Journal of Neuroscience Research

Persistent neural activity regulates Arc/Arg3.1 transcription in the dentate gyrus.

Publication Date: 2010 Aug 30 PMID: 20806412
Authors: Toyoda, T. - Nomura, H. - Hashikawa, K. - Nonaka, A. - Matsuki, N.
Journal: J Neurosci Res

The activity-regulated cytoskeleton-associated gene (Arc, also known as Arg3.1) is an effector immediate-early gene rapidly induced by strong neural activity. Although a number of studies have revealed significant functions of Arc and Arc has come into widespread use as a neural activity marker in behavioral studies, the mechanisms regulating Arc transcription remain unclear. Here, we examined the conditions of Arc transcription in acute slices of dentate gyrus. Surprisingly, kainic acid (1 muM to 10 mM) application to slices did not induce Arc transcription, although intraperitoneal injection of kainic acid (20 mg/kg) induced robust Arc transcription. No types of high-frequency stimulation examined induced Arc transcription in acute slices. These findings indicate that Arc transcription is dramatically suppressed in acute slices of the dentate gyrus, in which background neural activity is markedly reduced. Burst stimulation increased the number of Arc-expressing cells in the presence of picrotoxin, in which excitation was maintained even after the end of stimulation. Moreover, the involvement of background neural activity in Arc transcription was tested by application of carbachol, a muscarinic receptor agonist. Carbachol also increased the number of Arc-expressing cells, which was blocked by atropine, a muscarinic receptor antagonist. Taken together, these findings suggest that persistent background activity is critical for Arc transcription. (c) 2010 Wiley-Liss, Inc.

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Up-regulation of erythropoietin receptor by nitric oxide mediates hypoxia preconditioning.

Publication Date: 2010 Aug 30 PMID: 20806411
Authors: Chen, Z. Y. - Wang, L. - Asavaritkrai, P. - Noguchi, C. T.
Journal: J Neurosci Res

Erythropoietin (Epo), known to stimulate erythroid progenitor cell survival, proliferation, and differentiation, has been shown to be neuroprotective against brain ischemia in animal models. Both Epo and Epo receptor (EpoR) are expressed in the brain and are up-regulated by hypoxia. Brain Epo signaling can stimulate neural cell survival and prevent neuron apoptosis. Neurons from EpoR null mice exhibit marked increased sensitivity to hypoxia. In endothelial cells, Epo has been shown to stimulate nitric oxide (NO) production, particularly at low pO(2). We found here that the EpoR expression on neural cells and Epo's neuroprotective effect were regulated by NO. Hypoxia increased NO production as well as EpoR expression, and inhibition of NOS activity reduced the proportion of EpoR-expressing neurons induced at low pO(2). Conversely, addition of NO donor to cultures grown under normoxia induced EpoR. Similarly, NO donor increased EpoR promoter activity in a reporter gene assay, suggesting that NO regulates EpoR at the transcription level. Preincubation of neurons with NO results in induction of EpoR, which gives rise to protection against hypoxia even in the absence of exogenous Epo, although at high concentration NO is toxic. These data provide evidence of a role for NO in Epo activity in brain and suggest links between NO production, EpoR expression, and Epo signaling in neuroprotection. Published 2010 Wiley-Liss, Inc.

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I(h) "run-up" in rat neocortical neurons and transiently rat or human HCN1-expressing HEK293 cells.

Publication Date: 2010 Aug 30 PMID: 20806410
Authors: Battefeld, A. - Bierwirth, C. - Li, Y. C. - Barthel, L. - Velmans, T. - Strauss, U.
Journal: J Neurosci Res

Hyperpolarization-activated cyclic nucleotide-gated ion channels (HCN) are key determinants of CNS functions. Here we describe an increase in hyperpolarization-activated current (I(h)) at the beginning of whole-cell recordings in rat layer 5 cortical neurons. For a closer investigation of this I(h) increase, we overexpressed the predominant layer 5 rat subunit HCN1 in HEK293 cells. We characterized the resulting I(h) in the cell-attached and whole-cell configurations. Breaking into whole-cell configuration led to about a 30% enhancement of rat HCN1-mediated I(h) accompanied by a depolarizing shift in voltage dependence and an accelerated time course of activation. This current enhancement is not species specific; for human HCN1, the current similarly increases in amount and kinetics. Although the changes were bound to cytosolic solution exchange, they were independent of cAMP, ATP, GTP, and the phosphate group donor phosphocreatine. Together, these data provide a characterization of heterologous expression of rat HCN1 and suggest that cytosolic contents suppress I(h). Such a mechanism might constitute a reserve in h-channel function in vivo. (c) 2010 Wiley-Liss, Inc.

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Aberrant stress-induced Hsp70 expression in immune cells in multiple sclerosis.

Publication Date: 2010 Aug 30 PMID: 20806409
Authors: Cwiklinska, H. - Mycko, M. P. - Szymanska, B. - Matysiak, M. - Selmaj, K. W.
Journal: J Neurosci Res

Heat shock protein 70 (Hsp70), a prominent member of the heat shock protein family, is a stress-induced chaperone, contributing to the "protein triage" mechanism. However, we and others have previously shown that chaperonin activity of Hsp70 also promotes immune recognition of protein/peptide antigens, including myelin autoantigens. Hsp70 has been strikingly elevated in multiple sclerosis (MS) lesions. In a search for the mechanism of Hsp70 up-regulation in MS, we analyzed Hsp70 expression in peripheral blood mononuclear cells (PBMCs) from MS patients (n = 49), healthy controls (n = 40), and patients with rheumatoid arthritis, (RA; n = 13). Hsp70 was detected by Western blot, and Hsp70 levels were quantified by ELISA. We found that Hsp70 was expressed at low levels in ex vivo PBMCs. However, after heat shock, Hsp70 was up-regulated significantly more (up to sixfold) in MS patients compared with healthy controls. This significant overproduction of Hsp70 was also seen following another stress condition, LPS stimulation. Hsp70 is a product of several independent genes, and we found the HSPA1B gene product to be the major form responsible for Hsp70 protein overexpression in PBMCs. Hsp70 overexpression was preceded by increased nuclear presence of heat shock factor 1 (HSF1). HSF1 activation depends on phosphorylation, and we found that inhibition of the A group of protein kinase C isoenzymes significantly reduced inducible Hsp70 production. These results indicate that immune cells from MS patients are more prone to Hsp70 induction under stress conditions, suggesting a possible link between Hsp70 overexpression and development of autoimmunity. (c) 2010 Wiley-Liss, Inc.

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L10p and P158DEL DJ-1 mutations cause protein instability, aggregation, and dimerization impairments.

Publication Date: 2010 Aug 30 PMID: 20806408
Authors: Ramsey, C. P. - Giasson, B. I.
Journal: J Neurosci Res

A variety of mutations in the gene encoding DJ-1 protein cause autosomal recessive early-onset parkinsonism. Recently, a novel pathogenic homozygous DJ-1 missense mutation resulting in the L10P amino acid substitution was reported. In a separate study, a novel homozygous mutation resulting in the deletion of DJ-1 residue P158 was also reported to be causative of disease. The specific effects of the novel L10P and P158DEL mutations on protein function have not been studied. Here, L10P and P158DEL DJ-1 proteins were assessed for protein stability, dimerization, solubility, subcellular localization, and protective function in comparison with WT and the L166P DJ-1 pathogenic variant. It was discovered that, compared with WT protein, L10P, L166P, and P158DEL DJ-1 variants exhibited dramatically reduced protein stabilities. Degradation of each of the pathogenic mutants appeared to be mediated in part by the proteasome. Interestingly, unlike L166P DJ-1, the L10P and P158DEL DJ-1 variants retained the ability to dimerize with WT DJ-1 protein; however, neither of these mutants was able to form homodimers. Additionally, the L10P, L166P, and P158DEL DJ-1 variants exhibited altered profiles on size-exclusion chromatography and demonstrated reduced solubilities in comparison with WT protein, and the latter aberration could be exacerbated in the presence of MG-132. Furthermore, cells stably expressing L10P DJ-1 were more vulnerable to treatments with proteasome inhibitors, suggesting that L10P DJ-1 may be toxic to cells under conditions of proteasome stress. Taken together, these findings suggest that diverse aberrant mechanisms, including alterations in protein stability and protein folding, are associated with the pathogenicity of the L10P and P158DEL DJ-1 variants. (c) 2010 Wiley-Liss, Inc.

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Localized alteration of microtubule polymerization in response to guidance cues.

Publication Date: 2010 Aug 30 PMID: 20806407
Authors: Kelly, T. A. - Katagiri, Y. - Vartanian, K. B. - Kumar, P. - Chen, I. I. - Rosoff, W. J. - Urbach, J. S. - Geller, H. M.
Journal: J Neurosci Res

Inhibition of microtubule dynamic instability prevents growth cone turning in response to guidance cues, yet specific changes in microtubule polymerization as growth cones encounter boundaries have not been investigated. In this study, we examined the rate and direction of microtubule polymerization in response to soluble nerve growth factor (NGF) and immobilized chondroitin sulfate proteoglycans (CSPGs) by expressing enhanced GFP-EB3 in rat pheochromocytoma (PC12) cells. GFP-EB3 comets were monitored in live cells using time-lapse epifluorescent microscopy. With an automated tracking system, the rate of microtubule polymerization was calculated as the frame-to-frame displacement of EB3 comets. Our results demonstrate that the rate of microtubule polymerization is increased following NGF treatment, whereas contact with CSPGs decreases microtubule polymerization rates. This reduction in microtubule polymerization rates was specifically localized to neurites in direct contact with CSPGs and not at noncontacting neurites. Additionally, we found an increase in the percentage of microtubules polymerizing in the retrograde direction in neurites at CSPG boundaries, with a concomitant decrease in the rate of retrograde microtubule polymerization. These results implicate localized changes in microtubule dynamics as an important component of the growth cone response to guidance cues. (c) 2010 Wiley-Liss, Inc.

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Reduction of hippocampal N-acetyl aspartate level in aged APP(Swe)/PS1(dE9) transgenic mice is associated with degeneration of CA3 pyramidal neurons.

Publication Date: 2010 Aug 30 PMID: 20806406
Authors: Xu, W. - Zhan, Y. - Huang, W. - Wang, X. - Zhang, S. - Lei, H.
Journal: J Neurosci Res

Age-related metabolic changes in the hippocampus of APP(Swe)/PS1(dE9) mice were measured with long echo-time in vivo (1)H-magnetic resonance spectroscopy ((1)H-MRS). Thioflavine S staining and Nissl staining were used to characterize deposition of Abeta aggregates and neuronal degeneration in the transgenic animals, respectively. The results showed that the APP(Swe)/PS1(dE9) mice had significantly decreased hippocampal N-acetyl aspartate (NAA)/total creatine (tCr) level at 16 months of age, which was associated with degeneration of and intracellular deposition of thioflavine S-positive materials in hippocampal CA3 pyramidal neurons. The results of this study provide direct evidence showing association among Abeta pathology (intracellular deposition of thioflavine S-positive materials), neuronal degeneration, and metabolic changes observable with in vivo (1)H-MRS in the hippocampus of APP(Swe)/PS1(dE9) mice. (c) 2010 Wiley-Liss, Inc.

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Phosphorylation of ERK in the spinal dorsal horn following pancreatic pronociceptive stimuli with proteinase-activated receptor-2 agonists and hydrogen sulfide in rats: Evidence for involvement of distinct mechanisms.

Publication Date: 2010 Aug 30 PMID: 20806405
Authors: Fukushima, O. - Nishimura, S. - Matsunami, M. - Aoki, Y. - Nishikawa, H. - Ishikura, H. - Kawabata, A.
Journal: J Neurosci Res

Noxious stimuli cause prompt phosphorylation of extracellular signal-regulated kinase (ERK) in the spinal dorsal horn that contributes to facilitation of pain sensation and is often used as an immediate marker for excitation of spinal neurons following somatic and colonic nociception. Here we asked whether two distinct pronociceptive stimuli with proteinase-activated receptor-2 (PAR2) agonists and hydrogen sulfide (H(2)S) in the pancreas cause phosphorylation of ERK in the spinal dorsal horn and also examined involvement of their possible downstream signaling molecules, transient receptor potential vanilloid-1 (TRPV1) and T-type Ca(2+) channels, respectively. Capsaicin (a TRPV1 agonist), trypsin (an endogenous PAR2 agonist), SLIGRL-NH(2) (a PAR2-activating peptide), and NaHS (an H(2)S donor) were infused into the pancreatic duct in anesthetized rats, and phosphorylated ERK in the spinal cord was detected by immunohistochemistry. Intraductal administration of capsaicin and trypsin caused prompt phosphorylation of ERK in the superficial layers of T9, but not T5 or T12, spinal dorsal horn. SLIGRL-NH(2) and NaHS, administered in the same manner, also produced ERK phosphorylation in the corresponding spinal regions. Mibefradil, a T-type Ca(2+) channel blocker, abolished the phosphorylation of ERK caused by intraductal NaHS but not SLIGRL-NH(2). In contrast, capsazepine, an inhibitor of TRPV1, suppressed the phosphorylation of ERK caused by intraductal SLIGRL-NH(2) but not NaHS. Our data thus demonstrate that pancreatic pronociceptive stimuli with PAR2 agonists and H(2)S cause ERK phosphorylation in the spinal dorsal horn, through activation of TRPV1 and T-type Ca(2+) channels, respectively, and that those two pronociceptive pathways are independent of each other. (c) 2010 Wiley-Liss, Inc.

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Activation of calcium/calmodulin-dependent protein kinase IV and peroxisome proliferator-activated receptor gamma coactivator-1alpha signaling pathway protects against neuronal injury and promotes mitochondrial biogenesis in the hippocampal CA1 subfield after transient global ischemia.

Publication Date: 2010 Aug 26 PMID: 20799369
Authors: Chen, S. D. - Lin, T. K. - Lin, J. W. - Yang, D. I. - Lee, S. Y. - Shaw, F. Z. - Liou, C. W. - Chuang, Y. C.
Journal: J Neurosci Res

Delayed neuronal cell death occurs in the vulnerable CA1 subfield of the hippocampus after transient global ischemia (TGI). We demonstrated previously, based on an experimental model of TGI, that the significantly increased content of oxidized proteins in hippocampal CA1 neuron was observed as early as 30 min after TGI, followed by augmentation of PGC-1alpha expression at 1 hr, as well as up-regulation of mitochondrial uncoupling protein 2 (UCP2) and superoxide dismutases 2 (SOD2). Using the same animal model, the present study investigated the role of calcium/calmodulin-dependent protein kinase IV (CaMKIV) and PGC-1alpha in delayed neuronal cell death and mitochondrial biogenesis in the hippocampus. In Sprague-Dawley rats, significantly increased expression of nuclear CaMKIV was noted in the hippocampal CA1 subfield as early as 15 min after TGI. In addition, the index of mitochondrial biogenesis, including a mitochondrial DNA-encoded polypeptide, cytochrome c oxidase subunit 1 (COX1), and mitochondrial number significantly increased in the hippocampal CA1 subfield 4 hr after TGI. Application bilaterally into the hippocampal CA1 subfield of an inhibitor of CaMKIV, KN-93, 30 min before TGI attenuated both CaMKIV and PGC-1alpha expression, followed by down-regulation of UCP2 and SOD2, decrease of COX1 expression and mitochondrial number, heightened protein oxidation, and enhanced hippocampal CA1 neuronal damage. This study provides correlative evidence for the neuroprotective cascade of CaMKIV/PGC-1alpha which implicates at least in part the mitochondrial antioxidants UCP2 and SOD2 as well as mitochondrial biogenesis in ischemic brain injury. (c) 2010 Wiley-Liss, Inc.

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Glutamate-induced calcium increase mediates magnesium release from mitochondria in rat hippocampal neurons.

