parkin activation-enhanced Miro1 degradation and impaired mitochondrial anterograde transport drive tauopathy-linked synaptic pathogenesis and establish a foundation for future investigations into new therapeutic strategies to prevent synaptic deterioration in Alzheimer's disease and other tauopathies.

Maternal immune activation with staphylococcal enterotoxin A produces unique behavioral changes in C57BL/6 mouse offspring.

Stimulation of the immune system during pregnancy, known as maternal immune activation (MIA), can cause long-lasting neurobiological and behavioral changes in the offspring. This phenomenon has been implicated in the etiology of developmental psychiatric disorders, such as autism and schizophrenia. Much of this evidence is predicated on animal models using bacterial agents such as LPS and/or viral mimics such as Poly I:C, both of which act through toll-like receptors. However, fewer studies have examined the role of direct activation of maternal T-cells during pregnancy using microbial agents. Bacterial superantigens, such as Staphylococcal Enterotoxin A and B (SEA; SEB), are microbial proteins that activate CD4+ T-cells and cause prominent T-cell proliferation and cytokine production. We injected pregnant and non-pregnant adult female C57BL/6 mice with 200 µg/Kg of SEA, SEB, or 0.9% saline, and measured splenic T-cell-derived cytokine concentrations (viz., IL-2, IFN-γ, IL-6, and IL-4) 2 h later; animals injected with SEA were also measured for splenic concentrations of TNF-α and IL-17A. Half of the injected pregnant animals were brought to term, and their offspring were tested on a series of behavioral tasks starting at six weeks of age (postnatal day 42 [P42]). These tasks included social interaction, the elevated plus maze (EPM), an open field and object recognition (OR) task, prepulse inhibition (PPI) of sensorimotor gating, and the Morris water maze (MWM). Results showed that SEA and SEB induced significant concentrations of all measured cytokines, and in particular IFN-γ, although cytokine responses were greater following SEA exposure. In addition, pregnancy induced an inhibitory effect on cytokine production. Behavioral results showed distinct phenotypes among offspring from SEA- or SEB-injected mothers, very likely due to differences in the magnitude of cytokines generated in response to each toxin. Offspring from SEA-injected mothers displayed modest decreases in social behavior, but increased anxiety, locomotion, interest in a novel object, and short-term spatial memory, while offspring of SEB-injected mothers only exhibited increased anxiety and locomotion. There were no deficits in PPI, which was actually pronounced in SEA and SEB offspring. Overall, the novel use of SEA and SEB as prenatal immune challenges elicited distinct behavioral profiles in the offspring that both mirrors and diverges from previous models of maternal immune activation in important ways. We conclude that superantigen-induced T-cell-mediated maternal immune activation is a valid and valuable model for studying and expanding our understanding of the effects of prenatal immune challenge on neurodevelopmental and behavioral alterations in offspring.

Mitochondrial bioenergetics stimulates autophagy for pathological tau clearance in tauopathy neurons.

Hyperphosphorylation and aggregation of microtubule-associated tau is a pathogenic hallmark of tauopathies and a defining feature of Alzheimer's disease (AD). Pathological tau is targeted by autophagy for clearance, but autophagy dysfunction is indicated in tauopathy. While mitochondrial bioenergetic failure has been shown to precede the development of tau pathology, it is unclear whether energy metabolism deficiency is involved in tauopathy-related autophagy defects. Here, we reveal that stimulation of anaplerotic metabolism restores defective oxidative phosphorylation (OXPHOS) in tauopathy which, strikingly, leads to enhanced autophagy and pronounced tau clearance. OXPHOS-induced autophagy is attributed to increased ATP-dependent phosphatidylethanolamine biosynthesis in mitochondria. Excitingly, early bioenergetic stimulation boosts autophagy activity and reduces tau pathology, thereby counteracting memory impairment in tauopathy mice. Taken together, our study sheds light on a pivotal role of bioenergetic dysfunction in tauopathy-linked autophagy defects and suggests a new therapeutic strategy to prevent toxic tau buildup in AD and other tauopathies.

Neuronal DAMPs exacerbate neurodegeneration via astrocytic RIPK3 signaling.

