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Brain energy deficit has been a suggested cause of Huntington disease (HD), but ATP depletion has not reliably been shown in preclinical models, possibly because of the immediate post-mortem changes in cellular energy metabolism. To examine a potential role of a low energy state in HD, we measured, for the first time in a neurodegenerative model, brain levels of high energy phosphates using microwave fixation, which instantaneously inactivates brain enzymatic activities and preserves in vivo levels of analytes. We studied HD transgenic R6/2 mice at ages 4, 8, and 12 weeks. We found significantly increased creatine and phosphocreatine, present as early as 4 weeks for phosphocreatine, preceding motor system deficits and decreased ATP levels in striatum, hippocampus, and frontal cortex of R6/2 mice. ATP and phosphocreatine concentrations were inversely correlated with the number of CAG repeats. Conversely, in mice injected with 3-nitroproprionic acid, an acute model of brain energy deficit, both ATP and phosphocreatine were significantly reduced. Increased creatine and phosphocreatine in R6/2 mice was associated with decreased guanidinoacetate N-methyltransferase and creatine kinase, both at the protein and RNA levels, and increased phosphorylated AMP-dependent protein kinase (pAMPK) over AMPK ratio. In addition, in 4-month-old knock-in Hdh(Q111/+) mice, the earliest metabolic alterations consisted of increased phosphocreatine in the frontal cortex and increased the pAMPK/AMPK ratio. Altogether, this study provides the first direct evidence of chronic alteration in homeostasis of high energy phosphates in HD models in the earliest stages of the disease, indicating possible reduced utilization of the brain phosphocreatine pool.

Damage to the CNS results in a complex series of molecular and cellular changes involving the affected targets and the ensuing glial reaction. The initial gene expression events that underlie these cellular responses may serve as early biomarkers of neurotoxicity. Here, we examined gene expression profiles during the initial phase of hippocampal damage resulting from systemic exposure of rats to the organometallic neurotoxicant, trimethyltin (TMT, 8.0 mg/kg, i.p.). Using TMT as a neurodegeneration tool confers several advantages for evaluating molecular events associated with neural damage: 1) regional and cellular targets and time course of damage are known, 2) the blood-brain barrier is not compromised, which limits the contribution of blood-borne factors, e.g. immune, to neural injury responses and 3) known protein and mRNA signatures of TMT-induced neurotoxicity can be used as positive controls to validate novel expression events associated with exposure to this neurotoxicant. Using Affymetrix Gene Chip® to assess gene expression after TMT, combined with Ingenuity Pathway Analysis®, we observed changes consistent for genes known to be affected in hippocampus, while corresponding changes were not detected in cerebellum, a non-target region. In agreement with previous observations, limited changes in expression of inflammation-related genes were observed. Correlated expression profiles were found after exposure to TMT, including changes in gene ontologies associated with neurological disease, cellular assembly and maintenance, as well as signaling pathways associated with cellular stress, energy metabolism and glial activation. Selected gene changes were confirmed from each category by q-RT-PCR and immunoblot analysis. The canonical relationships identified implicate molecular pathways and processes relevant to detection of early stages of hippocampal damage in the TMT model. These observations provide new insight into early events associated with neuronal degeneration and associated glial activation that may serve as the basis for discovery and development of biomarkers of neurotoxicity.

A novel class of N-substituted tetrahydropyridine derivatives was found to have multiple kinetic mechanisms of monoamine oxidase A inhibition. Eleven structurally similar tetrahydropyridine derivatives were synthesized and evaluated as inhibitors of MAO-A and MAO-B. The most potent MAO-A inhibitor in the series, 2,4-dichlorophenoxypropyl analog 12, displayed time-dependent mixed noncompetitive inhibition. The inhibition was reversed by dialysis, indicating reversible enzyme inhibition. Evidence that the slow-binding inhibition of MAO-A with 12 involves a covalent bond was gained from stabilizing a covalent reversible intermediate product by reduction with sodium borohydride. The reduced enzyme complex was not reversible by dialysis. The results are consistent with slowly reversible, mechanism-based inhibition. Two tetrahydropyridine analogs that selectively inhibited MAO-A were characterized by kinetic mechanisms differing from the kinetic mechanism of 12. As reversible inhibitors of MAO-A, tetrahydropyridine analogs are at low risk of having an adverse effect of tyramine-induced hypertension.

Gestational lead exposure (GLE) produces supernormal scotopic electroretinograms (ERG) in children, monkeys and rats, and a novel retinal phenotype characterized by an increased number of rod photoreceptors and bipolar cells in adult mice and rats. Since the loss of dopaminergic amacrine cells (DA ACs) in GLE monkeys and rats contributes to supernormal ERGs, the retinal DA system was analyzed in mice following GLE. C57BL/6 female mice were exposed to low (27 ppm), moderate (55 ppm) or high (109 ppm) lead throughout gestation and until postnatal day 10 (PN10). Blood [Pb] in control, low-, moderate- and high-dose GLE was ≤ 1, ≤ 10, ~25 and ~40 μg/dL, respectively, on PN10 and by PN30 all were ≤ 1 μg/dL. At PN60, confocal-stereology studies used vertical sections and wholemounts to characterize tyrosine hydroxylase (TH) expression and the number of DA and other ACs. GLE dose-dependently and selectively decreased the number of TH-immunoreactive (IR) DA ACs and their synaptic plexus without affecting GABAergic, glycinergic or cholinergic ACs. Immunoblots and confocal revealed dose-dependent decreases in retinal TH protein expression and content, although monoamine oxidase-A protein and gene expression were unchanged. High-pressure liquid chromatography showed that GLE dose-dependently decreased retinal DA content, its metabolites and DA utilization/release. The mechanism of DA selective vulnerability is unknown. However, a GLE-induced loss/dysfunction of DA ACs during development could increase the number of rods and bipolar cells since DA helps regulate neuronal proliferation, whereas during adulthood it could produce ERG supernormality as well as altered circadian rhythms, dark/light adaptation and spatial contrast sensitivity.

The neurological effects of organophosphate (OP) pesticides, commonly used on foods and in households, are an important public health concern. Furthermore, subclinical exposure to combinations of organophosphates is implicated in Gulf War illness. Here, we characterized the effects of the broadly used insecticide chlorpyrifos (CPF) on dopamine and glutamatergic neurotransmission effectors in corticostriatal motor/reward circuitry. CPF potentiated protein kinase A (PKA)-dependent phosphorylation of the striatal protein dopamine- and cAMP-regulated phosphoprotein of M(r) 32 kDa (DARPP-32) and the glutamate receptor 1 (GluR1) subunit of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors in mouse brain slices. It also increased GluR1 phosphorylation by PKA when administered systemically. This correlated with enhanced glutamate release from cortical projections in rat striatum. Similar effects were induced by the sarin congener, diisopropyl fluorophosphate, alone or in combination with the putative neuroprotectant, pyridostigmine bromide and the pesticide N,N-diethyl-meta-toluamide (DEET). This combination, meant to mimic the neurotoxicant exposure encountered by veterans of the 1991 Persian Gulf War, also induced hyperphosphorylation of the neurofibrillary tangle-associated protein tau. Diisopropyl fluorophosphate and pyrodostigmine bromide, alone or in combination, also increased the aberrant activity of the protein kinase, Cdk5, as indicated by conversion of its activating cofactor p35 to p25. Thus, consistent with recent findings in humans and animals, organophosphate exposure causes dysregulation in the motor/reward circuitry and invokes mechanisms associated with neurological disorders and neurodegeneration.

Neuroinflammation is associated with glial activation following a variety of brain injuries, including stroke. While activation of perilesional astrocytes and microglia following ischemic brain injury is well documented, the influence of age on these cellular responses after stroke is unclear. This study investigated the influence of advanced age on neuronal degeneration, neuroinflammation, and glial activation in female Sprague-Dawley rats after reversible embolic occlusion of the middle cerebral artery (MCAO). Results indicate that in comparison to young adult rats (3 months), aged rats (18 months) showed enhanced neuronal degeneration, altered microglial response, and a markedly increased expression of proinflammatory cytokines/chemokines following MCAO. In addition, the time-course for activation of signal transducers and activators of transcription 3 (STAT3), the signaling mechanism that regulates astrocyte reactivity, was truncated in the aged rats after MCAO. Moreover, the expression of suppressor of cytokine signaling 3 (SOCS3), which is associated with termination of astrogliosis, was enhanced as a function of age after MCAO. These findings are suggestive of an enhanced proinflammatory response and a truncated astroglial response as a function of advanced age following MCAO. These data provide further evidence of the prominent role played by age in the molecular and cellular responses to ischemic stroke and suggest that astrocytes may represent targets for future therapies aimed at improving stroke outcome.