Publication Date: 2010 Aug 25 PMID: 20740499
Authors: Shindo, Y. - Fujimoto, A. - Hotta, K. - Suzuki, K. - Oka, K.
Journal: J Neurosci Res

Excess administration of glutamate is known to induce Ca(2+) overload in neurons, which is the first step in excitotoxicity. Although some reports have suggested a role for Mg(2+) in the excitotoxicity, little is known about its actual contribution. To investigate the role of Mg(2+) in the excitotoxicity, we simultaneously measured intracellular Ca(2+) and Mg(2+), using fluorescent dyes, Fura red, a fluorescent Ca(2+) probe, and KMG-104, a highly selective fluorescent Mg(2+) probe developed by our group, respectively. Administration of 100 muM glutamate supplemented with 10 muM glycine to rat hippocampal neurons induced an increase in intracellular Mg(2+) concentration ([Mg(2+)](i)). Extracellular Mg(2+) was not required for this glutamate-induced increase in [Mg(2+)](i), and no increase in intracellular Ca(2+) concentration ([Ca(2+)](i)) or [Mg(2+)](i) was observed in neurons in nominally Ca(2+)-free medium. Application of 5 muM carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone (FCCP), an uncoupler of mitochondrial inner membrane potential, also elicited increases in [Ca(2+)](i) and [Mg(2+)](i). Subsequent administration of glutamate and glycine following FCCP treatment did not induce a further increase in [Mg(2+)](i) but did induce an additive increase in [Ca(2+)](i). Moreover, the glutamate-induced increase in [Mg(2+)](i) was observed only in mitochondria localized areas. These results support the idea that glutamate is able to induced Mg(2+) efflux from mitochondria to the cytosol. Furthermore, pretreatment with Ru360, an inhibitor of the mitochondrial Ca(2+) uniporter, prevented this [Mg(2+)](i) increase. These results indicate that glutamate-induced increases in [Mg(2+)](i) result from the Mg(2+) release from mitochondria and that Ca(2+) accumulation in the mitochondria is required for this Mg(2+) release. (c) 2010 Wiley-Liss, Inc.

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Opioid system functional regulation in neurological disease management.

Publication Date: 2010 Aug 23 PMID: 20734417
Authors: Nandhu, M. S. - Naijil, G. - Smijin, S. - Jayanarayanan, S. - Paulose, C. S.
Journal: J Neurosci Res

There is increasing evidence to suggest a role for the opioid system in the control of pathophysiology of neurological disorders (Alzheimer's, Parkinson's, and Huntington's diseases, spinal cord injury, epilepsy, hypoxia, and autism). Resuscitation of the altered expression of the opioid system in various neurological disorders is of therapeutic importance. Such treatment may be beneficial in ameliorating the clinical symptoms of the disorder. This Mini-Review provides a brief update on opioid system regulation in neurological disorders and focuses on the opioids' pharmacological importance. (c) 2010 Wiley-Liss, Inc.

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The H63D HFE gene variant promotes activation of the intrinsic apoptotic pathway via mitochondria dysfunction following beta-amyloid peptide exposure.

Publication Date: 2010 Aug 23 PMID: 20734416
Authors: Mairuae, N. - Hall Ii, E. C. - Cheepsunthorn, P. - Lee, S. Y. - Connor, J. R.
Journal: J Neurosci Res

Numerous epidemiological studies suggest that the expression of the HFE allelic variant H63D may be a risk factor or genetic modifier for Alzheimer's disease (AD). The H63D variant alters cellular iron homeostasis and increases baseline oxidative stress. The elevated cellular stress milieu, we have proposed, may alter cellular responses to genetic and environmental determinants of AD. Accumulation of beta-amyloid peptides (Abeta) is one of the most prominent pathogenic characteristics of AD. Several studies have demonstrated that Abeta can induce neuronal cell death through apoptosis. In this study, we provide evidence that an Abeta(25-35) fragment, which contains the cytotoxic sequence of the amyloid peptide, activates the intrinsic apoptotic pathway in SH-SY5Y human neuroblastoma cells expressing the HFE allelic variant H63D to a greater extent than in cells with wild-type (WT) HFE. Specifically, Abeta(25-35) peptide exposure significantly induced Bax translocation from the cytosol to the mitochondria in H63D-expressing cells compared with WT cells. This translocation was associated with increased cytochrome c release from mitochondria and an increase in active caspase-9 and caspase-3 activity in H63D cells. Consequently, there is increased apoptosis in cells expressing the H63D variant as opposed to cells expressing WT HFE. We also found increased amyloid precursor protein (APP) and Abeta(1-42) peptide in the mitochondrial compartment as well as increased mitochondrial stress in H63D-expressing cells compared with WT. These findings support our hypothesis that the presence of the HFE H63D allele enables factors that trigger neurodegenerative processes associated with AD and predisposes cells to cytotoxcity. (c) 2010 Wiley-Liss, Inc.

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Environmental enrichment potentiates thalamocortical transmission and plasticity in the adult rat visual cortex.

Publication Date: 2010 Aug 18 PMID: 20722076
Authors: Mainardi, M. - Landi, S. - Gianfranceschi, L. - Baldini, S. - De Pasquale, R. - Berardi, N. - Maffei, L. - Caleo, M.
Journal: J Neurosci Res

It has been demonstrated that the complex sensorimotor and social stimulation achieved by rearing animals in an enriched environment (EE) can reinstate juvenile-like plasticity in the adult cortex. However, it is not known whether EE can affect thalamocortical transmission. Here, we recorded in vivo field potentials from the visual cortex evoked by electrical stimulation of the dorsal lateral geniculate nucleus (dLGN) in anesthetized rats. We found that a period of EE during adulthood shifted the input-output curves and increased paired-pulse depression, suggesting an enhanced synaptic strength at thalamocortical terminals. Accordingly, EE animals showed an increased expression of the vesicular glutamate transporter 2 (vGluT-2) in geniculocortical afferents to layer IV. Rats reared in EE also showed an enhancement of thalamocortical long-term potentiation (LTP) triggered by theta-burst stimulation (TBS) of the dLGN. To monitor the functional consequences of increased LTP in EE rats, we recorded visual evoked potentials (VEPs) before and after application of TBS to the geniculocortical pathway. We found that responses to visual stimulation were enhanced across a range of contrasts in EE animals. This was accompanied by an up-regulation of the intracortical excitatory synaptic marker vGluT-1 and a decrease in the expression of the vesicular GABA transporter (vGAT), indicating a shift in the excitation/inhibition ratio. Thus, in the adult rat, EE enhances synaptic strength and plasticity of the thalamocortical pathway associated with specific changes in glutamatergic and GABAergic neurotransmission. These data provide novel insights into the mechanisms by which EE shapes the adult brain. (c) 2010 Wiley-Liss, Inc.

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Regional and cellular distribution of mitochondrial ferritin in the mouse brain.

Publication Date: 2010 Aug 18 PMID: 20722075
Authors: Snyder, A. M. - Neely, E. B. - Levi, S. - Arosio, P. - Connor, J. R.
Journal: J Neurosci Res

Iron and mitochondrial dysfunction are important in many neurodegenerative diseases. Several iron transport proteins have been identified that are associated with mitochondria, most recently mitochondrial ferritin. Here we describe the cellular distribution of mitochondrial ferritin in multiple regions of the brain in C57/BL6 mice. Mitochondrial ferritin was found in all regions of the brain, although staining intensity varied between regions. Mitochondrial ferritin was detected throughout the layers of cerebral cortex and in the cerebellum, hippocampus, striatum, choroid plexus, and ependymal cells. The cell type in the brain that stains most prominently for mitochondrial ferritin is neuronal, but oligodendrocytes also stain strongly in both gray matter and in white matter tracts. Mice deficient in H-ferritin do not differ in the mitochondrial ferritin staining pattern or intensity compared with C57/BL6 mice, suggesting that there is no compensatory expression of these proteins. In addition, by using inbred mouse strains with differing levels of iron content, we have shown that regional brain iron content does not affect expression of mitochondria ferritin. The expression of mitochondria ferritin appears to be more influenced by mitochondrial density. Indeed, at an intracellular level, mitochondrial ferritin immunoreaction product is strongest where mitochondrial density is high, as seen in the ependymal cells. Given the importance and relationship between iron and mitochondrial activity, understanding the role of mitochondrial ferritin can be expected to contribute to our knowledge of mitochondrial dysfunction and neurodegenerative disease. (c) 2010 Wiley-Liss, Inc.

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Wnt-3a and Wnt-3 differently stimulate proliferation and neurogenesis of spinal neural precursors and promote neurite outgrowth by canonical signaling.

Publication Date: 2010 Aug 18 PMID: 20722074
Authors: David, M. D. - Canti, C. - Herreros, J.
Journal: J Neurosci Res

Wnt factors regulate neural stem cell development and neuronal connectivity. Here we investigated whether Wnt-3a and Wnt-3, expressed in the developing spinal cord, regulate proliferation and the neuronal differentiation of spinal cord neural precursors (SCNP). Wnt-3a promoted a sustained increase of SCNP proliferation and decreased the expression of cyclin-dependent kinase inhibitors. In contrast, Wnt-3 transiently enhanced SCNP proliferation and increased neurogenesis through beta-catenin signaling. Furthermore, both Wnt-3a and Wnt-3 stimulated neurite outgrowth in SCNP-derived neurons through beta-catenin- and TCF4-dependent transcription. Glycogen synthase kinase-3beta inhibitors mimicked Wnt signaling and promoted neurite outgrowth in established cultures. We conclude that Wnt-3a and Wnt-3 factors signal through the canonical Wnt/beta-catenin pathway to regulate different aspects of SCNP development. These findings may be of therapeutic interest for the treatment of neurodegenerative diseases and nerve injury. (c) 2010 Wiley-Liss, Inc.

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Different effects of erythropoietin in cisplatin- and docetaxel-induced neurotoxicity: An in vitro study.

Publication Date: 2010 Aug 18 PMID: 20722073
Authors: Maggioni, D. - Nicolini, G. - Chiorazzi, A. - Meregalli, C. - Cavaletti, G. - Tredici, G.
Journal: J Neurosci Res

Chemotherapy-induced peripheral neurotoxicity (CIPN) is a side effect limiting cisplatin (CDDP) and docetaxel (DOCE) treatment. Erythropoietin (EPO) is a hematopoietic growth factor also displaying neurotrophic properties. Evidence suggests that EPO's neuroprotective action may rely on PI3K/AKT pathway activation; however, data regarding the EPO neuroprotective mechanism are still limited. This study evaluated the effect of EPO on organotypic cultures of rat dorsal root ganglia (DRG) and in primary cultures of DRG-dissociated sensory neurons exposed to CDDP- and DOCE-induced neurotoxicity, aiming to investigate EPO's neuroprotective mechanism. Subsequently, the levels of AKT expression and activation were analyzed by Western blot in neurons exposed to CDDP or DOCE; AKT's role was further evaluated by using a chemical inhibitor of AKT activation, wortmannin. In these models EPO, was protective against both CDDP- and DOCE-induced cell death and against CDDP-induced neurite elongation reduction. A modulation of AKT activation was observed in CDDP-treated neurons, and the presence of wortmannin prevented EPO's neuroprotective action against CDDP toxicity but did not have any effect on EPO's protection against DOCE-induced toxicity, thus ruling out the PI3K-AKT pathway as the mechanism of EPO's effect in neuronal death prevention after DOCE exposure. Our results confirm in vitro the effectiveness of EPO as a neuroprotectant against both CDDP- and DOCE-induced neurotoxicity. In addition, a role of PI3K/AKT in EPO's protection against CDDP, but not against DOCE, neurotoxicity was shown, suggesting that alternative pathways could be involved in EPO's neuroprotective activity. (c) 2010 Wiley-Liss, Inc.

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Origin of quantal size variation and high-frequency miniature postsynaptic currents at the Caenorhabditis elegans neuromuscular junction.

Publication Date: 2010 Aug 18 PMID: 20722072
Authors: Wang, Z. W.
Journal: J Neurosci Res

The neuromuscular junction (NMJ) of Caenorhabditis elegans has proved to be a very useful model synapse for investigating molecular mechanisms of synaptic transmission. Intriguingly, miniature postsynaptic currents (minis) at this synapse occur at an unusually high frequency (50-90 Hz in wild-type worms) and show large variation in quantal size (from <10 pA to >200 pA). It is important to understand the cellular and molecular bases for these properties of minis in order to interpret electrophysiological data from this synapse properly. Existing data suggest that several factors may contribute to the high frequency and quantal size variation, including 1) the establishment of multiple NMJs with each body-wall muscle cell, 2) diversity of postsynaptic receptors (two acetylcholine receptors and one GABA receptor), 3) association of one presynaptic site with several body-wall muscle cells, 4) effects of Ca(2+) at the presynaptic site, and 5) a possibly elevated (less negative) resting membrane potential in motoneurons. Neither the frequency nor the quantal size of minis is affected by electrical coupling of body-wall muscle cells. Furthermore, quantal size variation is not due to synchronized multivesicular release. Analyses of the C. elegans NMJ may lead to a better understanding of the mechanisms controlling the frequency and quantal size of minis of other synapses as well. (c) 2010 Wiley-Liss, Inc.

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Cerebral tissue repair and atrophy after embolic stroke in rat: A magnetic resonance imaging study of erythropoietin therapy.

Publication Date: 2010 Aug 18 PMID: 20722071
Authors: Ding, G. - Jiang, Q. - Li, L. - Zhang, L. - Wang, Y. - Zhang, Z. G. - Lu, M. - Panda, S. - Li, Q. - Ewing, J. R. - Chopp, M.
Journal: J Neurosci Res

Using magnetic resonance imaging (MRI) protocols of T(2)-, T(2)*-, diffusion- and susceptibility-weighted imaging (T2WI, T2*WI, DWI, and SWI, respectively) with a 7T system, we tested the hypothesis that treatment of embolic stroke with erythropoietin (EPO) initiated at 24 hr and administered daily for 7 days after stroke onset has benefit in repairing ischemic cerebral tissue. Adult Wistar rats were subjected to embolic stroke by means of middle cerebral artery occlusion (MCAO) and were randomly assigned to a treatment (n = 11) or a control (n = 11) group. The treated group was given EPO intraperitoneally at a dose of 5,000 IU/kg daily for 7 days starting 24 hr after MCAO. Controls were given an equal volume of saline. MRI was performed at 24 hr and then weekly for 6 weeks. MRI and histological measurements were compared between groups. Serial T2WI demonstrated that expansion of the ipsilateral ventricle was significantly reduced in the EPO-treated rats. The volume ratio of ipsilateral parenchymal tissue relative to the contralateral hemisphere was significantly increased after EPO treatment compared with control animals, indicating that EPO significantly reduces atrophy of the ipsilateral hemisphere, although no significant differences in ischemic lesion volume were observed between the two groups. Angiogenesis and white matter remodeling were significantly increased and occurred earlier in EPO-treated animals than in the controls, as evident from T2*WI and diffusion anisotropy maps, respectively. These data indicate that EPO treatment initiated 24 hr poststroke promotes angiogenesis and axonal remodeling in the ischemic boundary, which may potentially reduce atrophy of the ipsilateral hemisphere. (c) 2010 Wiley-Liss, Inc.

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Sex differences in cerebral ischemia: possible molecular mechanisms.

Publication Date: 2010 Oct PMID: 20698025
Authors: Siegel, C. - Turtzo, C. - McCullough, L. D.
Journal: J Neurosci Res

Sex is emerging as an important factor in the etiology and expression of many different pathological conditions, including stroke. Initially, the levels of sex hormones were thought to be the major contributor to these sex differences, especially after puberty, when gonadal steroid levels sharply diverge between the sexes. More recently, it is recognized that sex differences also result from the organizational effects of sex hormone exposure early in development, even in the absence of hormone exposure later in life, as well as effects mediated by the sex chromosomes themselves. Epigenetic modifications of developmental genes important in sexual differentiation and the response to sex steroid hormones are also emerging as another important contributor to sex differences in disease expression. This review describes recent research on the relationship between hormones, organizational-activational effects of gonadal steroids, and epigenetic modifications in brain pathology, focusing specifically on cerebral ischemia.