Astrocyte activation is a common feature of neurodegenerative diseases. However, the ways in which dying neurons influence the activity of astrocytes is poorly understood. Receptor interacting protein kinase-3 (RIPK3) signaling has recently been described as a key regulator of neuroinflammation, but whether this kinase mediates astrocytic responsiveness to neuronal death has not yet been studied. Here, we used the 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine model of Parkinson's disease to show that activation of astrocytic RIPK3 drives dopaminergic cell death and axon damage. Transcriptomic profiling revealed that astrocytic RIPK3 promoted gene expression associated with neuroinflammation and movement disorders, and this coincided with significant engagement of damage-associated molecular pattern signaling. In mechanistic experiments, we showed that factors released from dying neurons signaled through receptor for advanced glycation endproducts to induce astrocytic RIPK3 signaling, which conferred inflammatory and neurotoxic functional activity. These findings highlight a mechanism of neuron-glia crosstalk in which neuronal death perpetuates further neurodegeneration by engaging inflammatory astrocyte activation via RIPK3.

Neuronal DAMPs exacerbate neurodegeneration via astrocytic RIPK3 signaling.

Astrocyte activation is a common feature of neurodegenerative diseases. However, the ways in which dying neurons influence the activity of astrocytes is poorly understood. RIPK3 signaling has recently been described as a key regulator of neuroinflammation, but whether this kinase mediates astrocytic responsiveness to neuronal death has not yet been studied. Here, we used the MPTP model of Parkinson's disease to show that activation of astrocytic RIPK3 drives dopaminergic cell death and axon damage. Transcriptomic profiling revealed that astrocytic RIPK3 promoted gene expression associated with neuroinflammation and movement disorders, and this coincided with significant engagement of DAMP signaling. Using human cell culture systems, we show that factors released from dying neurons signal through RAGE to induce RIPK3-dependent astrocyte activation. These findings highlight a mechanism of neuron-glia crosstalk in which neuronal death perpetuates further neurodegeneration by engaging inflammatory astrocyte activation via RIPK3.

The Effect of Valproic Acid Exposure throughout Development on Microglia Number in the Prefrontal Cortex, Hippocampus and Cerebellum.

Microglia serve as resident immune cells in the brain, responding to insults and pathological developments. They have also been implicated in shaping synaptic development and regulation. The present study examined microglial cell density in a number of brain regions across select postnatal (P) ages along with the effects of valproic acid (VPA) on microglia density. Specifically, C57BL/6JCx3CR1+/GFP mice were examined for microglial cell number changes on P7, P14, P30, and P60 under baseline conditions and following 400 mg/kg VPA or saline. The prefrontal cortex (PFC), hippocampus and cerebellum were observed. Under control conditions, the results showed a shift in the number of microglia in these brain areas throughout development with a peak density in the hippocampus at P14 and an increase in PFC microglial numbers from P15 to P30. Interestingly, VPA treatment enhanced microglial numbers in a region-specific manner. VPA at P7 increased microglial cell number in the hippocampus and cerebellum whereas P14 VPA treatment altered microglial density in the cerebellum only. Cerebellar increases also occurred after VPA at P30, and were attended by an effect of increased numbers in the PFC. Finally, animals treated with VPA at P60 exhibited decreased microglia density in the hippocampus only. These results suggest rapid VPA-induced increases in microglial cell density in a developmentally-regulated fashion which differs across distinct brain areas. Furthermore, in the context of prior reports that early VPA causes excitotoxic damage, the present findings suggest early VPA exposure may provide a model for studying altered microglial responses to early toxicant challenge.

Effects of acute and repeated administration of Staphylococcal enterotoxin A on Morris water maze learning, corticosterone and hippocampal IL-1ß and TNFα.

Staphylococcal enterotoxin A (SEA) is a bacterial superantigen that induces pronounced T cell expansion and cytokine production. In addition, SEA activates the HPA axis and forebrain regions relevant to cognitive functions. Since learning-related cognitive changes have not been assessed in response to SEA, spatial learning in the Morris water maze (MWM) was determined in male C57BL/6J mice subjected to acute or repeated injections of 5µg SEA or Saline. Injections were given 2h prior to 4-5days of hidden platform sessions. Animals were then rested for 1month and given retraining without further injections. In addition, splenic IL-1ß, IL-2 and TNFα, plasma corticosterone, and hippocampal IL-1ß and TNFα were measured after the regimen of treatment used in the behavioral experiments. The results showed no learning impairment following acute or repeated SEA challenge. Moreover, when retested 1month later, and without further injections, the SEA group showed more rapid relearning of the MWM. This suggested that coincidental superantigenic T cell activation and training served to promote long-term improvement in recovery of learning. Furthermore, repeated SEA challenge continued to drive increases in plasma corticosterone, but with a compensatory reduction in hippocampal IL-1ß. However, while hippocampal TNFα was reduced after acute and repeated SEA treatment, this was not statistically significant. In view of the importance of modest glucocorticoid elevations and hippocampal IL-1ß in promoting contextual learning, the data point to the hypothesis that SEA promotes long-term plasticity by restraining disruptive increases in hippocampal IL-1ß, and possibly TNFα, during learning.