Two distinct families of small molecules were discovered as novel alpha7 nicotinic acetylcholine receptor (nAChR) antagonists by pharmacophore-based virtual screening. These novel antagonists exhibited selectivity for the neuronal alpha7 subtype over other nAChRs and good brain penetration. Neuroprotection was demonstrated by representative compounds 7i and 8 in a mouse seizure-like behavior model induced by the nerve agent diisopropylfluorophosphate (DFP). These novel nAChR antagonists have potential use as antidote for organophosphorus nerve agent intoxication.

Organophosphorus (OP) compounds cause toxic symptoms, including convulsions, coma, and death, as the result of irreversible inhibition of acetylcholinesterase (AChE). The development of effective treatments to block these effects and attenuate long-term cognitive and motor disabilities that result from OP intoxication is hampered by a limited understanding of the CNS pathways responsible for these actions. We employed a candidate method (called CNSProfile™) to identify changes in the phosphorylation state of key neuronal phosphoproteins evoked by the OP compound, diisopropyl fluorophosphate (DFP). Focused microwave fixation was used to preserve the phosphorylation state of phosphoproteins in brains of DFP-treated mice; hippocampus and striatum were analyzed by immunoblotting with a panel of phospho-specific antibodies. DFP exposure elicited comparable effects on phosphorylation of brain phosphoproteins in both C57BL/6 and FVB mice. DFP treatment significantly altered phosphorylation at regulatory residues on glutamate receptors, including Serine897 (S897) of the NR1 NMDA receptor. NR1 phosphorylation was bi-directionally regulated after DFP in striatum versus hippocampus. NR1 phosphorylation was reduced in striatum, but elevated in hippocampus, compared with controls. DARPP-32 phosphorylation in striatum was selectively increased at the Cdk5 kinase substrate, Threonine75 (T75). Phencynonate hydrochloride, a muscarinic cholinergic antagonist, prevented seizure-like behaviors and the observed changes in phosphorylation induced by DFP. The data reveal region-specific effects of nerve agent exposure on intracellular signaling pathways that correlate with seizure-like behavior and which are reversed by the muscarinic receptor blockade. This approach identifies specific targets for nerve agents, including substrates for Cdk5 kinase, which may be the basis for new anti-convulsant therapies.

Serious questions have been raised by occupational health investigators regarding a possible causal association between neurological effects in welders and the presence of manganese (Mn) in welding fume. Male Sprague-Dawley rats were exposed by inhalation to 40 mg/m(3) of gas metal arc-mild steel (MS) welding fume for 3 h/day for 10 days. Generated fume was collected in the animal chamber during exposure, and particle size, composition, and morphology were characterized. At 1 day after the last exposure, metal deposition in different organ systems and neurological responses in dopaminergic brain regions were assessed in exposed animals. The welding particles were composed primarily of a complex of iron (Fe) and Mn and were arranged as chain-like aggregates with a significant number of particles in the nanometer size range. Mn was observed to translocate from the lungs to the kidney and specific brain regions (olfactory bulb, cortex, and cerebellum) after MS fume inhalation. In terms of neurological responses, short-term MS fume inhalation induced significant elevations in divalent metal ion transporter 1 (Dmt1) expression in striatum and midbrain and significant increases in expression of proinflammatory chemokines (Ccl2, Cxcl2) and cytokines (IL1beta, TNFalpha) in striatum. In addition, mRNA and protein expression of glial fibrillary acidic protein (GFAP) was significantly increased in striatum after MS fume exposure. However, the 10-day MS welding fume inhalation did not cause any changes in dopamine and its metabolites or GABA in dopaminergic brain regions nor did it produce overt neural cell damage as assessed by histopathology. In summary, short-term MS welding fume exposure led to translocation of Mn to specific brain regions and induced subtle changes in cell markers of neuroinflammatory and astrogliosis. The neurofunctional significance of these findings currently is being investigated in longer, more chronic welding fume exposure studies.

Neuroinflammation is a hot topic in contemporary neuroscience. A relatively new open-access journal, the Journal of Neuroinflammation, focuses on this field. As another example, abstracts to the 2007 Annual Meeting of the Society for Neuroscience could be submitted in several subcategories of neuroinflammation, a strong signal of growth in this research area. While it is becoming clear that activation of microglia and astroglia and the attendant expression of proinflammatory cytokines and chemokines often are associated with disease-, trauma-, and toxicant-induced damage to the CNS, it is by no means clear that a cause-and-effect relationship exists between the presence of a neuroinflammatory process and neural damage. We have explored this issue with two models of dopaminergic neurotoxicity. We used a single low-dose regimen of MPTP or METH, a paradigm that causes selective degeneration of striatal dopaminergic nerve terminals without affecting the cell body in the substantia nigra. Both compounds increased the expression of the microglia-associated factors, Il-1alpha, Il6, Ccl2, and Tnf-alpha, and also elicited morphologic evidence of microglial activation prior to induction of astrogliosis. Pharmacologic antagonism of MPTP and METH neurotoxicity prevented these proinflammatory responses, findings suggestive of a link between neuroinflammation and the observed neurotoxic outcomes. Nevertheless, when we used minocycline to suppress the expression of all these mediators, with the exception of Tnf-alpha, we failed to see neuroprotection. Likewise, when we examined the effects of MPTP or METH in transgenic mice lacking Il6, Ccl2, or Tnfr1/2 genes, deficiency of either Il6 or Ccl2 did not alter neurotoxicity, whereas deficiency in Tnfr1/2 was neuroprotective. Although these observations pointed to a role of the proinflammatory cytokine, TNF-alpha, in the neurotoxic effects of MPTP and METH, other observations did not support this supposition. For example, activation of NF-kappaB or induction of iNOS, known components of inflammatory responses and free radical formation, were not observed. Moreover, immunosuppressive regimens of glucocorticoids failed to suppress TNF-alpha or attenuate neurotoxicity. Taken together, our observations suggest that MPTP and METH neurotoxicity are associated with the elaboration of a "neuroinflammatory" response, yet this response lacks key features of inflammation and, with the exception of TNF-alpha, neurotoxicity appears to be the cause rather than the consequence of proinflammatory signals.

How early-life events "set the stage" for adult disease has emerged as a research focus. Historically, the epidemiology of disease risk factors has centered on adult life, with little scrutiny of early-life events. Here we review the concept that events in early life may contribute to late-life neurodegenerative disease development, with a focus on Parkinson disease (PD) and Alzheimer disease (AD). Suspect events in early life include infections, stress, poor nutrition, and environmental factors such as chemical and pesticide exposure. Adiposity appears to contribute to both PD and AD; and because early-life events contribute to the development of obesity, linkages may exist between early determinants of obesity and the subsequent development of these neurologic diseases. Many now suggest a life-course approach for determining the relative contributions of genetic and environmental factors in any chronic disease. This requires determining when during the life course that a given exposure has its greatest effect and how exposures may accumulate over the life span. The data for PD and AD suggest that a number of insults occurring early in life may lead or contribute to these diseases. More definitive knowledge of the key risk factors involved will be needed to implement intervention and preventative strategies early in life to dampen or prevent any adverse late-life outcomes.