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Cathepsin B contributes to traumatic brain injury-induced cell death through a mitochondria-mediated apoptotic pathway.

Publication Date: 2010 Oct PMID: 20653046
Authors: Luo, C. L. - Chen, X. P. - Yang, R. - Sun, Y. X. - Li, Q. Q. - Bao, H. J. - Cao, Q. Q. - Ni, H. - Qin, Z. H. - Tao, L. Y.
Journal: J Neurosci Res

It has been reported that lysosomal proteases play important roles in ischemic and excitotoxic neuronal cell death. We have previously reported that cathepsin B expression increased remarkably after traumatic brain injury (TBI). The present study sought to investigate the effects of a selective cathepsin B inhibitor (CBI) [N-L-3-trans-prolcarbamoyloxirane-2-carbonyl)-L-isoleucyl-L-proline] on cell death and behavioral deficits in our model. We examined the levels of cathepsin B enzymatic activity and its expression by double labelling damaged cells in the brain slice with propidium iodide (PI) and anticathepsin B. The results showed an elevated enzymatic activity associated with TBI-induced increase in a mature form of cathepsin B, suggesting that cathepsin B may play a role in TBI-induced cell injury. PI was found to label cells positive for the neuronal-specific nuclear marker NeuN, whereas fewer GFAP-positive cells were labelled by PI, suggesting that neurons are more sensitive to cell death induced by TBI. Additionally, we found that pretreatment with CBI remarkably attenuated TBI-induced cell death, lesion volume, and motor and cognitive dysfunction. To analyze the mechanism of action of cathepsin B in the cell death signaling pathway, we assessed DNA fragmentation by electrophoresis, Bcl-2/Bax protein expression levels, Bid cleavage, cytochrome c release, and caspase-3 activation. The results imply that cathepsin B contributes to TBI-induced cell death through the present programmed cell necrosis and mitochondria-mediated apoptotic pathways.

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Increased expression of urokinase-type plasminogen activator receptor in the frontal cortex of patients with intractable frontal lobe epilepsy.

Publication Date: 2010 Sep PMID: 20648659
Authors: Liu, B. - Zhang, B. - Wang, T. - Liang, Q. C. - Jing, X. R. - Zheng, J. - Wang, C. - Meng, Q. - Wang, L. - Wang, W. - Guo, H. - You, Y. - Zhang, H. - Gao, G. D.
Journal: J Neurosci Res

Urokinase-type plasminogen activator receptor (uPAR) is a glycosyl phosphatidylinositol-anchored protein involved in cell adhesion, proliferation, differentiation, migration, invasion, and tissue repair and remodeling. Our aim was to investigate uPAR expression in the frontal cortex of patients with intractable frontal lobe epilepsy and to explore the possible role of uPAR in intractable epilepsy. Tissue samples were obtained from the frontal cortex of 25 patients who had undergone surgery for intractable epilepsy and 15 histologically normal frontal cortex tissues from patients with orbital frontal lobe severe contusion (the control group). The frontal cortex expression of uPAR was studied by Western blot and immnohistochemistry. Double immunofluorescence was used to determine the expression of uPAR in astrocytes, microglia, and neurons. The normal frontal cortex uPAR protein level was shown to be low. In the brain tissue of patients with intractable epilepsy, the expression of uPAR protein increased dramatically. Based on the results of double immunofluorescence, many uPAR-positive cells are colocalized with the cell soma of NeuN-positive neurons, whereas only a few GFAP- and CD11b-positive cells colocalized with uPAR staining. These findings provide new information pertaining to the epileptogenesis of intractable epilepsy and suggest that increased expression of uPAR in human brain may be associated with human intractable epilepsy.

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Appearance of phagocytic microglia adjacent to motoneurons in spinal cord tissue from a presymptomatic transgenic rat model of amyotrophic lateral sclerosis.

Publication Date: 2010 Sep PMID: 20648658
Authors: Sanagi, T. - Yuasa, S. - Nakamura, Y. - Suzuki, E. - Aoki, M. - Warita, H. - Itoyama, Y. - Uchino, S. - Kohsaka, S. - Ohsawa, K.
Journal: J Neurosci Res

Microglial activation occurs early during the pathogenesis of amyotrophic lateral sclerosis (ALS). Recent evidence indicates that the expression of mutant Cu(2+)/Zn(2+) superoxide dismutase 1 (SOD1) in microglia contributes to the late disease progression of ALS. However, the mechanism by which microglia influence the neurodegenerative process and disease progression in ALS remains unclear. In this study, we revealed that activated microglia aggregated in the lumbar spinal cord of presymptomatic mutant SOD1(H46R) transgenic rats, an animal model of familial ALS. The aggregated microglia expressed a marker of proliferating cell, Ki67, and phagocytic marker proteins ED1 and major histocompatibility complex (MHC) class II. The motoneurons near the microglial aggregates showed weak choline acetyltransferase (ChAT) immunoreactivity and contained reduced granular endoplasmic reticulum and altered nucleus electron microscopically. Furthermore, immunopositive signals for tumor necrosis factor-alpha (TNFalpha) and monocyte chemoattractant protein-1 (MCP-1) were localized in the aggregated microglia. These results suggest that the activated and aggregated microglia represent phagocytic features in response to early changes in motoneurons and possibly play an important role in ALS disease progression during the presymptomatic stage.

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Peripheral purinergic receptor modulation influences the trigeminal ganglia nitroxidergic system in an experimental murine model of inflammatory orofacial pain.

Publication Date: 2010 Sep PMID: 20648657
Authors: Borsani, E. - Albertini, R. - Labanca, M. - Lonati, C. - Rezzani, R. - Rodella, L. F.
Journal: J Neurosci Res

ATP plays an important role as an endogenous pain mediator generating and/or modulating pain signaling from the periphery to the central nervous system. The aim of this study was to analyze the role of peripheral purinergic receptors in modulation of the nitroxidergic system at a trigeminal ganglia level by monitoring changes in nitric oxide synthase isoforms. We also evaluated Fos-positive neurons in brainstem (spinal trigeminal nucleus) and pain-related behavior. We found that local administration of the P2 purinergic receptor antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid (PPADS) decreased face-rubbing activity, nitric oxide synthase isoform expression in trigeminal ganglia, and Fos expression in spinal trigeminal nucleus after subcutaneous injection of formalin. These results suggest a role for peripheral P2 purinergic receptors in orofacial pain transmission through modulation of the nitroxidergic system. .

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Nuclear factor-kappaB/p65 responds to changes in the Notch signaling pathway in murine BV-2 cells and in amoeboid microglia in postnatal rats treated with the gamma-secretase complex blocker DAPT.

Publication Date: 2010 Sep PMID: 20648656
Authors: Cao, Q. - Li, P. - Lu, J. - Dheen, S. T. - Kaur, C. - Ling, E. A.
Journal: J Neurosci Res

Microglial cells constitutively express Notch-1 and nuclear factor-kappaB/p65 (NF-kappaB/p65), and both pathways modulate production of inflammatory mediators. This study sought to determine whether a functional relationship exists between them and, if so, to investigate whether they synergistically regulate common microglial cell functions. By immunofluorescence labeling, real-time polymerase chain reaction (RT-PCR), flow cytometry, and Western blot, BV-2 cells exhibited Notch-1 and NF-kappaB/p65 expression, which was significantly up-regulated in cells challenged with lipopolysaccharide (LPS). This was coupled with an increase in expression of Hes-1, tumor necrosis factor-alpha (TNF-alpha), and interleukin-1beta (IL-1beta). In BV-2 cells pretreated with N-[N-(3,5-difluorophenacetyl)-1-alany1]-S-phenyglycine t-butyl ester (DAPT), a gamma-secretase inhibitor, followed by LPS stimulation, Notch-1 expression level was enhanced but that of all other markers was suppressed. Additionally, Hes-1 expression and NF-kappaB nuclear translocation decreased as shown by flow cytometry. Notch-1's modulation of NF-kappaB/p65 was also evidenced in amoeboid microglial cells (AMC) in vivo. In 5-day-old rats given intraperitoneal injections of LPS, Notch-1, NF-kappaB/p65, TNF-alpha, and IL-1beta immunofluorescence in AMC was markedly enhanced. However, in rats given an intraperitoneal injection of DAPT prior to LPS, Notch-1 labeling was augmented, but that of TNF-alpha and IL-1beta was reduced. The results suggest that blocking of Notch-1 activation with DAPT would reduce the level of its downstream end product Hes-1 along with suppression of NF-kappaB/p65 translocation, resulting in suppressed production of proinflammatory cytokines. It is concluded that Notch-1 signaling can trans-activate NF-kappaB/p65 by amplifying NF-kappaB/p65-dependent proinflammatory functions in activated microglia.

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Sensitive and specific detection of alpha-synuclein in human plasma.

Publication Date: 2010 Sep PMID: 20648655
Authors: Tinsley, R. B. - Kotschet, K. - Modesto, D. - Ng, H. - Wang, Y. - Nagley, P. - Shaw, G. - Horne, M. K.
Journal: J Neurosci Res

alpha-Synuclein (alphasyn) mutations, overexpression, misfolding, and aggregation are associated with Parkinson's disease. This protein has been intensively studied in neuronal systems. However, alphasyn is also present in extracellular fluids, such as cerebrospinal fluid and blood plasma. Recent studies have attempted to quantify its levels and compare these in various extracellular fluids of control and Parkinson's disease subjects. Data from these studies have been difficult to interpret, suggesting that more sensitive, standardized, and well-characterized assays of larger cohorts are required. Here, we describe the development of a new ELISA specifically for quantifying alphasyn in human plasma. An initial assay, using a commercial anti-alphasyn monoclonal antibody (211; Santa Cruz Biotechnology, Santa Cruz, CA) and based on a published protocol, was adapted for use in human plasma. In addition, we have developed a novel alphasyn-specific antibody for the assay that has very high sensitivity and signal:noise characteristics. Assays with either antibody showed high specificity for alphasyn, and detected it in a variety of sample types, including plasma. These assays can now be employed on large cohorts of patients and control subjects to determine whether plasma levels are altered in disease. Although measuring extracellular alphasyn levels may prove to be a useful biomarker of Parkinson's disease, it should also be a powerful tool for basic research aimed at understanding the normal and pathological physiology of alphasyn secretion. .

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Investigation of annexin A5 as a biomarker for Alzheimer's disease using neuronal cell culture and mouse model.

Publication Date: 2010 Sep PMID: 20648654
Authors: Yamaguchi, M. - Kokai, Y. - Imai, S. - Utsumi, K. - Matsumoto, K. - Honda, H. - Mizue, Y. - Momma, M. - Maeda, T. - Toyomasu, S. - Ito, Y. M. - Kobayashi, S. - Hashimoto, E. - Saito, T. - Sohma, H.
Journal: J Neurosci Res

Alzheimer's disease (AD) differs from other forms of dementia in its relation to amyloid beta peptide (Abeta). Abeta, a proteolytic product of amyloid precursor proteins (APP), has a toxic effect on neuronal cells, which involves perturbation of their Ca(2+) homeostasis. This effect implies that changes of protein expression in neuronal cells with calcium stress should provide a molecular marker for this disease. In the present study, we used the supernatant from a neuronal cell culture after incubation with or without Abeta and isolated a Ca(2+)-dependent acidic phospholipid binding fraction to perform a proteomic study. Several unique proteins were identified after incubation with Abeta. We focused on annexin A5, among these proteins, because it binds both Ca(2+) and lipids likely to be involved in calcium homeostasis. Tg2576 transgenic mice (AD model) overexpressing mutant human APP showed a significant increase of annexin A5 in the brain cortex but not in other organs, including liver, kidney, lung, and intestine. In human plasma samples, the level of annexin A5 was significantly increased in a proportion of AD patients compared with a control group (P < 0.0001 in the logistic regression analysis). From the receiver operating characteristic (ROC) curve with plasma annexin A5 concentrations, the mean area under the curve (AUC 0.898) suggests that annexin A5 is a favorable marker for AD.

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BDNF-Akt-Bcl2 antiapoptotic signaling pathway is compromised in the brain of autistic subjects.

Publication Date: 2010 Sep PMID: 20648653
Authors: Sheikh, A. M. - Malik, M. - Wen, G. - Chauhan, A. - Chauhan, V. - Gong, C. X. - Liu, F. - Brown, W. T. - Li, X.
Journal: J Neurosci Res

Although the pathogenesis of autism is not understood, emerging evidence points to apoptotic mechanisms being involved in this disorder. However, it is not known whether apoptosis signaling is deregulated in the brain of autistic subjects. This study investigates how the apoptosis-related proteins are regulated in the autistic brain. Our studies show that Bcl2 is significantly decreased, whereas the expression of p53 is increased, in the brain of autistic subjects in comparison with age-matched controls. We also found that the expression and phosphorylation/activation of Akt kinase that regulates Bcl2 are significantly decreased in the autistic brain. The down-regulation of Akt may result from a decreased concentration of brain-derived neurotrophic factor (BDNF), the growth factor that modulates Akt activities. These results suggest that down-regulation of the BDNF-Akt-Bcl2 antiapoptotic signaling pathway in the autistic brain could be one of the underlying mechanisms responsible for the pathogenesis of autism.

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Potential in vivo amelioration by N-acetyl-L-cysteine of oxidative stress in brain in human double mutant APP/PS-1 knock-in mice: toward therapeutic modulation of mild cognitive impairment.

Publication Date: 2010 Sep PMID: 20648652
Authors: Huang, Q. - Aluise, C. D. - Joshi, G. - Sultana, R. - St Clair, D. K. - Markesbery, W. R. - Butterfield, D. A.
Journal: J Neurosci Res

Alzheimer's disease (AD) is the most prevalent form of dementia among the elderly. Although the underlying cause has yet to be established, numerous data have shown that oxidative stress is implicated in AD as well as in preclinical stages of AD, such as mild cognitive impairment (MCI). The oxidative stress observed in brains of subjects with AD and MCI may be due, either fully or in part, to increased free radicals mediated by amyloid-beta peptide (Abeta). By using double human mutant APP/PS-1 knock-in mice as the AD model, the present work demonstrates that the APP/PS-1 double mutation results in elevated protein oxidation (as indexed by protein carbonyls), protein nitration (as indexed by 3-nitrotyrosine), as well as lipid peroxidation (as indexed by protein-bound 4-hydroxy-2-nonenal) in brains of mice aged 9 months and 12 months. APP/PS-1 mice also exhibited lower levels of brain glutathione peroxidase (GPx) in both age groups studied, whereas glutathione reductase (GR) levels in brain were unaffected by the mutation. The activities of both of these antioxidant enzymes were significantly decreased in APP/PS-1 mouse brains, whereas the activity of glucose-6-phosphate dehydrogenase (G6PDH) was increased relative to controls in both age groups. Levels of peptidyl prolyl isomerase 1 (Pin1) were significantly decreased in APP/PS-1 mouse brain aged 9 and 12 months. Administration of N-acetyl-L-cysteine (NAC), a glutathione precursor, to APP/PS-1 mice via drinking water suppressed increased protein oxidation and nitration and also significantly augmented levels and activity of GPx in brain from both age groups. Oral administration of NAC also increased the diminished activity of GR and protected against lipid peroxidation in brains of 9-month-old APP/PS-1 mice only. Pin1 levels, GR levels, and G6PDH activity in brain were unaffected by oral administration of NAC in both age groups. These results are discussed with reference to the therapeutic potential of this brain-accessible glutathione precursor in the treatment of MCI and AD.

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Neuroserpin regulates the density of dendritic protrusions and dendritic spine shape in cultured hippocampal neurons.