Differential sensitivity to endotoxin exposure in young and middle-age mice.

Aging can have a profound effect on the neurobehavioral response to immune activation; aged subjects are predisposed to greater deficits in performance and cognitive function in conjunction with an exaggerated neuroinflammatory response. While increased reactivity to an immune insult has been well characterized in aged subjects, the alterations that may exist by middle-age have not been thoroughly investigated. The present study compared the reactions of young (4-month) and middle-age (12-month) male BALB/c mice to an acute or repeated lipopolysaccharide (LPS) challenge(s). The data suggest that in some respects middle-aged mice are more sensitive to endotoxin exposure, as they show enhanced weight loss, splenic cytokine levels, and c-fos expression in the brain following acute LPS administration compared to younger mice. However, acute LPS exposure led to comparable decreases in locomotor activity in young and middle-aged mice. Following repeated LPS administration both age groups showed diminished behavioral and neural reactions to the final LPS challenge, indicating tolerance development. However, the immune system of the middle-aged mice was still mildly responsive to the final LPS exposure, as splenic levels of IL-1beta were significantly elevated. Collectively, the data suggest that middle-age subjects are more sensitive to an immune insult.

Valproic acid induces nuclear factor erythroid 2-related factor 2 expression in fetal and neonatal brains but not in adult brain: evidence of the gamma-aminobutyric acid-shift hypothesis.

The gamma-aminobutyric acid (GABA)-shift hypothesis proposes that GABA agonist action is excitatory early in development and transitions to an inhibitory role later in life. In experiment 1, the nonspecific GABA agonist, valproic acid (VPA), was administered to pregnant C57BL/6 mice on embryonic day 13. Fetal and maternal brains were harvested 2 h post-VPA exposure and assayed for nuclear factor erythroid 2-related factor 2 (NRF2) and H3 expression through western blot analysis. In experiment 2, VPA was administered to neonatal pups on P14 and adult mice on P60. In both experiments, it was observed that NRF2 expression was increased in fetal and neonatal brains, but not in the adult brain. Because NRF2 expression is activated by oxidative stress, these results imply support of the GABA-shift hypothesis in that VPA may exert its developmental damage in the fetal and neonatal periods through excitotoxicity.

Long-lasting Behavioral and Neuroanatomical Effects of Postnatal Valproic Acid Treatment.

Valproic acid (VPA) administered to mice during the early postnatal period causes social, cognitive, and motor deficits similar to those observed in humans with autism spectrum disorder (ASD). However, previous studies on the effects of early exposure to VPA have largely focused on behavioral deficits occurring before or during the juvenile period of life. Given that ASD is a life-long condition, the present study ought to extend our understanding of the behavioral profile following early postnatal VPA into adulthood. Male mice treated with VPA on postnatal day 14 (P14) displayed increased aggression, decreased avoidance of the open arms in the elevated plus maze, and impaired reversal learning in the Y maze. This may indicate a disinhibited or impulsive phenotype in male, but not female, mice treated with VPA during the second week of postnatal life. Decreased dendritic spine density and dendritic spine morphological abnormalities in the mPFC of VPA-treated mice may be indicative of PFC hypofunction, consistent with the observed behavioral differences. Since these types of long-lasting deficits are not exclusively found in ASD, early life exposure to VPA may reflect dysfunction of a neurobiological domain common to several developmental disorders, including ASD, ADHD, and conduct disorder.

Influence of age on behavioral, immune and endocrine responses to the T-cell superantigen staphylococcal enterotoxin A.

Aged subjects are more vulnerable to administration of the endotoxin lipopolysaccharide, but research on age-associated sensitivity to other immune stimulants has been limited. The current study examined the effects of administering the superantigen, staphylococcal enterotoxin A (SEA), to young (4-month-old) and aged (20-month-old) male C57BL/6J mice on consumption of a novel liquid, cytokine production, corticosterone levels, and expression of central mRNA levels of cytokines and corticotropin-releasing hormone. SEA produced exaggerated hypophagia in aged mice, as they showed decreased consumption that persisted for 24 h. SEA increased hypothalamic mRNA levels of interleukin-1beta in the aged, but not the young, mice 2 h after administration. No differences in cytokine expression were observed 24 h after SEA. Both age groups showed increased plasma corticosterone levels 2 h after SEA administration. However, 24 h after SEA exposure the aged, but not the young, mice showed an augmented corticosterone response to the consumption test. Collectively, these data show that aging may exacerbate the behavioral and neuroinflammatory response to superantigen exposure. Further, the present study suggests that immune activation may result in delayed alterations in stress-induced corticosterone production in aged subjects.