The adverse effects of low-level developmental lead exposure [blood lead (BPb) ≤ 10 μg/dL] on cognitive, auditory, and visual-motor function are well documented (Canfield et al. 2003; Osman et al. 1999; Rothenberg et al. 2000; Wasserman et al. 2000), but only two studies have examined the impact of low-level developmental Pb exposure on retinal and visual function in children (Altmann et al. 1998; Rothenberg et al. 2002). This lack of research is despite findings that retinal and visual cortical structural–functional abnormalities occur in animals after moderate-level (BPb 11–39 μg/dL) and high-level (BPb ≥ 40 μg/dL) developmental Pb exposure (reviewed by Fox and Boyes 2007; Otto and Fox 1993). Persistent rod photoreceptor-mediated (scotopic) electroretinographic (ERG) and behavioral deficits occur in monkeys, rats, and mice after moderate-level and high-level post-natal Pb exposure (PLE) (Fox and Boyes 2007). The scotopic ERG alterations are characterized by decreases in a-wave and b-wave ERG amplitude (subnormality), sensitivity, and temporal resolution. Similar ERG changes occurred in Pb-exposed workers and isolated retinas exposed to Pb2+ (Otto and Fox 1993). In contrast, adult monkeys with high-level lifetime Pb exposure exhibited increased ERG b-wave amplitudes (supernormality) (Lilienthal et al. 1994). Moreover, our prospective epidemiologic study of children with lifetime low-level and moderate-level Pb exposure revealed that only gestational Pb exposure (GLE) showed a significant dose-dependent relationship with supernormal ERG a-waves and b-waves and increased b-wave sensitivity with no change in implicit times (Rothenberg et al. 2002). In experimental animals, supernormal scotopic ERGs are observed after the loss of retinal dopamine (DA) or zinc. For example, the destruction of dopaminergic amacrine cells by the neurotoxin 6-hydroxydopamine produced supernormal scotopic ERGs with normal implicit times (Olivier et al. 1986; Skrandies and Wassle 1988). Similarly, animals administered nonselective dopamine receptor antagonists had supernormal scotopic b-waves with normal implicit times (Jagadeesh et al. 1980; Schneider and Zrenner 1991). A persistent decrease in retinal tyrosine hydroxylase immunoreactivity after lifetime Pb exposure in monkeys suggests that a loss of DA produced supernormal ERGs (Kohler et al. 1997; Lilienthal et al. 1994). Chelation of retinal Zn produced supernormal scotopic ERGs and increased b-wave sensitivity (Redenti and Chappell 2003). Although no published reports have shown that scotopic ERG supernormality results from an increased number of rods, the log scotopic b-wave threshold and amplitude are linearly related to the number of rod photoreceptors and their rhodopsin content per eye (Dowling 1960). Therefore, we hypothesized that GLE produced supernormal scotopic ERGs by one of these three relatively independent mechanisms: an increased number of rod photoreceptors, decreased retinal DA metabolism, and/or decreased retinal Zn concentrations. To determine the sites and mechanisms of the GLE-induced supernormal scotopic ERGs, we developed a new dose–response model of GLE. Then, we conducted scotopic ERG studies and retinal histologic, morphometric, and neurochemical experiments in adult offspring. Low-level and moderate-level GLE produced supernormal scotopic ERGs, an increased neurogenesis of rods and rod bipolar cells (BCs), and decreased DA synthesis and utilization/release in the absence of retinal injury. GLE produced inverted U-shaped dose–response curves as the high-level GLE dose produced rod cell loss and thus ERG subnormality.

Neuropathic pain is a persistent pain condition that develops secondary to nerve injury. The two most common types of peripheral neuropathic pain are post-herpetic neuralgia (PHN) and painful diabetic neuropathy (PDN). Amitriptyline, nortriptyline, desipramine and imipramine are TCAs that have been shown to be effective for the symptomatic relief of PHN and PDN. Serotonin noradrenaline reuptake inhibitors (SNRIs) such as venlafaxine and duloxetine have been shown to be very promising for the treatment of PDN with fewer adverse effects than TCAs. Selective serotonin reuptake inhibitors (SSRIs) were shown in a number of studies to have some efficacy in relieving PDN-related pain, yet other studies of the SSRIs have demonstrated conflicting outcomes. Most of the older antiepileptic studies were performed in patients with PDN; consequently, little is known about the efficacy of these drugs in patients with PHN. Carbamazepine, phenytoin and valproic acid were shown to be effective in ameliorating PDN-related pain. Other antiepileptic agents, including lamotrigine, oxcarbazepine and topiramate, have demonstrated some beneficial effects for the treatment of PDN, although they were also found to be ineffective in some PDN studies. alpha2delta Ligands such as gabapentin and pregabalin have been proven to be effective for the treatment of PHN and PDN in a number of large placebo-controlled trials. These drugs are useful not only in relieving pain but also in improving quality of life. Although the use of opioids for the treatment of neuropathic pain is controversial, a number of studies support the efficacy and safety of opioids in the treatment of neuropathic pain. Of these, oxycodone and tramadol have been shown to be superior to placebo for the treatment of PHN and PDN. A number of small studies have shown that dextromethorphan was effective in patients with PDN but not in patients with PHN. Topical agents such as lidocaine 5% patches and topical capsaicin are useful in ameliorating pain in patients with PHN but these agents are unsatisfactory for use as a sole agent. Although a number of drug treatments are available for the symptomatic relief of neuropathic pain symptoms, these agents do not provide satisfactory relief in all patients. For these patients, other treatment alternatives such as combination drug therapy that produces pain relief via distinctly different mechanisms may be successful. The purpose of this review is to compare the efficacy and limitations of currently available pharmacological treatments for the symptomatic relief of PHN and PDN, and to discuss the potential of combination therapy in PHN and PDN.

Glutaric acidemia type I (GA-I) is an inherited disorder of lysine and tryptophan metabolism presenting with striatal lesions anatomically and symptomatically similar to Huntington disease. Affected children commonly suffer acute brain injury in the context of a catabolic state associated with nonspecific illness. The mechanisms underlying injury and age-dependent susceptibility have been unknown, and lack of a diagnostic marker heralding brain injury has impeded intervention efforts. Using a mouse model of GA-I, we show that pathologic events began in the neuronal compartment while enhanced lysine accumulation in the immature brain allowed increased glutaric acid production resulting in age-dependent injury. Glutamate and GABA depletion correlated with brain glutaric acid accumulation and could be monitored in vivo by proton nuclear magnetic resonance (1H NMR) spectroscopy as a diagnostic marker. Blocking brain lysine uptake reduced glutaric acid levels and brain injury. These findings provide what we believe are new monitoring and treatment strategies that may translate for use in human GA-I.

NFE2-related factor 2 (Nrf2), an oxidant-activated CNC bZip transcription factor, has been implicated in defense against oxidative stress and chemical insults in a range of cell and tissue types, including the central nervous system. Here, we report that deletion of the Nrf2 gene in mice caused vacuolar (spongiform) leukoencephalopathy with widespread astrogliosis. The leukoencephalopathy was present in all Nrf2-null mice more than 10 months of age, was characterized by vacuolar degeneration involving all major brain regions, and was most apparent in the white tracts of the cerebellum and pons. Vacuolar degeneration in white tracts was attributable to myelin unwinding and intramyelinic cysts, and double-label immunofluorescence for 4-hydroxy-2-nonenal and myelin basic protein localized free-radical-induced oxidative damage to the myelin sheath. Moreover, the brains of Nrf2-null mice exhibited widespread astrocyte activation with profusion of glial fibrillary acidic protein-immunoreactive glial processes. The study uncovered a possible physiological role for Nrf2 in maintaining central nervous system myelin. If this role is confirmed, it may suggest new approaches to treating genetically and chemically induced myelin degenerative diseases.

Proinflammatory cytokines and chemokines have been implicated in the pathogenesis of several neurological and neurodegenerative disorders. Prominent among such factors is the pleiotropic cytokine, tumor necrosis factor (TNF)-alpha. Under normal physiological conditions, TNF-alpha orchestrates a diverse array of functions involved in immune surveillance and defense, cellular homeostasis, and protection against certain neurological insults. However, paradoxical effects of this cytokine have been observed. TNF-alpha is elicited in the brain following injury (ischemia, trauma), infection (HIV, meningitis), neurodegeneration (Alzheimer's, Parkinson's), and chemically induced neurotoxicity. The multifarious identity for this cytokine appears to be influenced by several mechanisms. Among the most prominent are the regulation of TNFalpha-induced NF-kappaB activation by adapter proteins such as TRADD and TRAF, and second, the heterogeneity of microglia and their distribution pattern across brain regions. Here, we review the differential role of TNF-alpha in response to brain injury, with emphasis on neurodegeneration, and discuss the possible mechanisms for such diverse and region-specific effects.

There is increasing evidence that obesity and overweight may be related, in part, to adverse work conditions. In particular, the risk of obesity may increase in high-demand, low-control work environments, and for those who work long hours. In addition, obesity may modify the risk for vibration-induced injury and certain occupational musculoskeletal disorders. We hypothesized that obesity may also be a co–risk factor for the development of occupational asthma and cardiovascular disease that and it may modify the worker’s response to occupational stress, immune response to chemical exposures, and risk of disease from occupational neurotoxins. We developed 5 conceptual models of the interrelationship of work, obesity, and occupational safety and health and highlighted the ethical, legal, and social issues related to fuller consideration of obesity’s role in occupational health and safety.

Continuing our previous work in which we showed wide-ranging strain differences in MPTP neurotoxicity in male mice among ten BXD recombinant inbred strains, we replicated our work in females from nine of the same strains. Mice received a single s.c. injection of 12.5 mg/kg MPTP or saline. Forty-eight hours later the striatum was dissected for neurochemical analysis. Striatal dopamine (DA) and its metabolites, DOPAC and HVA, striatal serotonin (5-HT) and its metabolite, 5-HIAA, were analyzed using HPLC. Tyrosine hydroxylase (TH) and glial fibrillary acidic protein (GFAP), an astrocytic protein that increases during the astroglial response to neural injury, were measured using ELISA. There were wide genetic variations in the DA, DOPAC, HVA, TH and GFAP responses to MPTP. We also performed principal component analysis (PCA) on the difference values, saline minus MPTP, for DA, DOPAC, HVA and TH and mapped the dominant principal component to a suggestive QTL on chromosome 1 at the same location that we observed previously for males. Moreover, there were significant correlations between the sexes for the effect of MPTP on DA, HVA, and TH. Our findings suggest that the systems genetic approach as utilized here can help researchers understand the role of sex in individual differences. The same approach can pave the way to understand and pinpoint the genetic bases for individual differences in pathology attributable to toxicants. Such systems genetics approach has broad implications for elucidating gene-environment contributions to neurodegenerative diseases.