Publication Date: 2010 Sep PMID: 20648651
Authors: Borges, V. M. - Lee, T. W. - Christie, D. L. - Birch, N. P.
Journal: J Neurosci Res

Neuroserpin is a member of the serpin superfamily that is expressed principally in neurons of the central and peripheral nervous systems. Neuroserpin's spatial-temporal expression during development and in the adult brain suggests possible roles in synaptogenesis and synaptic plasticity. This is supported by behavioral changes in transgenic mice overexpressing neuroserpin. We have used an embryonic rat primary hippocampal neuron culture model to investigate whether neuroserpin can regulate elements of synaptic morphology that may be involved in these changes in cognitive function. Neuroserpin localized to axonal and dendritic compartments in cultured neurons and accumulated in synapsin-positive presynaptic terminals. Increased expression of neuroserpin resulted in an increase in the density of dendritic protrusions and alterations in dendritic spine shape. Our results identify neuroserpin as a new regulator of structural plasticity and suggest a cellular mechanism that may contribute to neuroserpin's effects on cognition.

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Role of adenosine kinase in cochlear development and response to noise.

Publication Date: 2010 Sep PMID: 20648650
Authors: Vlajkovic, S. M. - Guo, C. X. - Dharmawardana, N. - Wong, A. C. - Boison, D. - Housley, G. D. - Thorne, P. R.
Journal: J Neurosci Res

Adenosine signalling has an important role in cochlear protection from oxidative stress. In most tissues, intracellular adenosine kinase (ADK) is the primary route of adenosine metabolism and the key regulator of intracellular and extracellular adenosine levels. The present study provides the first evidence for ADK distribution in the adult and developing rat cochlea. In the adult cochlea, ADK was localized to the nuclear or perinuclear region of spiral ganglion neurons, lateral wall tissues, and epithelial cells lining scala media. In the developing cochlea, ADK was strongly expressed in multiple cell types at birth and reached its peak level of expression at postnatal day 21 (P21). Ontogenetic changes in ADK expression were evident in the spiral ganglion, organ of Corti, and stria vascularis. In the spiral ganglion, ADK showed a shift from predominantly satellite cell immunolabelling at P1 to neuronal expression from P14 onward. In contrast to the role of ADK in various aspects of cochlear development, the ADK contribution to the cochlear response to noise stress was less obvious. Transcript and protein levels of ADK were unaltered in the cochlea exposed to broadband noise (90-110 dBSPL, 24 hr), and the selective inhibition of ADK in the cochlea with ABT-702 failed to restore hearing thresholds after exposure to traumatic noise. This study indicates that ADK is involved in purine salvage pathways for nucleotide synthesis in the adult cochlea, but its role in the regulation of adenosine signalling under physiological and pathological conditions has yet to be established.

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Alzheimer's disease modifies progenitor cell expression of monoamine oxidase B in the subventricular zone.

Publication Date: 2010 Sep PMID: 20648649
Authors: Pugliese, M. - Rodriguez, M. J. - Gimeno-Bayon, J. - Pujadas, L. - Billett, E. E. - Wells, C. - Mahy, N.
Journal: J Neurosci Res

In the adult brain, progenitor cells remaining in the subventricular zone (SVZ) are frequently identified as glial fibrillary acidic protein (GFAP)-positive cells that retain attributes reminiscent of radial glia. Because the very high expression of monoamine oxidase B (MAO-B) in the subventricular area has been related to epithelial and astroglial expression, we sought to ascertain whether it was also expressed by progenitor cells of human control and Alzheimer's disease (AD) patients. In the SVZ, epithelial cells and astrocyte-like cells presented rich MAO-B activity and immunolabeling. Nestin-positive cells were found in the same area, showing a radial glia-like morphology. When coimmunostaining and confocal microscopy were performed, most nestin-positive cells showed MAO-B activity and labeling. The increased progenitor activity in SVZ proposed for AD patients was confirmed by the positive correlation between the SVZ nestin/MAO-B ratio and the progression of the disease. Nestin/GFAP-positive cells, devoid of MAO-B, can represent a distinct subpopulation of an earlier phase of maturation. This would indicate that MAO-B expression takes place in a further step of nestin/GFAP-positive cell differentiation. In the early AD stages, the discrete MAO-B reduction, different from the severe GFAP decrease, would reflect the capacity of this population of MAO-B-positive progenitor cells to adapt to the neurodegenerative process.

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Short-term low-frequency electrical stimulation enhanced remyelination of injured peripheral nerves by inducing the promyelination effect of brain-derived neurotrophic factor on Schwann cell polarization.

Publication Date: 2010 Sep PMID: 20648648
Authors: Wan, L. - Zhang, S. - Xia, R. - Ding, W.
Journal: J Neurosci Res

Electrical stimulation (ES) has been found to aid repair of nerve injuries and have been shown to increase and direct neurite outgrowth during stimulation. However, the effect of ES on peripheral remyelination after nerve damage has been investigated less well, and the mechanism underlying its action remains unclear. In the present study, the crush-injured sciatic nerves in rats were subjected to 1 hr of continuous ES (20 Hz, 100 microsec, 3 V). Electron microscopy and nerve morphometry were performed to investigate the extent of regenerated nerve myelination. The expression profiles of P0, Par-3, and brain-derived neurotrophic factor (BDNF) in the injuried sciatic nerves and in the dorsal root ganglion neuron/Schwann cell cocultures were examined by Western blotting. Par-3 localization in the sciatic nerves was determined by immunohistochemistry to demonstrate Schwann cell polarization during myelination. We reported that 20-Hz ES increased the number of myelinated fibers and the thickness myelin sheath at 4 and 8 weeks postinjury. P0 level in the ES-treated groups, both in vitro and in vivo, was enhanced compared with the controls. The earlier peak of Par-3 in the ES-treated groups indicated an earlier initiation of Schwann cell myelination. Additionally, ES significantly elevated BDNF expression in nerve tissues and in cocultures. ES on the site of nerve injury potentiates axonal regrowth and myelin maturation during peripheral nerve regeneration. Furthermore, the therapeutic actions of ES on myelination are mediated via enhanced BDNF signals, which drive the promyelination effect on Schwann cells at the onset of myelination.

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Interferon-gamma inhibits central nervous system myelination through both STAT1-dependent and STAT1-independent pathways.

Publication Date: 2010 Sep PMID: 20648647
Authors: Lin, W. - Lin, Y.
Journal: J Neurosci Res

The immune cytokine interferon-gamma (IFN-gamma) plays a crucial role in immune-mediated demyelinating diseases such as multiple sclerosis and experimental autoimmune encephalomyelitis (EAE). Our previous studies have shown that enforced expression of IFN-gamma in the central nervous system (CNS) inhibits developmental myelination or remyelination in EAE demyelinated lesions. Although many of the cellular actions of IFN-gamma result from its activation of the signal transducer and activator of transcription 1 (STAT1) pathway, recent studies have shown that STAT1-independent pathways regulate some facets of IFN-gamma biology. In this study, we dissected the role ofSTAT1-dependent and STAT1-independent pathways in IFN-gamma-induced hypomyelination using a genetic approach. We found that the induction of STAT1-dependent, IFN-gamma-responsive genes in response to this cytokine was abolished in the CNS of STAT1 null mice. Moreover, STAT1 deletion diminished oligodendrocyte loss, reduction of myelinated axons, and the inflammatory response in the CNS of transgenic mice that ectopically expressed IFN-gamma in the CNS. Nevertheless, IFN-gamma-induced reduction of myelin sheath thickness in the CNS of these mice was not altered by STAT1 deletion. Collectively, these data demonstrate that both STAT1-dependent and STAT1-independent pathways are involved in the detrimental effects of IFN-gamma on the myelination process.

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Reduction of Dicer impairs Schwann cell differentiation and myelination.

Publication Date: 2010 Sep PMID: 20648646
Authors: Verrier, J. D. - Semple-Rowland, S. - Madorsky, I. - Papin, J. E. - Notterpek, L.
Journal: J Neurosci Res

The process of Schwann cell myelination requires precisely coordinated gene expression. At the onset of myelination, there is an increase in the expression of differentiation-promoting transcription factors that regulate key Schwann cell genes. Further control of myelin gene expression occurs at the posttranscriptional level and, in part, is mediated by RNA binding proteins and micro-RNAs (miRNAs). miRNAs are small, endogenously derived RNA molecules that repress gene expression by specifically binding to their mRNA targets. In the experiments described here, we tested whether miRNAs were essential in controlling myelination by reducing the levels of Dicer, an essential endoribonuclease in miRNA biogenesis. We decreased the expression of Dicer by about 60% within Schwann cells using a lentiviral vector expressing an shRNA against Dicer. The reduced levels of Dicer led to a decrease in the steady-state expression of selected miRNAs and of the transcription factors Oct6 and Egr2/Krox20, both of which are critical for Schwann cells differentiation and myelination. In contrast, the levels of c-jun and Sox2 were up-regulated by the reduction in Dicer and were associated with an increase in Schwann cell proliferation. In dorsal root ganglion cocultures, Schwann cells transduced with Dicer shRNA synthesized less myelin, which was accompanied by significant reductions in the levels of myelin basic protein and protein zero. These findings support a critical role for Dicer and miRNAs in Schwann cell differentiation and myelination.

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Peptidylgycine alpha-amidating monooxygenase and copper: a gene-nutrient interaction critical to nervous system function.

Publication Date: 2010 Sep PMID: 20648645
Authors: Bousquet-Moore, D. - Mains, R. E. - Eipper, B. A.
Journal: J Neurosci Res

Peptidylgycine alpha-amidating monooxygenase (PAM), a highly conserved copper-dependent enzyme, is essential for the synthesis of all amidated neuropeptides. Biophysical studies revealed that the binding of copper to PAM affects its structure, and cell biological studies demonstrated that the endocytic trafficking of PAM was sensitive to copper. We review data indicating that genetic reduction of PAM expression and mild copper deficiency in mice cause similar alterations in several physiological functions known to be regulated by neuropeptides: thermal regulation, seizure sensitivity, and anxiety-like behavior.

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Expression of the IP3R1 promoter-driven nls-lacZ transgene in Purkinje cell parasagittal arrays of developing mouse cerebellum.

Publication Date: 2010 Oct PMID: 20632399
Authors: Furutama, D. - Morita, N. - Takano, R. - Sekine, Y. - Sadakata, T. - Shinoda, Y. - Hayashi, K. - Mishima, Y. - Mikoshiba, K. - Hawkes, R. - Furuichi, T.
Journal: J Neurosci Res

The cerebellar Purkinje cell monolayer is organized into heterogeneous Purkinje cell compartments that have different molecular compositions. Here we describe a transgenic mouse line, 1NM13, that shows heterogeneous transgene expression in parasagittal Purkinje cell arrays. The transgene consists of a nuclear localization signal (nls) fused to the beta-galactosidase (lacZ) composite gene driven by the type 1 inositol 1,4,5-trisphosphate receptor (IP(3)R1) gene promoter. IP(3)R1-nls-lacZ transgene expression was detected at a single Purkinje cell level over the surface of a whole-mount X-gal-stained cerebellum because of nuclear accumulation of the nls-lacZ activity. Developing cerebella of 1NM13 mice showed stripe-like X-gal staining patterns of parasagittal Purkinje cell subsets. The X-gal stripe pattern was likely determined by an intrinsic property as early as E15 and showed increasing complexity with cerebellar development. The X-gal stripe pattern was reminiscent of, but not identical to, the stripe pattern of zebrin II immunoreactivity. We designated the symmetrical X-gal-positive (transgene-positive, Tg(+)) Purkinje cell stripes about the midline as vermal Tg1(+), Tg2(a, b)(+) and Tg3(a, b)(+) stripes and hemispheric Tg4(a, b)(+), Tg5(a, b)(+), Tg6(a, b, c)(+), and Tg7(a, b)(+) stripes, where a, b, and c indicate substripes. We also assigned three parafloccular substripes Tg8(a, b, c)(+). The boundaries of X-gal stripes at P5 were consistent with raphes in the Purkinje cell layer through which granule cells migrate, suggesting a possible association of the X-gal stripes with raphe formation. Our results indicate that 1NM13 is a good mouse model with a reproducible and clear marker for the compartmentalization of Purkinje cell arrays.

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Inhibition of neutral sphingomyelinase-2 perturbs brain sphingolipid balance and spatial memory in mice.

Publication Date: 2010 Oct PMID: 20629193
Authors: Tabatadze, N. - Savonenko, A. - Song, H. - Bandaru, V. V. - Chu, M. - Haughey, N. J.
Journal: J Neurosci Res

The sphingolipid ceramide is a bioactive signaling lipid that is thought to play important roles in modulating synaptic activity, in part by regulating the function of excitatory postsynaptic receptors. However, the molecular mechanisms by which ceramide exerts its effects on synaptic activity remain largely unknown. We recently demonstrated that a rapid generation of ceramide by neutral sphingomyelinase-2 (nSMase2; also known as "sphingomyelin phosphodiesterase-3") played a key role in modulating excitatory postsynaptic currents by controlling the insertion and clustering of NMDA receptors (Wheeler et al. [2009] J. Neurochem. 109:1237-1249). We now demonstrate that nSMase2 plays a role in memory. Inhibition of nSMase2 impaired spatial and episodic-like memory in mice. At the molecular level, inhibition of nSMase2 decreased ceramide, increased PSD-95, increased the number of AMPA receptors, and altered the subunit composition of NMDA receptors. Our study identifies nSMase2 as an important component for efficient memory formation and underscores the importance of ceramide in regulating synaptic events related to learning and memory.

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Bone marrow mononuclear cells protect neurons and modulate microglia in cell culture models of ischemic stroke.

Publication Date: 2010 Oct PMID: 20629187
Authors: Sharma, S. - Yang, B. - Strong, R. - Xi, X. - Brenneman, M. - Grotta, J. C. - Aronowski, J. - Savitz, S. I.
Journal: J Neurosci Res

Although several studies have provided evidence for the therapeutic potential of bone marrow-derived mononuclear cells (MNCs) in animal models of stroke, the mechanisms underlying their benefits remain largely unknown. We have determined the neuroprotective potential of MNCs in primary neuronal cultures exposed to various injuries in vitro. Cortical neurons in culture were exposed to oxygen-glucose deprivation, hypoxia, or hydrogen peroxide, and cell death was assayed by MTT, caspase-3 activation or TUNEL labelling at 24 hrs. Cultures were randomized to cotreatment with MNC-derived supernatants or media before injury exposure. In separate experiments, macrophage or microglial cultures were exposed to lipopolypolysacharide (LPS) in the presence and absence of MNC-derived supernatants. Neuronal cultures were then exposed to conditioned media derived from activated macrophages or microglia. Cytokines from the supernantants of MNC cultures exposed to normoxia or hypoxia were also estimated by enzyme-linked immunosorbant assay (ELISA). MNC-derived supernatants attenuated neuronal death induced by OGD, hypoxia, hydrogen peroxide, and conditioned macrophage/microglial media and contain a number of trophic factors, including interleukin-10, insulin-like growth factor-1, vascular endothelial growth factor, and stromal cell-derived factor-1. MNCs provide broad neuroprotection against a variety of injuries relevant to stroke.

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Implication of Akt-dependent Prp19 alpha/14-3-3beta/Cdc5L complex formation in neuronal differentiation.

Publication Date: 2010 Oct PMID: 20629186
Authors: Urano-Tashiro, Y. - Sasaki, H. - Sugawara-Kawasaki, M. - Yamada, T. - Sugiyama, A. - Akiyama, H. - Kawasaki, Y. - Tashiro, F.
Journal: J Neurosci Res

PRP19alpha and CDC5L are major components of the active spliceosome. However, their association process is still unknown. Here, we demonstrated that PRP19 alpha/14-3-3beta/CDC5L complex formation is regulated by Akt during nerve growth factor (NGF)-induced neuronal differentiation of PC12 cells. Analysis of PRP19 alpha mutants revealed that the phosphorylation of PRP19 alpha at Thr 193 by Akt was critical for its binding with 14-3-3beta to translocate into the nuclei and for PRP19 alpha/14-3-3beta/CDC5L complex formation in neuronal differentiation. Forced expression of either sense PRP19 alpha or sense 14-3-3beta RNAs promoted NGF-induced neuronal differentiation, whereas down-regulation of these mRNAs showed a suppressive effect. The nonphosphorylation mutant PRP19 alpha T193A lost its binding ability with 14-3-3beta and acted as a dominant-negative mutant in neuronal differentiation. These results imply that Akt-dependent phosphorylation of PRP19 alpha at Thr193 triggers PRP19 alpha/14-3-3beta/CDC5L complex formation in the nuclei, likely to assemble the active spliceosome against neurogenic pre-mRNAs.