Relationship of varying patterns of cytokine production to the anorexic and neuroendocrine effects of repeated Staphylococcal enterotoxin A exposure.

Staphylococcal enterotoxin A (SEA) is a superantigen that stimulates T cells and induces the production of multiple cytokines. Previous studies have shown that SEA augments gustatory neophobia and activates the hypothalamic-pituitary-adrenal (HPA) axis. This study aimed to determine if the cytokine response, behavioral effects, and HPA axis activation persisted after repeated SEA treatment. Male C57BL/6J mice were given 1-4 intraperitoneal injections of 5 microg SEA, after which food intake, corticosterone, or peripheral cytokines were measured. In a series of experiments, it was found that secondary exposure to SEA two or three days after priming increased corticosterone, but attenuated splenic TNFalpha, while augmenting IL-1beta, IL-2, and IFNgamma. The anorexic response was intact after secondary exposure, but absent after a third injection, which was still able to elevate corticosterone. It is unlikely that IL-1 mediated the persistent effects on corticosterone, since this was increased in groups lacking corticosterone elevations. Similarly, TNFalpha was only modestly elevated under repeated SEA conditions that elevated plasma corticosterone. This attenuation appeared to be inversely related to the levels of IL-10, the production of which incrementally rose with each successive injection. In conclusion, repeated exposure to SEA activates the HPA axis and alters behavior. However, there may be dissociation between the behavioral and endocrine effects of SEA with increased SEA exposure. Furthermore, it is possible that while TNFalpha was previously shown to be important in response to acute SEA-induced HPA axis activation, further exposure to SEA elicits other cytokines that may exert neuromodulatory effects through sensitization and/or synergistic mechanisms.

Effects of urokinase-type plasminogen activator in the acquisition, expression and reinstatement of cocaine-induced conditioned-place preference.

Cocaine and many other psychostimulants strongly induce urokinase-type plasminogen activator (uPA) expression in the mesolimbic dopaminergic pathway, which plays a major role in drug-mediated behavioral plasticity [Bahi A, Boyer F, Gumy C, Kafri T, Dreyer JL. In vivo gene delivery of urokinase-type plasminogen activator with regulatable lentivirus induces behavioral changes in chronic cocaine administration. Eur J Neurosci 2004;20:3473-88; Bahi A, Boyer F, Kafri T, Dreyer JL. Silencing urokinase in the ventral tegmental area in vivo induces changes in cocaine-induced hyperlocomotion. J Neurochem 2006;98:1619-31; Bahi A, Dreyer JL. Overexpression of plasminogen activators in the nucleus accumbens enhances cocaine-, amphetamine- and morphine-induced reward and behavioral sensitization. Genes Brain Behav 2007]. In this study, the role of mesolimbic dopamine (DA) pathways in the development of cocaine reward was examined by conditioned-place preference in rats with bilateral intra-accumbens injections of uPA-expressing lentiviral vectors. We show that overexpression of uPA in the Nac significantly augments cocaine-induced place preference. Furthermore, while this did not affect the ability of preference to be extinguished, reinstatement with a low dose of cocaine produced significantly greater preference to the cocaine-associated context. Once CPP had been established, and the preference extinguished, reinstatement induced by a priming dose of cocaine was facilitated by uPA. Inhibition of this serine protease expression using doxycycline abolished the augmented acquisition produced by overexpression of uPA but not the expression of the cocaine-induced CPP. When uPA is inhibited during the acquisition phase, animals no longer demonstrate place preference for the environment previously paired with cocaine. B428, a specific uPA inhibitor does not affect drug reinstatement after extinction if uPA has been activated during acquisition, a clear indication that uPA is involved in the acquisition phase of conditioned-place preference. Our results suggest that that increased uPA expression with repeated drug exposure produces conditions for enhanced acquisition of cocaine-induced CPP, indicating that cocaine-induced CPP and reinstatement may be dependent on active extracellular uPA.

Strain differences in cuprizone induced demyelination.