Neurotoxicity has been linked to a number of common drugs and chemicals, yet efficient and accurate methods to detect it are lacking. There is a need for more sensitive and specific biomarkers of neurotoxicity that can help diagnose and predict neurotoxicity that are relevant across animal models and translational from nonclinical to clinical data. Fluid-based biomarkers such as those found in serum, plasma, urine, and cerebrospinal fluid (CSF) have great potential due to the relative ease of sampling compared with tissues. Increasing evidence supports the potential utility of fluid-based biomarkers of neurotoxicity such as microRNAs, F2-isoprostanes, translocator protein, glial fibrillary acidic protein, ubiquitin C-terminal hydrolase L1, myelin basic protein, microtubule-associated protein-2, and total tau. However, some of these biomarkers such as those in CSF require invasive sampling or are specific to one disease such as Alzheimer's, while others require further validation. Additionally, neuroimaging methodologies, including magnetic resonance imaging, magnetic resonance spectroscopy, and positron emission tomography, may also serve as potential biomarkers and have several advantages including being minimally invasive. The development of biomarkers of neurotoxicity is a goal shared by scientists across academia, government, and industry and is an ideal topic to be addressed via the Health and Environmental Sciences Institute (HESI) framework which provides a forum to collaborate on key challenging scientific topics. Here we utilize the HESI framework to propose a consensus on the relative potential of currently described biomarkers of neurotoxicity to assess utility of the selected biomarkers using a nonclinical model.

Neurological complications of human immunodeficiency virus (HIV) infection are a public health problem despite the availability of active antiretroviral therapies. The neuropathogenesis of HIV infection revolves around a complex cascade of events that include viral infection and glial immune activation, monocyte–macrophage brain infiltration, and secretion of a host of viral and cellular inflammatory and neurotoxic molecules. Although there is evidence that HIV-infected drug abusers experience more severe neurological disease, the biological basis for this finding is unknown. A scientific workshop organized by the National Institute on Drug Abuse (NIDA) was held on March 23–24, 2006 to address this question. The goal of the meeting was to bring together basic science and clinical researchers who are experts in NeuroAIDS, glial immunity, drugs of abuse, and/or pharmacology in order to find new approaches to understanding interactions between drug abuse and neuroAIDS. The format of the meeting was designed to stimulate open discussion and forge new multidisciplinary research collaborations. This report includes transcripts of active discussions and short presentations from invited participants. The presentations were separated into sections that included: Glial Biology, Inflammation, and HIV; Pharmacology, Neurotoxicology, and Neuroprotection; NeuroAIDS and Virology; and Virus–Drug and Immune–Drug Interactions. Research priorities were identified. Additional information about this meeting is available through links from the NIDA AIDS Research Program website ( http://www.nida.nih.gov/about/organization/arp/arp-websites.htm ).

Expression of the chemokine (C-C motif) ligand 2 (CCL2), also known as, monocyte chemoattractant protein (MCP)-1, increases in response to disease-, trauma-, or toxicant-induced damage to the central nervous system (CNS). In the periphery, endogenous and exogenous glucocorticoids are known to suppress CCL2 expression associated with inflammatory conditions. However, such actions of glucocorticoids on CCL2 expression in the CNS remain unknown. Here, we explored the effects of the glucocorticoid, corticosterone (CORT), on the expression of CCL2 and its receptors, CCR2 and CCR5, in the hippocampal formation using intact, adrenalectomized (ADX) and trimethyltin (TMT)-treated rats. An immunosuppressive regimen of CORT did not alter the mRNA expression of CCL2 or its receptors in the hippocampus. ADX, however, markedly increased the expression of CCL2 and CCR2 mRNAs in the hippocampus, while CORT replacement reversed the effects of ADX on CCL2 gene expression. Hippocampal damage resulting from systemic administration of the organometallic neurotoxicant, TMT, was associated with microglial activation, as evidenced by enhanced expression of microglial markers integrin alphaM (CD11b) and F4/80, as well as, microglia-associated factors, CCL2 and IL-1alpha. An immunosuppressive dose of CORT, suppressed TMT-induced expression of CCL2. Given the association of CCL2 with microglial activation, it appears that CORT may play a role in regulating microglial activation. However, CORT treatment did not alter TMT-mediated neuronal damage and astrogliosis. Such observations suggest that injury-related expression of microglia-associated chemokines and cytokines may subserve a role unrelated to neuronal damage. In summary, our data indicate that in the CNS, CCL2 gene expression is under negative regulation by glucocorticoids.

Enhanced expression of tumor necrosis factor (TNF) -alpha, is associated with the neuropathological effects underlying disease-, trauma- and chemically induced neurodegeneration. Previously, we have shown that deficiency of TNF receptors protects against MPTP-induced striatal dopaminergic neurotoxicity, findings suggestive of a role for TNF-alpha in neurodegeneration. Here, we demonstrate that deficiency of TNF receptors suppresses microglial activation and alters the susceptibility of brain regions to MPTP. MPTP-induced expression of microglia-derived factors, TNF-alpha, MCP-1, and IL-1alpha, preceded the degeneration of striatal dopaminergic nerve terminals and astrogliosis, as assessed by loss of striatal dopamine and TH, and an increase in striatal GFAP. Pharmacological neuroprotection with the dopamine reuptake inhibitor, nomifensine, abolished striatal dopaminergic neurotoxicity and associated microglial activation. Similarly, in mice lacking TNF receptors, microglial activation was suppressed, findings consistent with a role for TNF-alpha in striatal MPTP neurotoxicity. In the hippocampus, however, TNF receptor-deficient mice showed exacerbated neuronal damage after MPTP, as evidenced by Fluoro Jade-B staining (to identify degenerating neurons) and decreased microtubule-associated protein-2 (MAP-2) immunoreactivity. These effects were not accompanied by microglial activation, but were associated with increased oxidative stress (nitrosylation of tyrosine residues). These findings suggest that TNF-alpha exerts a neurotrophic/neuroprotective effect in hippocampus. The marked differences we observed in the regional density, distribution and/or activity of microglia and microglia-derived factors may influence the region-specific role for this cell type. Taken together, our results are indicative of a region-specific and dual role for TNF-alpha in the brain: a promoter of neurodegeneration in striatum and a protector against neurodegeneration in hippocampus.

A sample preparation method that combines a modified target plate with a nanoscale reversedphase column (nanocolumn) was developed for detection of neuropeptides by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). A gold-coated MALDI plate was modified with an octadecanethiol (ODT) self-assembled monolayer to create a hydrophobic surface that could concentrate peptide samples into a ∼200‐500-µm diameter spot. The spot sizes generated were comparable to those obtained for a substrate patterned with 200-µm hydrophilic spots on a hydrophobic substrate. The sample spots on the ODT-coated plate were 100-fold smaller than those formed on an unmodified gold plate with a 1-µl sample and generated 10 to 50 times higher mass sensitivity for peptide standards by MALDI-TOF MS. When the sample was deposited on an ODT-modified plate from a nanocolumn, thedetectionlimitforpeptideswasaslowas20 pMfor5-µlsamplescorresponding to80 amol deposited. This technique was used to analyze extracts of microwave-fixed tissue from rat brain striatum. Ninety-eight putative peptides were detected including several that had masses matching neuropeptides expected in this brain region such as substance P, rimorphin, and neurotensin. Twenty-three peptides had masses that matched peaks detected by capillary liquid chromatography with electrospray ionization MS. 1,2 Copyright  2005 John Wiley & Sons, Ltd.