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Aminoglycoside-mediated partial suppression of MECP2 nonsense mutations responsible for Rett syndrome in vitro.

Publication Date: 2010 Aug 15 PMID: 20623622
Authors: Popescu, A. C. - Sidorova, E. - Zhang, G. - Eubanks, J. H.
Journal: J Neurosci Res

Rett syndrome is a pediatric neurological condition that affects primarily girls. Approximately 30% of Rett syndrome cases arise from point mutations that introduce a premature stop codon into the MECP2 gene. Several studies have now shown that certain aminoglycosides can facilitate read-through of some types of nonsense mutations in a context-dependent manner and allow the generation of a full-length protein. It remains mostly unclear whether different nonsense mutations of MECP2 will be responsive to aminoglycoside treatment. In this study, we tested whether the common premature terminating mutations of MECP2 seen in Rett syndrome cases can be partially suppressed by aminoglycoside administration. Our results show that aminoglycosides allow different mutant forms of MECP2 to be overcome in transiently transfected HEK293 cells, but with differing levels of efficiency. In addition, we also show that aminoglycosides increased the prevalence of full-length MeCP2 protein in a dose-dependent manner in a lymphocyte cell line derived from a Rett syndrome girl with the R255X mutation. This study helps to establish the "proof of principle" that some nonsense mutations causing Rett syndrome can be at least partially suppressed by drug treatment.

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Calpeptin attenuated inflammation, cell death, and axonal damage in animal model of multiple sclerosis.

Publication Date: 2010 Aug 15 PMID: 20623621
Authors: Guyton, M. K. - Das, A. - Samantaray, S. - Wallace, G. C. 4th - Butler, J. T. - Ray, S. K. - Banik, N. L.
Journal: J Neurosci Res

Experimental autoimmune encephalomyelitis (EAE) is an animal model for studying multiple sclerosis (MS). Calpain has been implicated in many inflammatory and neurodegenerative events that lead to disability in EAE and MS. Thus, treating EAE animals with calpain inhibitors may block these events and ameliorate disability. To test this hypothesis, acute EAE Lewis rats were treated dose dependently with the calpain inhibitor calpeptin (50-250 microg/kg). Calpain activity, gliosis, loss of myelin, and axonal damage were attenuated by calpeptin therapy, leading to improved clinical scores. Neuronal and oligodendrocyte death were also decreased, with down-regulation of proapoptotic proteins, suggesting that decreases in cell death were due to decreases in the expression or activity of proapoptotic proteins. These results indicate that calpain inhibition may offer a novel therapeutic avenue for treating EAE and MS.

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X-linked mental retardation gene CASK interacts with Bcl11A/CTIP1 and regulates axon branching and outgrowth.

Publication Date: 2010 Aug 15 PMID: 20623620
Authors: Kuo, T. Y. - Hong, C. J. - Chien, H. L. - Hsueh, Y. P.
Journal: J Neurosci Res

Calcium/calmodulin-dependent serine kinase (CASK), a causative gene in X-linked mental retardation, carries out multiple functions in neurons, including vesicle trafficking of ion channels, synapse formation, and gene transcription. From a yeast two-hybrid screen, Kruppel-like zinc finger protein B cell lymphoma/COUP-TF-interacting protein 1 (Bcl11A/CTIP1) was identified as a CASK binding protein. Through alternative splicing, a single Bcl11A gene encodes two major protein products in neurons, Bcl11A-S and Bcl11A-L. CASK interacted with both Bcl11A-S and Bcl11A-L in transfected COS cells and brain. Immunofluorescence staining further indicated the colocalization of CASK and Bcl11A in the nuclei of neurons. These studies supported an interaction between CASK and Bcl11A in vivo. Bcl11A-L has previously been shown to play a role in gene transcription as well as axon outgrowth and branching. Here, we further show that Bcl11A-L rearranges the distribution of nuclear actin, which may be related to the function of Bcl11A-L in gene expression. More importantly, using cultured hippocampal neurons as a model system, we show that CASK enhances the ability of Bcl11A-L to restrict axon outgrowth and branching. Interruption of the interaction between CASK and Bcl11A increased the outgrowth and branching of axons, suggesting that the interaction between CASK and Bcl11A controls axon arborization. In conclusion, our results suggest that, through the interaction with Bcl11A, CASK plays a role in axonogenesis, which may be related to brain anatomical characteristics in humans.

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Novel D3 dopamine receptor-preferring agonist D-264: Evidence of neuroprotective property in Parkinson's disease animal models induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine and lactacystin.

Publication Date: 2010 Aug 15 PMID: 20623619
Authors: Li, C. - Biswas, S. - Li, X. - Dutta, A. K. - Le, W.
Journal: J Neurosci Res

Parkinson's disease (PD), a progressive neurodegenerative movement disorder, is known to be caused by diverse pathological conditions resulting from dysfunction of the ubiquitin-proteasome system (UPS), mitochondria, and oxidative stress leading to preferential nigral dopamine (DA) neuron degeneration in the substantia nigra. In the present study, we evaluated the novel D3 receptor-preferring agonist D-264 in a mouse model of PD to evaluate its neuroprotective properties against both the nigrostriatal dopaminergic toxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)- and the proteasome inhibitor lactacystin-induced dopaminergic degeneration. C57BL/6 male mice either were given MPTP by intraperitoneal injection twice per day for 2 successive days at a dose 20 mg/kg or were microinjected with lactacystin bilaterally (1.25 microg/side) into the medial forebrain bundle (MFB). Pretreatment with D-264 (1 mg/kg and 5 mg/kg, intraperitoneally, once per day), started 7 days before administration of MPTP or lactacystin. We found that D-264 significantly improved behavioral performance, attenuated both MPTP- and lactacystin-induced DA neuron loss, and blocked proteasomal inhibition and microglial activation in the substantia nigra (SN). Furthermore, D-264 treatment was shown to increase the levels of brain-derived neurotrophic factor (BDNF) and glial cell line-derived factor (GDNF) in MPTP- and lactacystin-treated mice, possibly indicating, at least in part, the mechanism of neuroprotection by D-264. Furthermore, pretreatment with the D3 receptor antagonist U99194 significantly altered the effect of neuroprotection conferred by D-264. Collectively, our study demonstrates that D-264 can prevent neurodegeneration induced by the selective neurotoxin MPTP and the UPS inhibitor lactacystin. The results indicate that D-264 could potentially serve as a symptomatic and neuroprotective treatment agent for PD.

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Preferential inhibition by antidiarrheic 2-methoxy-4-methylphenol of Ca(2+) influx across acquired N-methyl-D-aspartate receptor channels composed of NR1/NR2B subunit assembly.

Publication Date: 2010 Aug 15 PMID: 20623618
Authors: Nakamichi, N. - Fukumori, R. - Takarada, T. - Kambe, Y. - Yamamoto, T. - Matsushima, N. - Moriguchi, N. - Yoneda, Y.
Journal: J Neurosci Res

In our previous studies, particular phenolic ingredients, such as 2-methoxy-4-methylphenol (2M4MP), of the antidiarrheic drug wood creosote significantly prevented cell death by both hydrogen peroxide and glutamate in cultured rat hippocampal neurons. In this study, we further evaluated the pharmacological properties of 2M4MP on Ca(2+) influx across native and acquired N-methyl-D-aspartate (NMDA) receptor (NMDAR) channels. The addition of 2M4MP significantly prevented the loss of cellular viability and the increase in intracellular free Ca(2+) levels as determined by Fluo-3 in cultured rat hippocampal neurons briefly exposed to NMDA. Brief exposure to NMDA also led to a marked increase in mitochondrial free Ca(2+) levels determined by Rhod-2, in addition to intracellular free Ca(2+) levels, in HEK293 cells expressing either NR1/NR2A or NR1/NR2B subunit channels. The further addition of the general NMDAR channel blocker dizocilpine similarly inhibited the increase of intracellular Ca(2+) levels by NMDA in both types of acquired NMDAR channels, whereas the NR2B subunit selective antagonist ifenprodil drastically inhibited the increase by NMDA in HEK293 cells expressing NR1/NR2B, but not NR1/NR2A, subunits. Similarly, 2M4MP significantly and selectively inhibited the NMDA-induced influx of Ca(2+) across acquired NR1/NR2B channels in a concentration-dependent manner. Moreover, prior daily oral administration of 2M4MP significantly reduced the infarct volume in the ipsilateral cerebral hemisphere in rats with middle cerebral artery occlusion 1 day after reperfusion. These results suggest that 2M4MP may protect neurons from excitotoxicity through preferential inhibition of Ca(2+) influx across NMDAR channels composed of NR1/NR2B subunits.

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Role of proteoglycans and glycosaminoglycans in the pathogenesis of Alzheimer's disease and related disorders: amyloidogenesis and therapeutic strategies--a review.

Publication Date: 2010 Aug 15 PMID: 20623617
Authors: Ariga, T. - Miyatake, T. - Yu, R. K.
Journal: J Neurosci Res

The extracellular accumulation of amyloid beta proteins (Abetas) in neuritic plaques is one of the hallmarks of Alzheimer's disease (AD). The binding of Abetas to extracellular membranes (ECMs) is a critical step in developing AD. Abetas bind to many biomolecules, including lipids, proteins, and proteoglycans (PGs). PGs play several roles in amyloid formation, including promoting the aggregation of Abetas into insoluble amyloid fibrils, which contributes to the increased neurotoxicity of Abetas. Although Abetas readily self-aggregate to form amyloid fibrils in vitro, their binding to PGs and heparin enhances amyloid aggregation and fibril formation. The sulfate moiety in glycosaminoglycans (GAGs), the carbohydrate portion of PGs, is necessary for the formation of amyloid fibrils; no fibrils are observed in the presence of hyaluronic acid (HA), a nonsulfated GAG. PGs and Abetas are known to colocalize in senile plaques (SPs) and neurofibrillary tangles (NFTs) in the AD brain. The binding site of PGs to Abetas has been identified in the 13-16-amino-acid region (His-His-Gln-Lys) of Abetas and represents a unique target site for inhibition of amyloid fibril formation; His13 in particular is an important residue critical for interaction with GAGs. The sulfate moieties of GAGs play a critical role in the binding to Abetas and enhance Abeta fibril formation. Low-molecular-weight heparins (LMWHs) can reverse the process of amyloidosis to inhibit fibril formation by blocking the formation of beta-plated structures, suggesting a possible therapeutic approach using LMWHs to interfere with the interaction between PGs and Abetas and to arrest or prevent amyloidogenesis.

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Potential roles of D-serine and serine racemase in experimental temporal lobe epilepsy.

Publication Date: 2010 Aug 15 PMID: 20623543
Authors: Ryu, H. J. - Kim, J. E. - Yeo, S. I. - Kim, D. S. - Kwon, O. S. - Choi, S. Y. - Kang, T. C.
Journal: J Neurosci Res

To confirm the roles of D-serinergic gliotransmission in epilepsy, we investigated the relationship between spatiotemporally specific glial responses and the D-serine/serine racemase system in mesial temporal structures following status epilepticus (SE). In control animals, D-serine and serine racemase immunoreactivities were detected mainly in astrocytes. After SE, D-serine and serine racemase immunoreactivities were increased in astrocytes. Double-immunofluorescence study revealed that up-regulation of serine racemase immunoreactivity was relevant not to D-serine immunoreactivity but to nestin or vimentin immunoreactivity. Neither D-serine nor serine racemase was found in naive or reactive microglia. In addition, phosphorylated N-methyl-D-aspartate (NMDA) receptor subunit 1 (pNR1-Ser896) immunoreactivity in the hippocampus was increased compared with controls. Increased D-serine immunoreactivity showed direct correlation with the phosphorylation of Ser896 of NR1. Given the findings of our previous study, these findings suggest that D-serine and serine racemase in astrocytes may play roles in neuronal hyperexcitability via a cooperative activation of NMDA receptors. Furthermore, serine racemase may be involved in migration and differentiation of immature astrocytes, which is relevant to reactive astrogliosis.

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Delta-catenin/NPRAP: A new member of the glycogen synthase kinase-3beta signaling complex that promotes beta-catenin turnover in neurons.

Publication Date: 2010 Aug 15 PMID: 20623542
Authors: Bareiss, S. - Kim, K. - Lu, Q.
Journal: J Neurosci Res

Through a multiprotein complex, glycogen synthase kinase-3beta (GSK-3beta) phosphorylates and destabilizes beta-catenin, an important signaling event for neuronal growth and proper synaptic function. delta-Catenin, or NPRAP (CTNND2), is a neural enriched member of the beta-catenin superfamily and is also known to modulate neurite outgrowth and synaptic activity. In this study, we investigated the possibility that delta-catenin expression is also affected by GSK-3beta signaling and participates in the molecular complex regulating beta-catenin turnover in neurons. Immunofluorescent light microscopy revealed colocalization of delta-catenin with members of the molecular destruction complex: GSK-3beta, beta-catenin, and adenomatous polyposis coli proteins in rat primary neurons. GSK-3beta formed a complex with delta-catenin, and its inhibition resulted in increased delta-catenin and beta-catenin expression levels. LY294002 and amyloid peptide, known activators of GSK-3beta signaling, reduced delta-catenin expression levels. Furthermore, delta-catenin immunoreactivity increased and protein turnover decreased when neurons were treated with proteasome inhibitors, suggesting that the stability of delta-catenin, like that of beta-catenin, is regulated by proteasome-mediated degradation. Coimmunoprecipitation experiments showed that delta-catenin overexpression promoted GSK-3beta and beta-catenin interactions. Primary cortical neurons and PC12 cells expressing delta-catenin treated with proteasome inhibitors showed increased ubiquitinated beta-catenin forms. Consistent with the hypothesis that delta-catenin promotes the interaction of the destruction complex molecules, cycloheximide treatment of cells overexpressing delta-catenin showed enhanced beta-catenin turnover. These studies identify delta-catenin as a new member of the GSK-3beta signaling pathway and further suggest that delta-catenin is potentially involved in facilitating the interaction, ubiquitination, and subsequent turnover of beta-catenin in neuronal cells.

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Arachidonic acid inhibits neuronal nitric oxide synthase elicited by proinflammatory stimuli and promotes astrocyte survival with both exogenous and endogenous peroxynitrite via different mechanisms.

Publication Date: 2010 Aug 15 PMID: 20623541
Authors: Palomba, L. - Cerioni, L. - Cantoni, O.
Journal: J Neurosci Res

Cytosolic phospholipase A(2)-inhibited astrocytes respond to the cocktail lipopolysaccharide/interferon-gamma with an immediate formation of peroxynitrite (ONOO(-)) and a delayed lethal response. Low concentrations of arachidonic acid (ARA; i.e., <0.1 microM) cause tyrosine kinase-dependent inhibition of neuronal nitric oxide synthase (nNOS) activity, thereby suppressing formation of ONOO(-) and the ensuing lethal response. ARA promoted its effects only when given to the cultures just prior to, or in parallel with, the proinflammatory mixture. High concentrations of ARA, i.e., >3 microM, promoted cytoprotection when applied to the cultures up to 50 min after the formation of endogenous ONOO(-) had been completed or up to 30 min after addition of exogenous ONOO(-). The mechanism(s) involved in these responses was, however, independent of tyrosine kinase activation and was in fact mediated by ARA metabolites of the lipoxygenase pathway. These results are consistent with a scenario in which astrocytes respond to low or high amounts of ARA with the triggering of different pathways involved in the inflammatory response. Early nNOS inhibition mediated by very low levels of ARA is indeed critical for nuclear factor-kappaB activation, which is otherwise effectively inhibited by constitutive nitric oxide, and for preventing early formation of ONOO(-). Greater ARA concentrations promote survival in astrocytes committed to death by ONOO(-), a species extensively released under inflammatory conditions, via a mechanism dependent on lipoxygenase metabolism and inhibition of downstream events leading to cell demise.