BACKGROUND: Multiple sclerosis (MS) is a severe neurological disorder, characterized by demyelination of the central nervous system (CNS), and with a prevalence of greater than 2 million people worldwide. In terms of research in MS pathology, the cuprizone toxicity model is widely used. Here we investigated the contribution of genetic differences in response to cuprizone-induced demyelination in two genetically different mouse strains: CD1 and C57BL/6. RESULTS: We demonstrate that exposure to a diet containing 0.2% cuprizone resulted in less severe demyelination in the midline of the corpus callosum over the fornix in CD1 mice than C57BL/6 mice. With continuous cuprizone feeding, demyelination in CD1 mice was not prominent until after 7 weeks, in contrast to C57BL/6 mice, which showed prominent demyelination after 4 weeks of exposure. Concomitantly, immunohistochemical analysis demonstrated more oligodendrocytes, as well as fewer oligodendrocyte progenitor cells, microglia and astrocytes in cuprizone treated CD1 mice. We also analyzed 4-weeks-cuprizone treated corpus callosum tissue samples and found that cuprizone treated CD1 mice showed a smaller reduction of myelin-associated glycoprotein (MAG) and a smaller increase of Iba1 and NG2. CONCLUSIONS: These observations suggest that CD1 mice are less vulnerable to cuprizone-induced demyelination than C57BL/6 mice and thus genetic background factors appear to influence the susceptibility to cuprizone-induced demyelination.

Neuronal, endocrine, and anorexic responses to the T-cell superantigen staphylococcal enterotoxin A: dependence on tumor necrosis factor-alpha.

Staphylococcal enterotoxin A (SEA) is a microbial superantigen that activates T-lymphocytes and induces production of various cytokines, including tumor necrosis factor-alpha (TNFalpha). Previously, it was shown that SEA activates the hypothalamic-pituitary-adrenal axis and augments gustatory neophobic behaviors. In the present study, it was hypothesized that these effects involve neuronal activation in forebrain regions mediating fear and/or anxiety and are dependent on the production of TNFalpha. Male C57BL/6J mice were given intraperitoneal injections of 10 microg of SEA and 5 microg of lipopolysaccharide (LPS) or saline and perfused 2 h later for histochemical determination of brain c-Fos immunoreactivity (IR). The results showed increased c-Fos IR in the paraventricular nucleus, arcuate nucleus, central nucleus of the amygdala, bed nucleus of the stria terminalis, and lateral septum. Challenge of TNF-/- mice with SEA did not produce a significant increase in brain c-Fos IR, although c-Fos was increased after exposure to a psychogenic stressor (i.e., open field). In additional experiments, the elevated corticosterone response to SEA was abrogated in TNF-/- mice and was shown to be corticotropin-releasing hormone dependent. Finally, the augmented reduction in novel food intake after SEA challenge was attenuated in TNF-/- mice as well as in wild-type mice administered antibody to TNFalpha. In conclusion, challenge with SEA recruits brain regions mediating stress and anxiety responses, an effect that requires endogenous TNFalpha. Whether this is indicative of all T-cell superantigens remains to be determined, although it stands in contrast to other models of neuroimmunomodulation (e.g., LPS) that involve multiple cytokine influences.

Orphanin FQ/nociceptin interactions with the immune system in vivo: gene expression changes in lymphoid organs and regulation of the cytokine response to staphylococcal enterotoxin A.

Orphanin FQ/nociceptin (OFQ/N) is a neuropeptide implicated in immunomodulation. Here, we show that endogenous and exogenous OFQ/N modulated the cytokine response to the bacterial superantigen staphylococcal enterotoxin A (SEA). Specifically, OFQ/N enhanced TNFalpha and IFNgamma transcripts in the spleen when injected prior to SEA challenge. Moreover, mice lacking the OFQ/N precursor gene showed diminished TNFalpha and IFNgamma mRNA induction in the spleen following SEA challenge. In addition, mRNA levels of ppOFQ/N and the OFQ/N receptor, NOP, were altered in thymus and spleen after SEA challenge. Overall, this suggests that the OFQ/N system can influence immune function and is itself influenced by immune stimuli.

Neural and behavioral responses to systemic immunologic stimuli: a consideration of bacterial T cell superantigens.