The effect of sample extraction and preparation on neuropeptidomic analysis of brain tissue by capillary liquid chromatography with tandem mass spectrometry was investigated. In agreement with previous reports, analysis of peptide extracts of brain tissue from animals sacrificed by microwave irradiation, which fixes tissue, allows identification of neuronally derived peptides whereas similar analysis of tissue from animals sacrificed without fixation does not. A comparison of a physical method for cell lysis (sonication) to physical combined with chemical cell lysis (sonication with detergent treatment) revealed that the latter method increased the number of neuronally derived peptides positively identified by approximately 3-fold, from 16 to 44, for analysis of microwave-fixed rat striatum. Use of synaptosome preparations also allowed detection of neuronally derived peptides (23 positively identified) without a requirement of microwave fixation, suggesting that this method may be a useful alternative for sample preparation. Although numerous peptides were identified in these experiments, several known neuropeptides were not detected including neuropeptide Y and neurotensin. Chemical properties such as hydrophobicity and atypical gas-phase fragmentation were found to account for the inability to detect these peptides. These results suggest that further improvement in sample preparation and automated spectral interpretation are needed to provide better coverage of neuropeptides in mammalian tissues. A total of 39 novel neuronally derived peptides, including some originating from proenkephalin and phosphatidylethanolamine binding protein, were identified in striatum and synaptosome.

It has been known for some time that cytokines made and released during systemic illness can result in a constellation of symptoms strikingly similar to those observed in depression. The overlap of the symptoms of depression and systemic illness raises the intriguing possibility that cytokines may be involved in the development and maintenance of mood disorders. Cytokines are small ubiquitous pleiotropic molecules that are made and released in response to a variety of stimuli. They have a multitude of actions throughout the body, including actions on the central and peripheral nervous systems. Alterations in the levels of circulating cytokines, especially the key proinflammatory cytokines, interleukin 6 and tumor necrosis factor alpha , have been linked to a variety of disease states including those involving central nervous system depression. In this brief review, epidemiological and clinical data on depression, as well as findings from relevant animal models, are examined for links between cytokine expression and depression. We suggest that glial cells, both as a source and target of cytokines, represent the overlooked targets involved in the etiology of depression.

We examined 204 decedents of the autopsy component of the Honolulu-Asia Aging Study, a longitudinal cohort study, who had been clinically assessed for dementia. A sensitive ELISA technique was used to quantify glial fibrillary acidic protein (GFAP), a marker for astrogliosis, in four specific cortical brain regions and assess associations between GFAP and 1) a measure of cognitive function, 2) several clinical dementia conditions, and 3) neuritic plaque (NP) and neurofibrillary tangle (NFT) formation. Cognitive function was inversely associated with GFAP in the occipital, parietal and temporal lobes, but not in the frontal lobe. This relationship remained significant when the contribution of NP and NFT counts was removed. Further, compared to brain samples from non-demented individuals, significantly greater GFAP levels were found in samples from individuals diagnosed with Alzheimer's disease, mixed dementia, and vascular mediated dementia. Because elevated levels of GFAP reflect astroglial responses to even subtle forms of neural damage, our data indicate that increments in GFAP may provide independent, supporting evidence for the damage underlying dementia, even in the absence of other evidence of neuropathology such as the presence of NPs or NFTs. Our findings underscore the need to look beyond standard neuropathological measures putatively linked to specific neuropathological conditions in efforts to identify common cellular and molecular processes that contribute to dementia.

At any point in time, net protein phosphorylation represents the contribution of protein kinase and protein phosphatase activities affecting a specific site on a given substrate. Preservation of phosphorylated proteins in neural tissues has traditionally included flash-freezing or fresh tissue processing following tissue isolation. Rapid heat inactivation of protein kinases and phosphatases by focused microwave irradiation sacrifice represents another method to preserve, in vivo, brain protein phosphorylation state. In this study, we compared preservation of the phosphorylation state of a variety of phosphoproteins in the brain following sacrifice of mice by decapitation, decapitation into liquid nitrogen and focused microwave irradiation. We found that microwave irradiation generally provided the highest and most consistent levels of protein phosphorylation, regardless of the substrates examined in striatum and hippocampus. In general, flash-freezing resulted in the least preservation of phospho-state with ERK1/2 and CREB showing almost complete dephosphorylation. When regions of freshly decapitated brains were homogenized and incubated on ice for 30 min, ERK1/2 phosphorylation was completely lost, whereas it was well preserved in microwaved samples left at room temperature for 2 h. Loss of ERK1/2 phosphorylation in the fresh samples could not be attributed to substrate proteolysis. Our results indicate that focused microwave irradiation sacrifice may be required to achieve biologically relevant data for the in vivo protein phosphorylation state of many phosphoproteins.

In the mouse but not the rat, d-3,4-methylenedioxymethamphetamine (d-MDMA) is a dopaminergic neurotoxicant. Various stressors and hypothermia protect against d-MDMA-induced neurotoxicity through unknown mechanisms, one of which could be a reduction in the distribution of d-MDMA to the brain. We determined striatal levels of d-MDMA in relation to body temperature in mice exposed to a neurotoxic regimen of d-MDMA in the presence or absence of various stressors. Female C57BL6/J mice received a neurotoxic regimen of d-MDMA (15.0 mg/kg s.c. as the base every 2 h×4) alone or in combination with manipulations with a known neuroprotective status. d-MDMA levels were determined by HPLC with fluorometric detection while rectal temperature provided core temperature status. Levels of dopamine, tyrosine hydroxylase and GFAP were used to assess neurotoxicity. Restraint, ethanol co-treatment and cold stress were neuroprotective, caused hypothermia and increased striatal d-MDMA levels by 4- to 7-fold. Corticosterone treatment, as a stress mimic, did not alter striatal d-MDMA or temperature and was not protective. The protective glutamate receptor antagonist, MK-801, doubled striatal d-MDMA levels and caused hypothermia. Although stress and other protective manipulations can alter the striatal concentration of d-MDMA their hypothermia-inducing properties appear a more likely determinant of their neuroprotection against the striatal dopaminergic neurotoxicity of d-MDMA.

1-Methyl-4-(2'-aminophenyl)-1,2,3,6-tetrahydropyridine (2'-NH2-MPTP) causes long-term loss of forebrain serotonin (5-HT) and norepinephrine (NE) and consequently, is unlike 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and its other 2'-analogs that primarily deplete striatal dopamine (DA). In the present investigation into the acute effects of 2'-NH2-MPTP in mice, profound decreases in cortical and hippocampal 5-HT and NE to 10 to 40% of control were observed as early as 30 min post-treatment and lasted throughout the ensuing 21 days. Striatal DA was decreased to 60 to 80% of control during the first 48 h but returned to normal by 72 h. Reactive gliosis, which occurs in response to neurodegeneration was not evident by immunocytochemistry but was detected by enzyme-linked immunosorbent assay, where glial fibrillary acidic protein (GFAP) was increased to 130% of control in cortex, hippocampus, and brain stem 48 to 72 h post-treatment. To explore the possibility that 5-HT modulates the astrocytic response to injury, 2'-NH2-MPTP was used to damage 5-HT axons 2 weeks before administration of the potent DA neurotoxin 1-methyl-4-(2'-methylphenyl)-1,2,3,6-tetrahydropyridine (2'-CH3-MPTP). Despite a 90% decrement in striatal DA in 2'-NH2-MPTP/2'-CH3-MPTP-treated mice, increases in GFAP were attenuated compared to mice treated with 2'-CH3-MPTP alone. Thus, 2'-NH2-MPTP causes severe and immediate decrements in 5-HT and NE in frontal cortex and hippocampus, yet induces a modest GFAP response compared with other MPTP analogs that have their primary effect on DA. These results demonstrate the importance of obtaining quantitative assessments of GFAP to detect astroglial responses associated with selective damage to neurotransmitter systems with low-density innervation and suggest that serotonin may facilitate the astrocytic response to striatal injury.

Obesity is a major risk factor associated with a variety of human disorders. While its involvement in disorders such as diabetes, coronary heart disease and cancer have been well characterized, it remains to be determined if obesity has a detrimental effect on the nervous system. To address this issue we determined whether obesity serves as a risk factor for neurotoxicity. Model neurotoxicants, methamphetamine (METH) and kainic acid (KA), which are known to cause selective neurodegeneration of anatomically distinct areas of the brain, were evaluated using an animal model of obesity, the ob/ob mouse. Administration of METH and KA resulted in mortality among ob/ob mice but not among their lean littermates. While METH caused dopaminergic nerve terminal degeneration as indicated by decreased striatal dopamine (49%) and tyrosine hydroxylase protein (68%), as well as an increase in glial fibrillary acidic protein by 313% in the lean mice, these effects were exacerbated under the obese condition (96%, 86% and 602%, respectively). Similarly, a dosage of KA that did not increase glial fibrillary acidic protein in lean mice increased the hippocampal content of this protein (93%) in ob/ob mice. KA treatment resulted in extensive neuronal degeneration as determined by Fluoro-Jade B staining, decreased hippocampal microtubule-associated protein-2 immunoreactivity and increased reactive gliosis in ob/ob mice. The neurotoxic outcome in ob/ob mice remained exacerbated even when lean and ob/ob mice were dosed with METH or KA based only on a lean body mass. Administration of METH or KA resulted in up-regulation of the mitochondrial uncoupling protein-2 to a greater extent in the ob/ob mice, an effect known to reduce ATP yield and facilitate oxidative stress and mitochondrial dysfunction. These events may underlie the enhanced neurotoxicity seen in the obese mice. In summary, our results implicate obesity as a risk factor associated with chemical- and possibly disease-induced neurodegeneration.