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Expression of plasminogen activator inhibitor-1 and protease nexin-1 in human astrocytes: Response to injury-related factors.

Publication Date: 2010 Aug 15 PMID: 20623540
Authors: Hultman, K. - Blomstrand, F. - Nilsson, M. - Wilhelmsson, U. - Malmgren, K. - Pekny, M. - Kousted, T. - Jern, C. - Tjarnlund-Wolf, A.
Journal: J Neurosci Res

Astrocytes play a diverse role in central nervous system (CNS) injury. Production of the serine protease inhibitors (serpins) plasminogen activator inhibitor-1 (PAI-1) and protease nexin-1 (PN-1) by astrocytes may counterbalance excessive serine protease activity associated with CNS pathologies such as ischemic stroke. Knowledge regarding the regulation of these genes in the brain is limited, so the objective of the present study was to characterize the effects of injury-related factors on serpin expression in human astrocytes. Native human astrocytes were exposed to hypoxia or cytokines, including interleukin-6 (IL-6), IL-1beta, tumor necrosis factor-alpha (TNF-alpha), IL-10, transforming growth factor-alpha (TGF-alpha), and TGF-beta for 0-20 hr. Serpin mRNA expression and protein secretion were determined by real-time RT-PCR and ELISA, respectively. Localization of PAI-1 and PN-1 in human brain tissue was examined by immunohistochemistry. Hypoxia and all assayed cytokines induced a significant increase in PAI-1 expression, whereas prolonged treatment with IL-1beta or TNF-alpha resulted in a significant down-regulation. The most pronounced induction of both PAI-1 and PN-1 was observed following early treatment with TGF-alpha. In contrast to PAI-1, the PN-1 gene did not respond to hypoxia. Positive immunoreactivity for PAI-1 in human brain tissue was demonstrated in reactive astrocytes within gliotic areas of temporal cortex. We show here that human astrocytes express PAI-1 and PN-1 and demonstrate that this astrocytic expression is regulated in a dynamic manner by injury-related factors.

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Endogenous expression of interleukin-4 regulates macrophage activation and confines cavity formation after traumatic spinal cord injury.

Publication Date: 2010 Aug 15 PMID: 20623539
Authors: Lee, S. I. - Jeong, S. R. - Kang, Y. M. - Han, D. H. - Jin, B. K. - Namgung, U. - Kim, B. G.
Journal: J Neurosci Res

Traumatic spinal cord injury (SCI) triggers inflammatory reactions in which various types of cells and cytokines are involved. Several proinflammatory cytokines are up-regulated after SCI and play crucial roles in determining the extent of secondary tissue damage. However, relatively little is known about antiinflammatory cytokines and their roles in spinal cord trauma. Recent studies have shown that an antiinflammatory cytokine, interleukin-4 (IL-4), is expressed and exerts various modulatory effects in CNS inflammation. We found in the present study that IL-4 was highly expressed at 24 hr after contusive SCI in rats and declined thereafter, with concurrent up-regulation of IL-4 receptor subunit IL-4alpha. The majority of IL-4-producing cells were myeloperoxidase-positive neutrophils. Injection of neutralizing antibody against IL-4 into the contused spinal cord did not significantly affect the expression levels of proinflammatory cytokines such as IL-1beta, IL-6, and tumor necrosis factor-alpha or other antiinflammatory cytokines such as IL-10 and transforming growth factor-beta. Instead, attenuation of IL-4 activity led to a marked increase in the extent of ED1-positive macrophage activation along the rostrocaudal extent at 7 days after injury. The enhanced macrophage activation was preceded by an increase in the level of monocyte chemoattractant protein-1 (MCP-1/CCL2). Finally, IL-4 neutralization resulted in more extensive cavitation at 4 weeks after injury. These results suggest that endogenous expression of antiinflammatory cytokine IL-4 regulates the extent of acute macrophage activation and confines the ensuing secondary cavity formation after spinal cord trauma.

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Immunodeficiency reduces neural stem/progenitor cell apoptosis and enhances neurogenesis in the cerebral cortex after stroke.

Publication Date: 2010 Aug 15 PMID: 20623538
Authors: Saino, O. - Taguchi, A. - Nakagomi, T. - Nakano-Doi, A. - Kashiwamura, S. - Doe, N. - Nakagomi, N. - Soma, T. - Yoshikawa, H. - Stern, D. M. - Okamura, H. - Matsuyama, T.
Journal: J Neurosci Res

Acute inflammation in the poststroke period exacerbates neuronal damage and stimulates reparative mechanisms, including neurogenesis. However, only a small fraction of neural stem/progenitor cells survives. In this report, by using a highly reproducible model of cortical infarction in SCID mice, we examined the effects of immunodeficiency on reduction of brain injury, survival of neural stem/progenitor cells, and functional recovery. Subsequently, the contribution of T lymphocytes to neurogenesis was evaluated in mice depleted for each subset of T lymphocyte. SCID mice revealed the reduced apoptosis and enhanced proliferation of neural stem/progenitor cells induced by cerebral cortex after stroke compared with the immunocompetent wild-type mice. Removal of T lymphocytes, especially the CD4(+) T-cell population, enhanced generation of neural stem/progenitor cells, followed by accelerated functional recovery. In contrast, removal of CD25(+) T cells, a cell population including regulatory T lymphocytes, impaired functional recovery through, at least in part, suppression of neurogenesis. Our findings demonstrate a key role of T lymphocytes in regulation of poststroke neurogenesis and indicate a potential novel strategy for cell therapy in repair of the central nervous system.

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Chronic social isolation suppresses proplastic response and promotes proapoptotic signalling in prefrontal cortex of Wistar rats.

Publication Date: 2010 Aug 15 PMID: 20623537
Authors: Djordjevic, A. - Adzic, M. - Djordjevic, J. - Radojcic, M. B.
Journal: J Neurosci Res

Successful adaptation to stress involves synergized actions of glucocorticoids and catecholamines at several levels of the CNS, including the prefrontal cortex (PFC). Inside the PFC, hormonal signals trigger concerted actions of transcriptional factors, such as glucocorticoid receptor (GR) and nuclear factor kappa B (NFkappaB), culminating in a balanced, proadaptive expression of their common genes, such as proplastic NCAM and/or apoptotic Bax and Bcl-2. In the present study, we hypothesized that chronic stress may compromise the balance between GR and NFkappaB signals and lead to an altered/maladaptive expression of their cognate genes in the PFC. Our results obtained with Wistar rats exposed to chronic social isolation indicated alterations of the GR relative to the NFkappaB, in favor of the GR, in both the cytoplasmic and the nuclear compartments of the PFC. Although these alterations did not affect the induction of proplastic NCAM gene, they decreased the NCAM sialylation necessary for plastic response and caused marked relocation of the mitochondrial membrane antiapoptotic Bcl-2 protein to its cytoplasmic form. Moreover, the compromised PSA-NCAM plastic response found under chronic stress was sustained after exposure of animals to the subsequent acute stress, whereas the proapoptotic signals were further emphasized. It is concluded that chronic social isolation of Wistar animals leads to a maladaptive response of the PFC, considering the diminishment of its plastic potential and potentiating of apoptosis. Such conditions in the PFC are likely to compromise its ability to interact with other CNS structures, such as the hippocampus, which is necessary for successful adaptation to stress.

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Long-lasting post-mortem activity of spinal microglia in situ in mice.

Publication Date: 2010 Aug 15 PMID: 20623536
Authors: Dibaj, P. - Steffens, H. - Nadrigny, F. - Neusch, C. - Kirchhoff, F. - Schomburg, E. D.
Journal: J Neurosci Res

As CNS macrophages, microglia show a high spontaneous motility of their processes, continuously surveying their microenvironment. Upon CNS injury, microglia react by immediate cellular polarization and process extension toward the lesion site as well as by subsequent amoeboid lesion-directed migration and phagocytosis. To determine the ability of microglia to fulfill their role within distinctively lesioned tissue in the absence of life support, we investigated microglial activity and responsiveness to laser-induced axonal injuries in the spinal dorsal columns in situ after cardiac and respiratory arrest, i.e., post-mortem, in the progressively degrading nervous tissue. For this purpose, we used time-lapse two-photon laser scanning microscopy in double transgenic mice expressing enhanced green fluorescent protein in microglia and enhanced yellow fluorescent protein in projection neurons. Depending on the premortal condition of the animal, microglial activity and responsiveness remain for up to5-10 hr post-mortem. Thereby, the continuously decreasing glial reaction is independent of oxygen and glucose supply but requires residual ATP, suggesting a parasitic form of energy, such as a transmembrane uptake of ATP released from injured nervous tissue. Even though initially microglia are able to detect axonal injury after disruption of the blood supply, the later aspects of glial reaction, for example amoeboid conversion and migration, are absent post- mortem, corresponding to the failure of microglia to prevent secondary damage after injury of nervous tissue.

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Interaction of dopamine D1 receptor with N-ethylmaleimide-sensitive factor is important for the membrane localization of the receptor.

Publication Date: 2010 Aug 15 PMID: 20623535
Authors: Chen, S. - Liu, F.
Journal: J Neurosci Res

The dopamine D1 receptor (D1R) plays important roles in regulating motor coordination, working memory, learning, and reward. In the mammalian brain, D1R is localized predominantly in dendritic spines. However, the molecular mechanisms involved in the transport, sorting, and targeting of D1R to dendritic spines are largely unknown. Here, we characterize the interaction between D1R and N-ethylmaleimide-sensitive factor (NSF) and show that the interaction is mediated by aa 387-401 of the D1R C-terminal tail. Interfering D1R and NSF interaction by coexpressing GFP-D1R aa 387-401 fusion protein reduces D1R membrane localization and inhibits D1R mediated cAMP accumulation. Treatment of hippocampal neurons with Tat-D1R aa 387-401 decreases the synaptic localization of D1R and the cell surface expression of D1R, but not the cell surface expression of alpha7 nicotinic receptor. Our data indicate that the interaction between NSF and D1R is important for the membrane localization of D1R.

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Role of mitogen-activated protein kinases in the mechanism of oxidant-induced cell swelling in cultured astrocytes.

Publication Date: 2010 Aug 15 PMID: 20623534
Authors: Moriyama, M. - Jayakumar, A. R. - Tong, X. Y. - Norenberg, M. D.
Journal: J Neurosci Res

Cytotoxic brain edema, usually a consequence of astrocyte swelling, is an important complication of stroke, traumatic brain injury, hepatic encephalopathy, and other neurological disorders. Although mechanisms underlying astrocyte swelling are not fully understood, oxidative stress (OS) has generally been considered an important factor in its pathogenesis. To better understand the mechanism(s) by which OS causes cell swelling, we examined the potential involvement of mitogen-activated protein kinases (MAPKs) in this process. Cultures exposed to theoxidant H(2)O(2) (10, 25, 50 microM) for different time periods (1-24 hr) significantly increased cell swelling in a triphasic manner. Swelling was initially observed at 10 min (peaking at 30 min), which was followed by cell shrinkage at 1 hr. A subsequent increase in cell volume occurred at approximately 6 hr, and the rise lasted for at least 24 hr. Cultures exposed to H(2)O(2) caused the activation of MAPKs (ERK1/2, JNK and p38-MAPK), whereas inhibition of MAPKs diminished cell swelling induced by 10 and 25 microM H(2)O(2). These findings suggest that activation of MAPKs is an important factor in the mediation of astrocyte swelling following oxidative stress.

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Spatiotemporal distribution and function of N-cadherin in postnatal Schwann cells: A matter of adhesion?

Publication Date: 2010 Aug 15 PMID: 20623533
Authors: Corell, M. - Wicher, G. - Limbach, C. - Kilimann, M. W. - Colman, D. R. - Svenningsen, A. F.
Journal: J Neurosci Res

During embryonic development of the peripheral nervous system (PNS), the adhesion molecule neuronal cadherin (N-cadherin) is expressed by Schwann cell precursors and associated with axonal growth cones. N-cadherin expression levels decrease as precursors differentiate into Schwann cells. In this study, we investigated the distribution of N-cadherin in the developing postnatal and adult rat peripheral nervous system. N-cadherin was found primarily in ensheathing glia throughout development, concentrated at neuron-glial or glial-glial contacts of the sciatic nerve, dorsal root ganglia (DRG), and myenteric plexi. In the sciatic nerve, N-cadherin decreases with age and progress of myelination. In adult animals, N-cadherin was found exclusively in nonmyelinating Schwann cells. The distribution of N-cadherin in developing E17 DRG primary cultures is similar to what was observed in vivo. Functional studies of N-cadherin in these cultures, using the antagonist peptide INPISGQ, show a disruption of the attachment between Schwann cells, but no interference in the initial or long-term contact between Schwann cells and axons. We suggest that N-cadherin acts primarily in the adhesion between glial cells during postnatal development. It may form adherents/junctions between nonmyelinating glia, which contribute to the stable tubular structure encapsulating thin caliber axons and thus stabilize the nerve structure as a whole.

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Impairment of nuclear factor-kappaB activation increased glutamate excitotoxicity in a motoneuron-neuroblastoma hybrid cell line expressing mutant (G93A) Cu/Zn-superoxide dismutase.

Publication Date: 2010 Aug 15 PMID: 20623531
Authors: Pyo, J. S. - Ko, Y. S. - Kim, W. H. - Kim, M. - Lee, K. W. - Nam, S. Y. - Chung, H. Y. - Cho, S. J. - Baik, T. K. - Lee, B. L.
Journal: J Neurosci Res

Mutations in the superoxide dismutase 1 (SOD1) gene are linked to glutamate excitotoxicity in familial amyotrophic lateral sclerosis (fALS), but the underlying mechanism remains unclear. We investigated whether nuclear factor-kappaB (NF-kappaB) activation is involved in glutamate excitotoxicity by using motor neuron-neuroblastoma hybrid cells that expressed a mutant (G93A) SOD1 (mtSOD1) or wild-type SOD1 (wtSOD1). MtSOD1 cells were more vulnerable to glutamate excitotoxicity than wtSOD1 cells and showed higher NF-kappaB activity, higher nuclear cRel expression, and lower nuclear RelA expression under basal conditions. Glutamate treatment increased NF-kappaB activation along with nuclear expressions of RelA and cRel in wtSOD1 cells but induced only weak nuclear RelA expression in mtSOD1 cells. Suppression of NF-kappaB activation using transfection of the superrepressive mutant form of IkappaBalpha (mIkappaBalpha) inhibited nuclear RelA expression in both types of SOD1 cells, which increased glutamate excitotoxicity in wtSOD1 cells but not in mtSOD1 cells. Furthermore, immunohistochemistry confirmed stronger RelA immunoreactivity in the nuclei of motor neurons of spinal cord in wild-type SOD1 transgenic mice than in those in SOD1 G93A transgenic mice. In addition, we found that glutamate treatment decreased XIAP expression and increased caspase-3 activity in mtSOD1 cells and mIkappaBalpha-overexpressing wtSOD1 cells. Our results suggest that glutamate excitotoxicity in motor neurons of SOD1-linked fALS is attributable, at least in part, to the impairment of IkappaBalpha-dependent RelA activation and subsequent apoptosis mediated by XIAP inhibition and caspase-3 activation.

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The inhibitory neurotransmitter glycine modulates macrophage activity by activation of neutral amino acid transporters.