Immune responses represent a source of systemic stress which impacts the brain and modifies various neuroendocrine and behavioral functions. Therefore, the immune system has been conceived of as a potential contributor to stress-related behavioral abnormalities, such as depression. Much of this knowledge has been gained through research focused largely on the administration of cytokines and/or bacterial endotoxin (eg., LPS), which targets innate immune cells, such as macrophages. However, fewer studies have addressed the effects of T cell activation on central nervous system (CNS) function. The discovery and characterization of bacterial superantigens (SAgs) has introduced an important opportunity for studying how T cell activation influences CNS function. Superantigens target unique variable regions of the beta chain of the mouse and human T cell receptor. This is restricted by the class II molecule of the major histocompatibility complex (MHC), and results in the production of a cytokine cascade that includes interleukin-2 (IL-2), interferon-gamma (IFNgamma), tumor necrosis factor (TNF) and many other cytokines, including IL-6. The best studied SAgs are the staphylococcal enterotoxins, of which staphylococcal enterotoxins A and B (SEA and SEB), have been shown to produce significant changes in behavior and activation of the hypothalamic-pituitary-adrenal (HPA) axis. Importantly, a T cell requirement was necessary to produce these changes. Furthermore, the anorexic or hypophagic effects of SAg challenge in mice appears to be related to anxiety-like processes, since challenge with both SEA or SEB reduces consumption of mainly novel food or food presented in a novel context. In the present paper, these studies are reviewed and related to known alterations in both anxiogenic and anxiolytic neuropeptides. It is suggested that immunologically-induced changes in the brain activate both categories of neuropeptides, thereby sustaining an adaptive state of arousal that promotes appropriate behavioral adjustments during infectious illness.

Effects of staphylococcal enterotoxin A on pituitary-adrenal activation and neophobic behavior in the C57BL/6 mouse.

Bacterial superantigens, such as the staphylococcal enterotoxins (SE), exert a strong capacity for in vivo stimulation of T cell proliferation and cytokine production. Among these superantigens, staphylococcal enterotoxin B (SEB) has been shown to promote anxiety-like properties, possibly mediated by activation of central corticotropin-releasing hormone. In the present study, using male C57BL/6J mice, it was shown that challenge with another prominent superantigen, staphylococcal enterotoxin A (SEA), produced a dose-dependent (0.2-50 microg/mouse) increase in corticosterone and ACTH levels. Interestingly, while the adrenocorticoid response to SEA persisted in cyclosporine A-pretreated mice, it was completely abolished in RAG-1 deficient mice that lack functional T and B lymphocytes. The latter is consistent with the need for cellular interactions involving T cells and B cells (probably in an antigen-presenting capacity) that will initiate events leading to pituitary-adrenal activation by SEA. Since pituitary-adrenal activation typically alters "emotional" reactivity in animals, a final set of experiments assessed behavioral responses to an open field, exposure to a novel object, and a novel appetitive stimulus. These tests revealed a significant augmentation of reactivity to the novel object in SEA-challenged mice, although activity in the open field was not affected. Furthermore, consumption of a novel solution was reduced only if testing involved unfamiliar contextual circumstances. This suggested that anorexic effects per se were not induced by SEA at the dose used, but that attentional mechanisms focused on novelty were enhanced.

Immunological challenge modulates brain orphanin FQ/nociceptin and nociceptive behavior.

Orphanin FQ/Nociceptin (OFQ/N), an endogenous peptide found throughout the central nervous system, has been attributed with a wide range of functions, including modulation of motivational and emotional behavior, but most prominently, facilitation of hyperalgesia. It has also been shown that brain OFQ/N is stimulated during locally-induced peripheral inflammation, a condition well known to increase pain sensitivity. However, few studies have addressed whether specific immunological challenge using T-cell dependent and independent stimuli alters OFQ/N gene activation in the brain. Consequently, male C57BL/6J mice were challenged with 5 microg of lipopolysaccharide (LPS) or a T-cell-activating bacterial superantigen, Staphyloccocal enterotoxin A (SEA), and levels of brain OFQ/N precursor, pNOC, mRNA were analyzed by semi-quantitative RT-PCR. In addition, nociceptive thresholds were examined in immunologically challenged mice using the hotplate test. Initial results on a combined region of the brain containing various limbic components, revealed increased levels of pNOC mRNA in response to SEA challenge, but not to LPS. Further analysis of more discrete brain regions revealed increased pNOC mRNA in the hypothalamus and amygdala in response to SEA. Interestingly, challenge with SEA, but not LPS, significantly reduced hindpaw-lick latency in the hot plate test, although this effect was observed only if the hotplate environment was unfamiliar, suggesting an interaction between immunological stimulation and novelty-induced stress. Since SEA induces various cytokines, including TNF-alpha, these results are consistent with a growing literature documenting the effects of cytokines on nociceptive functions, and a possible involvement of the OFQ/nociceptin system.