Enhanced expression of proinflammatory cytokines and chemokines has long been linked to neuronal and glial responses to brain injury. Indeed, inflammation in the brain has been associated with damage that stems from conditions as diverse as infection, multiple sclerosis, trauma, and excitotoxicity. In many of these brain injuries, disruption of the blood-brain barrier (BBB) may allow entry of blood-borne factors that contribute to, or serve as the basis of, brain inflammatory responses. Administration of trimethyltin (TMT) to the rat results in loss of hippocampal neurons and an ensuing gliosis without BBB compromise. We used the TMT damage model to discover the proinflammatory cytokines and chemokines that are expressed in response to neuronal injury. TMT caused pyramidal cell damage within 3 days and a substantial loss of these neurons by 21 days post dosing. Marked microglial activation and astrogliosis were evident over the same time period. The BBB remained intact despite the presence of multiple indicators of TMT-induced neuropathology. TMT caused large increases in whole hippocampal-derived monocyte chemoattractant protein (MCP)-1 mRNA (1000%) by day 3 and in MCP-1 (300%) by day 7. The mRNA levels for tumor necrosis factor (TNF)-α, interleukin (IL)-1β and IL-6, cytokines normally expressed during the earliest stage of inflammation, were not increased up to 21 days post dosing. Lipopolysaccharide, used as a positive control, caused large inductions of cytokine mRNA in liver, as well as an increase in IL-1β in hippocampus, but it did not result in the induction of astrogliosis. The data suggest that enhanced expression of the proinflammatory cytokines, TNF-α, IL-1β and IL-6, is not required for neuronal and glial responses to injury and that MCP-1 may serve a signaling function in the damaged CNS that is distinct from its role in proinflammatory events.

Premarin, which contains several equine estrogens, as well as estradiol (E2) as a minor component, is widely used for replacement therapy of estrogen deficits, but little is known of its direct actions on brain cells. In mixed glial cultures, apolipoprotein E (apoE) and glial fibrillary acidic protein (GFAP) are induced by estrogens. GFAP induction showed an inverted-U shape E2 dose response, with a maximum induction at 1 pM, whereas apoE mRNA induction was greatest at 100 pM. GFAP and ApoE mRNAs were induced by equine estrogens in the following order: E2=equilinestrone17 alpha-dihydroequilenin. However, the induction of apoE secretion by 17 alpha-dihydroequilenin was as effective as by the other estrogens. The greater response of apoE secretion than GFAP mRNA induction to 17 alpha-dihydroequilenin might be therapeutically important because of the glial scarring during brain lesions, in which GFAP induction has a major role in inhibiting neurite outgrowth, whereas apoE secretion supports neurite outgrowth.

Sporadic or idiopathic Parkinson's disease (PD) is an age-related neurodegenerative disorder of unknown origin that ranks only second behind Alzheimer's disease (AD) in prevalence and its consequent social and economic burden. PD neuropathology is characterized by a selective loss of dopaminergic neurons in the substantia nigra pars compacta; however, more widespread involvement of other CNS structures and peripheral tissues now is widely documented. The onset of molecular and cellular neuropathology of PD likely occurs decades before the onset of the motor symptoms characteristic of PD. The hallmark symptoms of PD, resting tremors, rigidity and postural disabilities, are related to dopamine (DA) deficiency. Current therapies treat these symptoms by replacing or boosting existing DA. All current interventions have limited therapeutic benefit for disease progression because damage likely has progressed over an estimated period of ~5 to 15 years to a loss of 60%–80% of the nigral DA neurons, before symptoms emerge. There is no accepted definitive biomarker of PD. An urgent need exists to develop early diagnostic biomarkers for two reasons: (1) to intervene at the onset of disease and (2) to monitor the progress of therapeutic interventions that may slow or stop the course of the disease. In the context of disease development, one of the promises of personalized medicine is the ability to predict, on an individual basis, factors contributing to the susceptibility for the development of a given disease. Recent advances in our understanding of genetic factors underlying or contributing to PD offer the potential for monitoring susceptibility biomarkers that can be used to identify at-risk individuals and possibly prevent the onset of disease through treatment. Finally, the exposome concept is new in the biomarker discovery arena and it is suggested as a way to move forward in identifying biomarkers of neurological diseases. It is a two-stage scheme involving a first stage of exposome-wide association studies (EWAS) to profile omic features in serum to discover molecular biomarkers. The second stage involves application of this knowledge base in follow-up studies. This strategy is unique in that it promotes the use of data-driven (omic) strategies in interrogating diseased and healthy populations and encourages a movement away from using only reductionist strategies to discover biomarkers of exposure and disease. In this short review we will examine 1) advances in our understanding of the molecular mechanisms underlying PD that have led to candidate biomarkers for diagnosis and treatment efficacy and 2) new technologies on the horizon that will lead to novel approaches in biomarker development.

Epidemiological studies show a positive association between adequate sleep and good health. Further, disrupted sleep may increase the risk for CNS diseases, such as stroke and Alzheimer’s disease. However, there has been limited progress in determining how sleep is linked to brain health or how sleep disruption may increase susceptibility to brain insult and disease. Animal studies can aid in understanding these links. In reviewing the animal literature related to the effects of sleep disruption on the brain, we found most of the work was directed toward investigating and characterizing the role of various brain areas or structures in initiating and regulating sleep. In contrast, limited effort has been directed towards understanding how sleep disruption alters the brain’s health or susceptibility to insult. We also note many current studies have determined the changes in the brain following compromised sleep by examining, for example, the brain transcriptome or to a more limited extent the proteome. However, these studies have utilized almost exclusively total sleep deprivation (e.g., 24 out of 24 hours) paradigms or single short periods of limited acute sleep deprivation (e.g., 3 out of 24 hours). While such strategies are beneficial in understanding how sleep is controlled, they may not have much translational value for determining links between sleep and brain health or for determining how sleep disruption may increase brain susceptibility to insult. Surprisingly, few studies have determined how the duration and recurrence of sleep deprivation influence the effects seen after sleep deprivation. Our aim in this review was to identify relevant rodent studies from 1980 through 2012 and analyze those that use varying durations of sleep deprivation or restriction in their effort to evaluate the effects of sleep deprivation on the brain transcriptome and to a more limited extent the proteome. We examined how differences in the duration of sleep deprivation affect gene and protein expression to better understand the full consequences of repeated sleep disruption on the brain. Future research needs to consider and emphasize how the type and extent of the sleep deprivation exposure impacts the conclusions reached concerning the influence of sleep disruption on the brain. We identified relevant studies between 1980 and 2012 by searching the electronic databases of PubMed, Medline (Ovid), Embase (Ovid), and Web of Science using the terms “sleep” AND “disrupt”, “deprivation”, “restrict”, “fragment”, “loss”, “disturb”, “disorder”, “dysfunction”, “brain”, “cortex”, striatum”, hypothalamus”, “hippocampus”, “gene”, “protein”, “genomics”, “proteomics”, “polymerase chain reaction”, “pcr”, “microarray”, “molecular”, “rodent” “rat”, “rats”, “mouse”, “mice”. All searches were limited to rodent studies in English and the reference lists of retrieved articles were searched for additional pertinent studies. Electronic supplementary material The online version of this article (doi:10.1186/2193-1801-3-728) contains supplementary material, which is available to authorized users.

The goal of the GWI consortium is to develop a better understanding of GWI and identify specific disease targets to find treatments that will address the cause of the disease. The consortium will integrate our clinical understanding of the disease process with basic research efforts using a novel mathematical model. The computational biology approach will enable the consortium to quickly identify targets of dysfunction and find treatments that will address the causes of the disease. The project will combine animal models of GWI with focus on the immune, cardiovascular and autonomic systems.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a pro-neurotoxicant that must be metabolized to 1-methyl-4-phenylpyridinium (MPP + ) and taken up into striatal dopaminergic neurons to produce neurodegeneration. Recently, we showed wide genetic variability in MPTP-associated neuronal damage in a panel of recombinant inbred mouse strains. Here we examined the amount of MPP + produced in the striatum in the same strains of inbred BXD mice. This allowed us to determine if the differences in the dopaminergic neurotoxicity and associated astrogliosis among the BXD mouse strains were due to differential metabolism of MPTP to MPP + . Using the same BXD mouse strains examined previously (Jones et al., 2013) we found that the extent of the striatal damage produced following MPTP treatment is not correlated quantitatively with the production of MPP + in the striatum. Our findings also extend those of others regarding strain differences in MPTP-induced dopaminergic neurotoxicity. Importantly, our finding suggests that additional factors influence the neurodegenerative response other than the presence and amount of the toxicant at the target site.