Publication Date: 2010 Aug 15 PMID: 20623529
Authors: Carmans, S. - Hendriks, J. J. - Thewissen, K. - Van den Eynden, J. - Stinissen, P. - Rigo, J. M. - Hellings, N.
Journal: J Neurosci Res

Glycine, an important inhibitory neurotransmitter in the mammalian central nervous system (CNS), has been shown to modulate peripheral immune cell responses. In that respect, glycine levels are increased in several neuroinflammatory disorders, such as amyotrophic lateral sclerosis (ALS) and multiple sclerosis (MS). In this study, we show that glycine modulates macrophage effector functions implicated in CNS inflammation and in other, related inflammatory conditions. We demonstrate that glycine does not affect the production of reactive oxygen species but stimulates myelin phagocytosis and the production of the proinflammatory mediators nitric oxide (NO) and tumor necrosis factor (TNF)-alpha by rat macrophages. These effects of glycine are not mediated by the glycine receptor (GlyR) or by glycine transporters (GlyTs), as neither the GlyR antagonist strychnine nor the antagonist of GlyT1 (ALX5407) reverses the observed effects. In contrast, 2-aminoisobutyric acid, a substrate of neutral amino acid transporters (NAATs), inhibits the glycine-mediated enhancement of myelin phagocytosis as well as of NO and TNF-alpha production. In conclusion, our findings demonstrate that glycine modulates macrophage function through activation of NAATs. Glycine may thereby influence immunological processes in inflammatory diseases involving macrophage activation and demyelination, including MS and related conditions associated with altered glycine levels.

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Enhanced adenoviral gene delivery to motor and dorsal root ganglion neurons following injection into demyelinated peripheral nerves.

Publication Date: 2010 Aug 15 PMID: 20623527
Authors: Zhang, Y. - Zheng, Y. - Zhang, Y. P. - Shields, L. B. - Hu, X. - Yu, P. - Burke, D. A. - Wang, H. - Jun, C. - Byers, J. - Whittemore, S. R. - Shields, C. B.
Journal: J Neurosci Res

Injection of viral vectors into peripheral nerves may transfer specific genes into their dorsal root ganglion (DRG) neurons and motoneurons. However, myelin sheaths of peripheral axons block the entry of viral particles into nerves. We studied whether mild, transient peripheral nerve demyelination prior to intraneural viral vector injection would enhance gene transfer to target DRG neurons and motoneurons. The right sciatic nerve of C57BL/6 mice was focally demyelinated with 1% lysolecithin, and the left sciatic nerve was similarly injected with saline (control). Five days after demyelination, 0.5 microl of Ad5-GFP was injected into both sciatic nerves at the site of previous injection. The effectiveness of gene transfer was evaluated by counting GFP(+) neurons in the DRGs and ventral horns. After peripheral nerve demyelination, there was a fivefold increase in the number of infected DRG neurons and almost a 15-fold increase in the number of infected motoneurons compared with the control, nondemyelinated side. Focal demyelination reduced the myelin sheath barrier, allowing greater virus-axon contact. Increased CXADR expression on the demyelinated axons facilitated axoplasmic viral entry. No animals sustained any prolonged neurological deficits. Increased gene delivery into DRG neurons and motoneurons may provide effective treatment for amyotrophic lateral sclerosis, pain, and spinal cord injury.

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Combination of olfactory ensheathing cells with local versus systemic cAMP treatment after a cervical rubrospinal tract injury.

Publication Date: 2010 Oct PMID: 20568293
Authors: Bretzner, F. - Plemel, J. R. - Liu, J. - Richter, M. - Roskams, A. J. - Tetzlaff, W.
Journal: J Neurosci Res

The failure of CNS axons to regenerate following traumatic injury is due in part to a growth-inhibitory environment in CNS as well as a weak intrinsic neuronal growth response. Olfactory ensheathing cell (OECs) transplants have been reported to create a favorable environment promoting axonal regeneration, remyelination, and functional recovery after spinal cord injury. However, in our previous experiments, OEC transplants failed to promote regeneration of rubrospinal axons through and beyond the site of a dorsolateral funiculus crush in rats. Rubrospinal neurons undergo massive cell atrophy and limited expression of regeneration-associated genes after axotomy. Using the same injury model, we tested the hypothesis that treatment of the red nucleus with cAMP, known to stimulate the intrinsic growth response in other neurons, will promote rubrospinal regeneration in combination with OEC transplants. In addition, we assessed a systemic increase of cAMP using the phosphodiesterase inhibitor rolipram. OECs prevented cavity formation, attenuated astrocytic hypertrophy and the retraction of the axotomized rubrospinal axons, and tended to reduce the overall lesion size. OEC transplantation lowered the thresholds for thermal sensitivity of both forepaws. None of our treatments, alone or in combination, promoted rubrospinal regeneration through the lesion site. However, the systemic elevation of cAMP with rolipram resulted in greater numbers of OECs and axonal density within the graft and improved motor performance in a cylinder test in conjunction with enhanced rubrospinal branching and attenuated astrocytic hypertrophy.

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Progesterone regulates the phosphorylation of protein phosphatases in the brain.

Publication Date: 2010 Oct PMID: 20568292
Authors: Amorim, M. A. - Guerra-Araiza, C. - Pernia, O. - da Cruz e Silva, E. F. - Garcia-Segura, L. M.
Journal: J Neurosci Res

Previous studies have shown that progesterone modulates the activity of different kinases and the phosphorylation of Tau in the brain. These actions of progesterone may be involved in the hormonal regulation of neuronal differentiation, neuronal function, and neuroprotection. However, the action of progesterone on protein phosphatases in the nervous system has not been explored previously. In this study we have assessed the effect of the administration of progesterone to adult ovariectomized rats on protein phosphatase 2A (PP2A) and phosphatase and tensin homolog deleted on chromosome 10 (PTEN) in the hypothalamus, the hippocampus, and the cerebellum. Total levels of PP2A, the state of methylation of PP2A, and total levels of PTEN were unaffected by the hormone in the three brain regions studied. In contrast, progesterone significantly increased the levels of PP2A phosphorylated in tyrosine 307 in the hippocampus and the cerebellum and significantly decreased the levels of PTEN phosphorylated in serine 380 in the hypothalamus and in the hippocampus compared with control values. Estradiol priming blocked the effect of progesterone on PP2A phosphorylation in the hippocampus and on PTEN phosphorylation in the hypothalamus and the hippocampus. In contrast, the action of progesterone on PP2A phosphorylation in the cerebellum was not modified by estradiol priming. These findings suggest that the regulation of the phosphorylation of PP2A and PTEN may be involved in the effects of progesterone on the phosphorylation of Tau and on the activity of phophoinositide-3 kinase and mitogen-activated protein kinase in the brain.

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Astrocyte-derived interleukin-6 promotes specific neuronal differentiation of neural progenitor cells from adult hippocampus.

Publication Date: 2010 Oct PMID: 20568291
Authors: Oh, J. - McCloskey, M. A. - Blong, C. C. - Bendickson, L. - Nilsen-Hamilton, M. - Sakaguchi, D. S.
Journal: J Neurosci Res

The purpose of this study was to investigate the ability of astrocyte-derived factors to influence neural progenitor cell differentiation. We previously demonstrated that rat adult hippocampal progenitor cells (AHPCs) immunoreactive for the neuronal marker class III beta-tubulin (TUJ1) were significantly increased in the presence of astrocyte-derived soluble factors under noncontact coculture conditions. Using whole-cell patch-clamp analysis, we observed that the cocultured AHPCs displayed two prominent voltage-gated conductances, tetraethyl ammonium (TEA)-sensitive outward currents and fast transient inward currents. The outward and inward current densities of the cocultured AHPCs were approximately 2.5-fold and 1.7-fold greater, respectively, than those of cells cultured alone. These results suggest that astrocyte-derived soluble factors induce neuronal commitment of AHPCs. To investigate further the activity of a candidate neurogenic factor on AHPC differentiation, we cultured AHPCs in the presence or absence of purified rat recombinant interleukin-6 (IL-6). We also confirmed that the astrocytes used in this study produced IL-6 by ELISA and RT-qPCR. When AHPCs were cultured with IL-6 for 6-7 days, the TUJ1-immunoreactive AHPCs and the average length of TUJ1-immunoreactive neurites were significantly increased compared with the cells cultured without IL-6. Moreover, IL-6 increased the inward current density to an extent comparable to that of coculture with astrocytes, with no significant differences in the outward current density, apparent resting potential, or cell capacitance. These results suggest that astrocyte-derived IL-6 may facilitate AHPC neuronal differentiation. Our findings have important implications for understanding injury-induced neurogenesis and developing cell-based therapeutic strategies using neural progenitors.

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Modulation of sensory neuron-specific receptors in the development of morphine tolerance and its neurochemical mechanisms.

Publication Date: 2010 Oct PMID: 20568289
Authors: Chen, P. - Wang, D. - Li, M. - Zhang, Y. - Quirion, R. - Hong, Y.
Journal: J Neurosci Res

Prevention of opiate tolerance is a critical issue in pain management. The present study was designed to characterize the pharmacological properties of sensory neuron-specific receptors (SNSR; also known as Mas-related gene receptors, or Mrg) for their modulation in the development of morphine tolerance and to investigate the underlying mechanism(s). Daily coadministration of the SNSR agonist BAM8-22 at a dose of 0.01 or 0.001, but not 1.0, nmol with morphine (intrathecally, or i.t., 20 microg/day) for 6 days significantly decreased the development of morphine tolerance. Coadministration of BAM8-22 (i.t., 1.0 nmol) on days 1, 3, and 5 completely blocked tolerance to morphine-induced analgesia. Intermittent coadministration of the structurally dissimilar SNSR agonist (Tyr(6))-2-MSH-6-12 (MSH; 5 nmol) also produced similar modulation. Chronic administration of morphine (20 microg, i.t.) increased expression of neuronal nitric oxide synthase (nNOS) and calcitonin gene-related peptide (CGRP) in superficial layers of the spinal cord and dorsal root ganglia. All these increases were abolished when BAM8-22 or MSH was intermittently coadministered. Furthermore, intermittent administration of BAM8-22 inhibited morphine-induced increase in protein kinase C gamma (PKC gamma) in both membrane and cytosol of spinal dorsal horn neurons. These results suggest that moderate activation of SNSR modulated morphine tolerance by inhibition of the PKC signaling pathway, leading to abolishment of enhancement of nNOS and CGRP. As SNSR are uniquely located ina subset of small-sized neurons in dorsal root and trigeminal ganglia, intermittent combination of SNSR agonist could be a promising adjunct for sustained use of opiates without central nervous system side effects.

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Stromal cell-derived factor-1 promotes migration of cells from the upper rhombic lip in cerebellar development.

Publication Date: 2010 Oct PMID: 20568288
Authors: Yu, T. - Huang, H. - Li, H. F.
Journal: J Neurosci Res

During cerebellar development, the chemokine stromal cell-derived factor-1 alpha (SDF-1 alpha) has been shown to play an important role in recruiting cells from the upper rhombic lip (URL) and external granule cell layer (EGL). However, its function in cerebellar development is still poorly understood. Our results have demonstrated that SDF-1 is necessary for EGL development, and URL cells stream to the SDF-1 source in vitro. Results of embryonic URL explant assays and transwell assays indicated that SDF-1 induces neural cell migration from the URL region in chemotactic and chemokinetic responses. The time-lapse results showed that the migration speed of granule cell progenitors out of the URL was accelerated by the addition of recombinant SDF-1 alpha. Collectively, our study shows that SDF-1 increases the motility of URL cells in the absence of a gradient and promotes the migration of granule cell progenitors during cerebellar development.

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Analysis of gene expression changes associated with long-lasting synaptic enhancement in hippocampal slice cultures after repetitive exposures to glutamate.

Publication Date: 2010 Oct PMID: 20568283
Authors: Kawaai, K. - Tominaga-Yoshino, K. - Urakubo, T. - Taniguchi, N. - Kondoh, Y. - Tashiro, H. - Ogura, A. - Tashiro, T.
Journal: J Neurosci Res

We have previously shown that repetitive exposures to glutamate (100 muM, 3 min, three times at 24-hr intervals) induced a long-lasting synaptic enhancement accompanied by synaptogenesis in rat hippocampal slice cultures, a phenomenon termed RISE (for repetitive LTP-induced synaptic enhancement). To investigate the molecular mechanisms underlying RISE, we first analyzed the time course of gene expression changes between 4 hr and 12 days after repetitive stimulation using an original oligonucleotide microarray: "synaptoarray." The results demonstrated that changes in the expression of synapse-related genes were induced in two time phases, an early phase of 24-96 hr and a late phase of 6-12 days after the third stimulation. Comprehensive screening at 48 hr after the third stimulation using commercially available high-density microarrays provided candidate genes responsible for RISE. From real-time PCR analysis of these and related genes, two categories of genes were identified, 1) genes previously reported to be induced by physiological as well as epileptic activity (bdnf, grm5, rgs2, syt4, ania4/carp/dclk) and 2) genes involved in cofilin-based regulation of actin filament dynamics (ywhaz, ssh1l, pak4, limk1, cfl). In the first category, synaptotagmin 4 showed a third stimulation-specific up-regulation also at the protein level. Five genes in the second category were coordinately up-regulated by the second stimulation, resulting in a decrease in cofilin phosphorylation and an enhancement of actin filament dynamics. In contrast, after the third stimulation, they were differentially regulated to increase cofilin phosphorylation and enhance actin polymerization, which may be a key step leading to the establishment of RISE.

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Deficiency of telomerase activity aggravates the blood-brain barrier disruption and neuroinflammatory responses in a model of experimental stroke.

Publication Date: 2010 Oct PMID: 20564349
Authors: Zhang, B. - Chen, L. - Swartz, K. R. - Bruemmer, D. - Eum, S. Y. - Huang, W. - Seelbach, M. - Choi, Y. J. - Hennig, B. - Toborek, M.
Journal: J Neurosci Res

Epidemiology and genetic studies indicate that patients with telomere length shorter than average are at higher risk of dying from heart disease or stroke. Telomeres are located at the ends of eukaryotic chromosomes, which demonstrate progressive length reduction in most somatic cells during aging. The enzyme telomerase can compensate for telomere loss during cell replication. The present study sought to investigate the contribution of telomerase to stroke and blood-brain barrier (BBB) dysfunction. Telomerase reverse transcriptase knockout (TERT(-/-)) mice and littermate controls with normal TERT expression were subjected to a 24-hr permanent middle cerebral artery occlusion (pMCAO). The stroke outcomes were assessed in terms of neurological scores and infarct volumes. In addition, we evaluated oxidative stress, permeability across the BBB, and integrity of tight junctions in brain microvessels. Neurological testing revealed that TERT(-/-) mice showed enhanced deficits compared with controls. These changes were associated with a greater infarct volume. The expression of tight junction protein ZO-1 decreased markedly in ischemic hemispheres of TERT(-/-) mice. The brain microvessels of TERT(-/-) mice also were more susceptible to oxidative stress, revealing higher superoxide and lower glutathione levels compared with mice with normal TERT expression. Importantly, TERT deficiency potentiated the production of inflammatory mediators, such as tumor necrosis factor-alpha, interleukin-1 beta, and intercellular adhesion molecule-1, in the ischemic hemispheres of mice with pMCAO. Our study suggests that TERT deficiency can predispose to the development of stroke in an experimental model of this disease.

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Involvement of nerve injury and activation of peripheral glial cells in tetanic sciatic stimulation-induced persistent pain in rats.

Publication Date: 2010 Oct PMID: 20544834
Authors: Liang, L. - Wang, Z. - Lu, N. - Yang, J. - Zhang, Y. - Zhao, Z.
Journal: J Neurosci Res

Tetanic stimulation of the sciatic nerve (TSS) produces long-lasting pain hypersensitivity in rats. Long-term potentiation (LTP) of C- and A-fiber-evoked field potentials in the spinal cord has been explored as contributing to central sensitization in pain pathways. However, the peripheral mechanism underlying TSS-induced pain hypersensitivity remains largely unknown. We investigated the effect of TSS on peripheral nerve and the expression of activating transcription factor 3 (ATF3) in dorsal root ganglion (DRG) as a marker of neuronal injury. TSS induced a mechanical allodynia for at least 35 days and induced ATF3 expression in the ipsilateral DRG. ATF3 is colocalized with NF200-labeled myelinated DRG neurons or CGRP- and IB4-labeled unmyelinated ones. Furthermore, we found that TSS induced Wallerian degeneration of sciatic nerve at the level of myelinisation by S100 protein (to label Schwann cells) immunohistochemistry, luxol fast blue staining, and electron microscopy. TSS also elicited the activation of satellite glial cells (SGCs) and enhanced the colocalization of GFAP and P2X7 receptors. Repeated local treatment with tetrodotoxin decreased GFAP expression in SGCs and behavioral allodynia induced by TSS. Furthermore, reactive microglia and astrocytes were found in the spinal dorsal horn after TSS. These results suggest that TSS-induced nerve injury and glial activation in the DRG and spinal dorsal horn may be involved in cellular mechanisms underlying the development of persistent pain after TSS and that TSS-induced nerve injury may be used as a novel neuropathic pain model.