Reports of methamphetamine-related emergency room visits suggest that elevated body temperature is a universal presenting symptom, with lethal overdoses generally associated with extreme hyperthermia. This review summarizes the available information on methamphetamine toxicity as it pertains to elevations in body temperature. First, a brief overview of thermoregulatory mechanisms is presented. Next, central and peripheral targets that have been considered for potential involvement in methamphetamine hyperthermia are discussed. Finally, future areas of investigation are proposed, as further studies are needed to provide greater insight into the mechanisms that mediate the alterations in body temperature elicited by methamphetamine.

Astrogliosis, characterized by the enhanced expression of GFAP, represents a remarkably homotypic response of astrocytes to all types of injuries of the CNS, including injuries of the developing CNS. As such, astrocytes serve as microsensors of the injured microenvironment regardless of their location in the CNS. The diversity of insults that engender astrogliosis and the brain-wide nature of the astrocytic response suggest that common injury factors serve as the trigger of this cellular reaction. One prominent theme that has emerged in recent years is that proinflammatory cytokines and chemokines serve as a stimulus for induction of astrogliosis. Here we present a brief critique of this hypothesis based on a review of literature and some of our own recentfindings. Studies of astrocytes, in vitro, clearly indicate that these cell types are responsive to a variety of growth factors, including cytokines and chemokines. A somewhat different picture, however, can be seen from data obtained in vivo. It is true that trauma and diseases of the nervous system, as well as some exposures to neurotoxic chemicals, can be associated with the expression in brain of large varieties of cytokines and chemokines. That these same conditions result in astrogliosis has fostered the circumstantial link between cytokine/chemokine expression and the induction of astrogliosis. Several lines of evidence argue against this view, including (a) suppression of cytokine expression does not suppress gliosis, (b) gliosis can occur in the absence of enhanced expression of cytokines, (c) elevations in brain cytokines can occur in the absence of gliosis and (d) the patterns of cytokine expression in the adult and developing CNS are more consistent with a trophic role for these chemical messengers rather than a role in the induction of inflammation. Enhanced expression of cytokines and chemokines after brain injury appear to be signal transduction events unrelated to the induction of astrogliosis.

A long-term induction of Fos-related antigens has been shown in neurons after brain injury, suggesting that Fos-related antigens are involved in enhancing the transcription of genes related to the process of regeneration and repair. In the present study, we report that levels of Fos-related antigen-2 are elevated in several models of chemically induced brain injury. Trimethyltin, which causes degeneration of neurons primarily in the hippocampus and other limbic regions, results in a five-fold induction of Fos-related antigen-2 immunoreactivity in neurons in the pyramidal and dentate layers of the hippocampus starting at seven days post-treatment and persisting for 60 days. Methamphetamine and methylenedioxymethamphetamine, agents which cause degeneration of dopaminergic nerve terminals in the striatum of the mouse, cause an increase in Fos-related antigen-2 immunoreactivity which begins at three days post-treatment and returns to basal levels by days 5 and 15, respectively. Treatment with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine elevated levels of Fos-related antigen-2 in the mouse striatum at three days post-treatment. This abbreviated time-course of Fos-related antigen-2 induction is consistent with less severe insult (terminal damage) relative to trimethyltin (cell death), but induction occurs during the period of regeneration and repair in both models. Dexfenfluramine, a non-neurotoxic amphetamine, does not induce Fos-related antigen-2 expression. Decreasing core temperature of the mouse, which blocks amphetamine-induced neurotoxicity, also blocks Fos-related antigen-2 induction. In summary, Fos-related antigen-2 is induced in models of both cell death and terminal degeneration, suggesting that this transcription factor may serve as a universal signal transduction molecule involved in the regulation of genes related to regeneration and repair in the CNS.

A number of substituted amphetamines, including methamphetamine (METH) are considered dopaminergic neurotoxicants. METH causes long-term depletions of striatal dopamine (DA) and its metabolites (DOPAC and HVA) that are accompanied by other changes indicative of nerve terminal degeneration. These include argyrophilia as detected by silver degeneration stains and an elevation in glial fibrillary acidic protein (GFAP), a marker of reactive gliosis in response to injury, as well as a long-term decrease in tyrosine hydroxylase (TH) protein levels. The susceptibility to the dopaminergic neurotoxicity of METH and the other amphetamines can be affected by a number of factors including age, gender, stress, and environment. Many of these susceptibility factors have been extensively investigated in the rat but less so in the mouse. As the availability of genetically altered mice continues to expand, this species is increasingly selected for study. Thus, in previous work we determined that stress, gender, and the environment can significantly impact the neurotoxicity of the amphetamines. Here we determined how age affects the striatal DA depletion and GFAP elevation induced by d-METH in C57BL/6 mice. Age was a significant determinant of the ability of a known neurotoxic regimen of d-METH (10 mg/kg x 4) to produce striatal DA depletion with one-month-old C57BL16 mice displaying minimal and nonpersistent depletion of DA or its metabolites while mice 12 months of age displayed large and persistent depletions of DA (87%), DOPAC (71%), and HVA (94%). Large elevations in striatal GFAP were induced in mice 2-23 months of age by d-METH, with lower dosages of d-METH being effective in the older mice. In contrast, the usual neurotoxic regimen of d-METH was minimally effective in inducing GFAP elevations (49% over control) in one-month-old mice, despite elevations in body temperature equivalent to those observed in older mice. Although increasing the dosage of d-METH (20 to 80 mg/kg) did increase the GFAP response (100% over control), it was still well below that usually exhibited at the usual neurotoxic dosage (300-400% over control) in fully mature mice. These data suggest maturity of striatal dopamine systems may be an essential element in the striatal damage induced by the neurotoxic amphetamines.

AP-1 transcription factors have been shown to be induced in the brain after ischemic injury. However, their roles in neuronal survival or death have yet to be defined. Here, we report the discovery of elevated nuclear levels of fos-related antigen-2 (FRA-2) in the nuclei of hippocampal neurons seven days after middle cerebral artery occlusion (MCAO). Expression of FRA-2 and AP-1 DNA binding activity is elevated in hippocampi ipsilateral as well as contralateral to MCAO. Using Fluoro-Jade staining as a marker of neurodegeneration, FRA-2 was not found to be expressed in degenerating neurons. Thus, FRA-2 is expressed in neurons that survive ischemic insult suggesting a role for this transcription factor in neuronal adaptation to the post-injury state.

Specific protein phosphorylation pathways have been shown to play a role in cellular adaptation responses underlying addiction to psychostimulants such as methamphetamine and cocaine. Transcriptional regulation through fos-related antigens constitutes one element through which these dopaminergic agonists exert their persistent actions. In addition to their addictive properties, amphetamines are known to damage dopaminergic nerve terminals. Although not widely appreciated, protein phosphorylation cascades and fos-related antigens also may play a role in the neurotoxic actions of substituted amphetamines such as methamphetamine. Here we document the involvement of the dopaminoceptive phosphoprotein, DARPP-32, the fos-related antigen, FRA-2, and the growth associated protein kinase, MAP kinase, in the neurotoxic action of known dopaminergic neurotoxicants, including methamphetamine. The addictive and neurotoxic properties of psychostimulants may share some molecular signaling mechanisms.

We compared the effects of treatment with high doses of fluoxetine, sibutramine, sertraline, and dexfenfluramine for 4 days on brain serotonergic nerve terminals in rats. Methylenedioxymethamphetamine (MDMA) and 5,7-dihydroxytryptamine (5,7-DHT) were used as positive controls because both compounds deplete brain serotonin. Food intake and body weight changes were also monitored and yoked, pair-fed animals were used to control for possible changes in morphology due to nutritional deficits. Fluoxetine, sibutramine, sertraline and dexfenfluramine all produced a significant reduction in body weight. Fluoxetine, sibutramine and sertraline treatment resulted in no depletion of brain serotonin but produced morphological abnormalities in the serotonergic immunoreactive nerve network. In contrast, dexfenfluramine and MDMA depleted brain serotonin and produced morphological changes in the serotonin nerve network. These results indicate that even though fluoxetine, sibutramine and sertraline do not deplete brain serotonin, they do produce morphological changes in several brain regions (as identified by serotonin immunohistochemistry). Dexfenfluramine and MDMA, on the other hand, markedly deplete brain serotonin and also produce morphological changes. Collectively, these results lend support to the concept that all compounds acting on brain serotonin systems, whether capable of producing serotonin depletion or not, could produce similar effects on the morphology of cerebral serotonin systems.

Dopaminergic neurons exert a major modulatory effect on the forebrain. Dopamine and adenosine 3′,5′-monophosphate–regulated phosphoprotein (32 kilodaltons) (DARPP-32), which is enriched in all neurons that receive a dopaminergic input, is converted in response to dopamine into a potent protein phosphatase inhibitor. Mice generated to contain a targeted disruption of the DARPP-32 gene showed profound deficits in their molecular, electrophysiological, and behavioral responses to dopamine, drugs of abuse, and antipsychotic medication. The results show that DARPP-32 plays a central role in regulating the efficacy of dopaminergic neurotransmission.

The reproductive properties of estrogen are well established, but it is now evident that this steroid hormone has substantial modulatory capabilities in nonreproductive systems. For example, estrogen may be neuroprotective as Alzheimer's disease progresses more slowly in women receiving hormone replacement therapy, and Parkinson's disease affects more men than women. Gender affects both the functional and biochemical responses of the nigral-striatal pathway to dopaminergically active compounds. To begin to evaluate the possible neuroprotective effects of estrogen in this pathway, we first determined if gender affected the dopaminergic striatal neurotoxicity induced by two different neurotoxicants, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and methamphetamine (METH). Both agents induced greater neurotoxicity in males than females as evidenced by greater striatal dopamine (DA) depletions. An examination of striatal levels of 1-methyl-4-phenylpyridium ion (MPP+) following MPTP treatment established that the observed gender differences were not due to metabolic/pharmacokinetic variables. The neurotoxicity of MPTP was then examined in ovariectomized (OVX) mice. Estrogen replacement reduced the DA, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) depletions as well as the glial fibrillary acidic protein (GFAP) elevation induced by MPTP, which indicates that estrogen has neuroprotective properties in this model of striatal dopaminergic neurotoxicity. Surprisingly, estrogen supplementation did not protect against the neurotoxic effects of MPTP in intact 2-yr-old intact female mice, suggesting that low endogenous levels of estrogen may provide neuroprotection.

Responses to injury in the ageing hippocampus were assessed utilizing the synaptic markers glial fibrillary acidic protein and synaptosomal-associated protein (mol. wt 25 000) following administration of the neurotoxin, 5,7-dihydroxytryptamine, into the fimbria–fornix and cingulum bundle to denervate serotonergic afferent input to the dorsal hippocampus. Age-dependent alterations in hippocampal immunohistochemical localization of glial fibrillary acidic protein and synaptosomal-associated protein were evaluated in female Fischer 344 rats following serotonergic deafferentation with 5,7-dihydroxytryptamine. Across the lifespan, as indicated by measurements taken at three, 18, 21 and 29 months, marked increases in glial fibrillary acidic protein, but not synaptosomal-associated protein immunoreactivity, occurred throughout the hippocampus at 21 and 29 months compared to three and 18 months. Following three weeks pretreatment with 5,7-dihydroxytryptamine (20 μ g total dose) or vehicle (0.1% ascorbic saline; 2 μ l total volume) infused in the fimbria–fornix/cingulum bundle, immunohistochemical analysis demonstrated marked increases of glial fibrillary acidic protein, but not synaptosomal-associated protein, in the 18-month 5,7-dihydroxytryptamine group compared to the 18-month vehicle and 3-month 5,7-dihydroxytryptamine groups. Additionally, a significant increase in glial fibrillary acidic protein concentration was found by enzyme-linked immunosorbent asssay in the 18-month 5,7-dihydroxytryptamine group compared to the 18-month vehicle and three-month 5,7-dihydroxytryptamine groups. These results demonstrate that selective neurotoxicant damage of the hippocampal serotonergic system differentially alters the expression of glial fibrillary acidic protein. This approach may provide a valuable tool to determine the ability of the hippocampus to respond to age-related neurodegenerative injury.

Brain damage following administration of domoic acid, a structural analog of the excitatory amino acids glutamic acid and kainic acid, was compared using different anatomic markers in adult rats. Seven days after administration of domoic acid (2.25 mg/kg i.p.) or vehicle, brains were collected and sectioned and stained to visualize Nissl substance using thionin, argyrophilia using a cupric silver staining method, astroglia using immunohistochemistry to detect glial fibrillary acidic protein-like immunoreactivity (GFAP-ir), and activated microglia using lectin histochemistry to detect Griffonia simplicifolia I-B4 isolectin (GSI-B4) binding in adjacent sections. In approximately 60% of rats to which it was administered, domoic acid caused stereotyped behavior within 60 min, followed by convulsions within 2-3 h. Brains of domoic acid-administered rats that did not manifest stereotyped behavior or convulsions did not differ from brains from vehicle-administered controls. In animals that had manifested stereotyped behavior and convulsions, Nissl staining was mostly unremarkable in brain sections. In contrast, there was intense argyrophilia in anterior olfactory nucleus, CA1 hippocampus, lateral septum, parietal (layer IV), piriform, and entorhinal cortices, ventral posterolateral thalamus, and amygdala. This pattern was reminiscent of that seen in postmortem specimens from humans who consumed domoic acid-tainted mussels and in experimental animals after kainic acid administration. Adjacent sections displayed astrogliosis, evidenced by increased GFAP-ir, which was more diffuse than the argyrophilic reaction. Activated microglia were revealed using GSI-B4 histochemistry. These data suggest activation of discrete brain circuits in rats that convulse following domoic acid administration and subsequent pathological alterations. The data strongly suggest that neuropathology following domoic acid occurs only in animals manifesting domoic acid-induced sterotypy and convulsions. The data do not rule out more insidious damage in behaviorally normal rats that receive domoic acid.

Methamphetamine (METH) exposure results in dopaminergic neurotoxicity in striatal regions of the brain, an effect that has been linked to an increased risk of Parkinson's disease. Various aspects of neuroinflammation, including astrogliosis, are believed to be contributory factors in METH neurotoxicity. METH interacts with sigma receptors at physiologically relevant concentrations and treatment with sigma receptor antagonists has been shown to mitigate METH-induced neurotoxicity in rodent models. Whether these compounds alter the responses of glial cells within the central nervous system to METH however has yet to be determined. Therefore, the purpose of the current study was to determine whether the sigma receptor antagonist, SN79, mitigates METH-induced striatal reactive astrogliosis. Male, Swiss Webster mice treated with a neurotoxic regimen of METH exhibited time-dependent increases in striatal gfap mRNA and concomitant increases in GFAP protein, indicative of astrogliosis. This is the first report that similar to other neurotoxicants that induce astrogliosis through the activation of JAK2/STAT3 signaling by stimulating gp-130-linked cytokine signaling resulting from neuroinflammation, METH treatment also increases astrocytic oncostatin m receptor (OSMR) expression and the phosphorylation of STAT3 (Tyr-705) in vivo. Pretreatment with SN79 blocked METH-induced increases in OSMR, STAT3 phosphorylation and astrocyte activation within the striatum. Additionally, METH treatment resulted in striatal cellular degeneration as measured by Fluoro-Jade B, an effect that was mitigated by SN79. The current study provides evidence that sigma receptor antagonists attenuate METH-induced astrocyte activation through a pathway believed to be shared by various neurotoxicants.

We analyzed genetic variation in severity of neuronal damage using the known dopaminergic neurotoxicant, MPTP, as a prototypical chemical denervation agent. Male mice from ten members of the BXD family of recombinant inbred strains received 12.5 mg/kg MPTP s.c. (vs. saline) and 48 h later brains were taken for multiple related biochemical analyses. Striatal dopamine (DA) and its metabolites, DOPAC and HVA, and serotonin and its metabolite, 5-HIAAA, were analyzed by HPLC. DA turnover was assessed using DOPAC/DA and HVA/DA ratios. Striatal tyrosine hydroxylase (TH), glial fibrilary acidic protein (GFAP), and iron content in ventral midbrain were quantified. All dopamine measures, as well as TH and GFAP, demonstrated wide, genotype-dependent differences in response to MPTP. Serotonin was largely unaffected. Principal components analysis (PC) on difference values, saline minus MPTP, for DA, DOPAC, HVA, and TH, yielded a dominant principal component. The PC trait residuals for each genotype were compared against complementary expression data for striatum of the same strains. Three transcripts representing Mtap2, Lancl 1, and Kansl1l were highly correlated with the PC, as was the difference score, MPTP minus saline for GFAP. This systems approach to the study of environmental neurotoxicants holds promise to define individual genetic differences that contribute to variability in susceptibility to risk factors for diseases such as Parkinson's disease.