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An angiotensin II type 1 receptor blocker can preserve endothelial function and attenuate brain ischemic damage in spontaneously hypertensive rats.

Publication Date: 2010 Oct PMID: 20544833
Authors: Oyama, N. - Yagita, Y. - Sasaki, T. - Omura-Matsuoka, E. - Terasaki, Y. - Sugiyama, Y. - Sakoda, S. - Kitagawa, K.
Journal: J Neurosci Res

Hypertension reduces endothelial nitric oxide synthase (eNOS) expression and leads to endothelial dysfunction. However, few studies have demonstrated the influences of hypertension on eNOS function in the cerebral cortex. The present study investigates the influences of hypertension on endothelial function in the cerebral cortex and the protective effects of antihypertensive agents against brain ischemia through the preservation of endothelial function. Five- and ten-week-old male Wistar rats and spontaneously hypertensive rats (SHR) were used for experiments. Five-week-old SHR received olmesartan, hydralazine, or vehicle for 5 weeks in drinking water. eNOS activation in the cerebral cortex was evaluated by analyzing levels of total and Ser(1177)-phosphorylated eNOS protein by Western blot. Blood pressure of 10-week-old SHR without treatment was clearly high, and the ratio of phospho-eNOS/total eNOS protein was significantly low. Five-week treatment with olmesartan or hydralazine suppressed the elevation of blood pressure and the reduction of phosphorylated eNOS-Ser(1177) in SHR, and olmesartan was more effective in maintaining phosphorylation of eNOS-Ser(1177) than hydralazine. To assess the contribution of eNOS to maintaining cerebral blood flow (CBF), we monitored CBF by laser-Doppler flowmetry after L-N(5)-(1-iminoethyl)ornithine (L-NIO) infusion. CBF response to L-NIO was preserved in olmesartan-treated SHR but not in hydralazine-treated SHR. Furthermore, infarct volume 48 hr after transient focal brain ischemia in olmesartan-treated SHR was significantly reduced compared with vehicle-treated SHR. These findings indicate that chronic prehypertensive treatment with olmesartan could attenuate brain ischemic injury through the maintenance of endothelial function in the cerebral cortex in SHR.

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Chronic caloric restriction reduces tissue damage and improves spatial memory in a rat model of traumatic brain injury.

Publication Date: 2010 Oct PMID: 20544832
Authors: Rich, N. J. - Van Landingham, J. W. - Figueiroa, S. - Seth, R. - Corniola, R. S. - Levenson, C. W.
Journal: J Neurosci Res

Although it has been known for some time that chronic caloric or dietary restriction reduces the risk of neurodegenerative disorders and injury following ischemia, the possible role of chronic restriction in improving outcomes after traumatic brain injury (TBI) has not been previously studied. Therefore, 2-month-old male Sprague-Dawley rats were divided into two dietary groups, an ad libitum fed group (AL) and a caloric-restriction group (CR) that was provided with 70% of the food intake of AL rats (n = 10/group). After 4 months, a weight-drop device (300 g) was used to produce a 2-mm bilateral medial frontal cortex contusion following craniotomy. Additional animals in each dietary group (n = 10) were used as sham-operated controls. The CR diet resulted in body weights that were reduced by 30% compared with AL controls. Not only did CR decrease the size of the cortical lesion after injury, there were marked improvements in spatial memory as measured by Morris water maze that included an increase in the number of animals successfully finding the platform as well as significantly reduced time to finding the hidden platform. Western analysis, used to examine the expression of proteins that play a role in neuronal survival, revealed significant increases in brain-derived neurotrophic factor (BDNF) in the cortical region around the site of injury and in the hippocampus in CR rats after injury. These findings suggest that molecular mechanisms involved in cell survival may play a role in reducing tissue damage and improving cognition after TBI and that these mechanisms can be regulated by dietary interventions.

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Amyloid-beta oligomers impair fear conditioned memory in a calcineurin-dependent fashion in mice.

Publication Date: 2010 Oct PMID: 20544830
Authors: Dineley, K. T. - Kayed, R. - Neugebauer, V. - Fu, Y. - Zhang, W. - Reese, L. C. - Taglialatela, G.
Journal: J Neurosci Res

Soluble oligomeric aggregates of the amyloid-beta (A beta) peptide are believed to be the most neurotoxic A beta species affecting the brain in Alzheimer disease (AD), a terminal neurodegenerative disorder involving severe cognitive decline underscored by initial synaptic dysfunction and later extensive neuronal death in the CNS. Recent evidence indicates that A beta oligomers are recruited at the synapse, oppose expression of long-term potentiation (LTP), perturb intracellular calcium balance, disrupt dendritic spines, and induce memory deficits. However, the molecular mechanisms behind these outcomes are only partially understood; achieving such insight is necessary for the comprehension of A beta-mediated neuronal dysfunction. We have investigated the role of the phosphatase calcineurin (CaN) in these pathological processes of AD. CaN is especially abundant in the CNS, where it is involved in synaptic activity, LTP, and memory function. Here, we describe how oligomeric A beta treatment causes memory deficits and depresses LTP expression in a CaN-dependent fashion. Mice given a single intracerebroventricular injection of A beta oligomers exhibited increased CaN activity and decreased pCREB, a transcription factor involved in proper synaptic function, accompanied by decreased memory in a fear conditioning task. These effects were reversed by treatment with the CaN inhibitor FK506. We further found that expression of hippocampal LTP in acutely cultured rodent brain slices was opposed by A beta oligomers and that this effect was also reversed by FK506. Collectively, these results indicate that CaN activation may play a central role in mediating synaptic and memory disruption induced by acute oligomeric A beta treatment in mice.

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Melatonin's protective action against ischemic neuronal damage is associated with up-regulation of the MT2 melatonin receptor.

Publication Date: 2010 Sep PMID: 20544829
Authors: Lee, C. H. - Yoo, K. Y. - Choi, J. H. - Park, O. K. - Hwang, I. K. - Kwon, Y. G. - Kim, Y. M. - Won, M. H.
Journal: J Neurosci Res

Melatonin is a potent free radical scavenger and antioxidant and has protective effects against ischemic damage. In the present study, we examined the relationship between the neuroprotective effects of melatonin and the activation of MT2 melatonin receptor in the hippocampal CA1 region (CA1) after transient cerebral ischemia. MT2 immunoreactivity and protein levels were increased in the CA1 after ischemic damage. Most of MT2-immunoreactive cells were colocalized with astrocytes, not microglia, in the ischemic CA1. In the melatonin-sham group, MT2 immunoreaction and protein levels were increased compared with the sham group, and MT2 immunoreactivity and its protein levels in the melatonin-ischemia group were similar to those in the melatonin-sham group. In addition, melatonin treatment attenuated the activation of astrocytes and microglia. These results indicate that MT2 are increased and expressed in astrocytes in the ischemic region after an ischemic insult. The activation of MT2 melatonin receptor in the CA1 after melatonin treatment may be involved in the neuroprotective effect associated with melatonin after ischemic injury.

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3R tau expression modifies behavior in transgenic mice.

Publication Date: 2010 Sep PMID: 20544828
Authors: Shiryaev, N. - Jouroukhin, Y. - Gozes, I.
Journal: J Neurosci Res

Tauopathy is a group of disorders characterized by the accumulation of hyperphosphorylated tau protein in the brain, resulting in dementia. Here, tau-related behavior was evaluated in a mouse model with brain overexpression of the shortest human tau isoform (0N3R). Two groups of animals [tau-transgenic (tau-tg) and control littermates] were tested for learning and memory at 1 and 7 months. In the Morris water maze, all mice learned the task at 1 month of age and did not learn at 7 months. In contrast, at 7 months, the tau-tg animals demonstrated better retention of the passive avoidance response compared with their control littermates, which did not learn. In the open field test, no differences were measured between transgenic and nontransgenic young mice, but significantly higher locomotion was observed in the 7-month-old tau-tg mice compared with controls. Behavior during the elevated plus maze test was the same at 1 month, but at 7 months increased entrance to the different arms was observed in the tau-tg group. Tau expression and phosphorylation levels were analyzed at 8 months. In the subcortical brain region associated with passive avoidance behavior, the tau-tg mice demonstrated increased brain tau expression coupled with reduced relative phosphorylation. In contrast, increased tau expression and phosphorylation were measured in the cerebral cortex of the tau-tg mice. In conclusion, 7-8-month-old tau-tg mice overexpressing nonmutated 0N3R human tau isoform demonstrated enhanced behavior in the passive avoidance test, paralleled by relative tau hypophosphorylation in the subcortical brain region.

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Corticotropin-releasing factor-2 activation prevents gentamicin-induced oxidative stress in cells derived from the inner ear.

Publication Date: 2010 Oct PMID: 20544827
Authors: Basappa, J. - Turcan, S. - Vetter, D. E.
Journal: J Neurosci Res

Generation of reactive oxygen species (ROS) is a common denominator in many conditions leading to cell death in the cochlea, yet little is known of the cochlea's endogenous mechanisms involved in preventing oxidative stress and its consequences in the cochlea. We have recently described a corticotropin-releasing factor (CRF) signaling system in the inner ear involved in susceptibility to noise-induced hearing loss. We use biochemical and proteomics assays to define further the role of CRF signaling in the response of cochlear cells to aminoglycoside exposure. We demonstrate that activity via the CRF(2) class of receptors protects against aminoglycoside-induced ROS production and activation of cell death pathways. This study suggests for the first time a role for CRF signaling in protecting the cochlea against oxidative stress, and our proteomics data suggest novel mechanisms beyond induction of free radical scavengers that are involved in its protective mechanisms.

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Effects of zona incerta lesions on striatal neurochemistry and behavioral asymmetry in 6-hydroxydopamine-lesioned rats.

Publication Date: 2010 Oct PMID: 20544826
Authors: Walker, R. H. - Davies, G. - Koch, R. J. - Haack, A. K. - Moore, C. - Meshul, C. K.
Journal: J Neurosci Res

Analysis of optimal sites for neurosurgical interventions in patients with Parkinson's disease (PD) suggests that significant clinical benefits may be achieved by involvement of the zona incerta (ZI). Unilateral electrolytic ZI lesions were made in intact and ipsilaterally 6-hydroxydopamine (6OHDA)-lesioned rats. Extracellular levels of glutamate, dopamine, and its metabolites in the ipsilateral striatum of awake rats were measured by using microdialysis, and tests of behavioral asymmetry were performed. In intact rats, ZI lesions had no effect on striatal extracellular glutamate or absolute levels of dopamine or metabolites, but dopamine metabolism decreased. After ZI lesions, contralateral forepaw use decreased in the forepaw adjusting steps test, but there was no change in response to vibrissa stimulation or cylinder exploration. There was no development of rotational asymmetry with amphetamine. In 6OHDA-lesioned rats, striatal extracellular glutamate levels were elevated compared with controls. ZI lesions reduced the increased levels of glutamate back to normal values. ZI lesions reduced dopamine and homovanillic acid levels and showed a trend toward a decrease in dopamine metabolism. 6OHDA-lesioned rats demonstrated the expected asymmetry of motor behaviors. After ZI lesions, ipsilateral turns following amphetamine injection were reduced, and there was a trend toward improved symmetry of forepaw use as determined with the forepaw adjusting steps test. There was no change in forepaw use with vibrissa stimulation or cylinder exploration. These data indicate that lesions of the ZI can affect striatal neurochemistry and motor behavioral asymmetry and suggest potential mechanisms by which ZI lesions may improve symptoms in PD.

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Glial cell line-derived neurotrophic factor-secreting genetically modified human bone marrow-derived mesenchymal stem cells promote recovery in a rat model of Parkinson's disease.

Publication Date: 2010 Sep PMID: 20544825
Authors: Glavaski-Joksimovic, A. - Virag, T. - Mangatu, T. A. - McGrogan, M. - Wang, X. S. - Bohn, M. C.
Journal: J Neurosci Res

Parkinson's disease (PD) is a neurodegenerative disease characterized by progressive degeneration of nigrostriatal dopaminergic (DA) neurons. The therapeutic potential of glial cell line-derived neurotrophic factor (GDNF), the most potent neurotrophic factor for DA neurons, has been demonstrated in many experimental models of PD. However, chronic delivery of GDNF to DA neurons in the brain remains an unmet challenge. Here, we report the effects of GDNF-releasing Notch-induced human bone marrow-derived mesenchymal stem cells (MSC) grafted into striatum of the 6-hydroxydopamine (6-OHDA) progressively lesioned rat model of PD. Human MSC, obtained from bone marrow aspirates of young, healthy adult volunteers, were transiently transfected with the intracellular domain of the Notch1 gene (NICD) to generate SB623 cells. SB623 cells expressing GDNF and/or humanized Renilla green fluorescent protein (hrGFP) following lentiviral transduction or nontransduced cells were stereotaxically placed into rat striatum 1 week after a unilateral partial 6-OHDA striatal lesion. At 4 weeks, rats that had received GDNF-transduced SB623 cells had significantly decreased amphetamine-induced rotation compared with control rats, although this effect was not observed in rats that received GFP-transduced or nontransduced SB623 cells. At 5 weeks, rejuvenated tyrosine hydroxylase-immunoreactive (TH-IR) fibers that appeared to be host DA axons were observed in and around grafts. This effect was more prominent in rats that received GDNF-secreting cells and was not observed in controls. These observations suggest that human bone-marrow derived MSC, genetically modified to secrete GDNF, hold potential as an allogeneic or autologous stem cell therapy for PD.

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Hypothyroid state does not protect but delays neuronal death in the hippocampal CA1 region following transient cerebral ischemia: Focus on oxidative stress and gliosis.

Publication Date: 2010 Sep PMID: 20544824
Authors: Lee, C. H. - Yoo, K. Y. - Hwang, I. K. - Choi, J. H. - Park, O. K. - Li, H. - Kang, I. J. - Kwon, Y. G. - Kim, Y. M. - Won, M. H.
Journal: J Neurosci Res

We investigated protective effects of hypothyroidism on delayed neuronal death, gliosis, lipid peroxidation and Cu,Zn-superoxide dismutase (SOD1) in the gerbil hippocampal CA1 region (CA1) after 5 min of transient cerebral ischemia. The hypothyroidism was induced by 0.025% methimazole treatment. Free triiodothyronine and thyroxine levels were markedly decreased in the hypothyroid group. Four days after ischemia/reperfusion, only a few NeuN-immunoreactive (+) neurons were detected in the CA1 of euthyroid-ischemia (eu-ischemia) group; however, at this time point, the number of NeuN(+) neurons was significantly higher in the hypothyroid-ischemia (hypo-ischemia) group than in the eu-ischemia group. At 5 days postischemia, NeuN(+) neurons were significantly decreased in the hypo-ischemia group: The number of NeuN(+) neurons in this group was similar to that in the eu-ischemia group. Activations of GFAP(+) astrocytes and Iba-1(+) microglia in the CA1 were higher in the eu-ischemia group 3 and 4 days after ischemia/reperfusion. At 5 days postischemia, the activations of both the glial cells in the CA1 were similar between the two groups. 4-Hydroxy-2-nonenal (HNE), a marker for lipid peroxidation, immunoreactivity in the eu-ischemia group was higher than in the hypo-ischemia group; at 5 days postischemia, the immunoreactivity was similar between the two groups. In contrast, SOD1 level was lower in the CA1 of the eu-ischemia group. These results suggest that hypothyroid state does not protect against delayed neuronal death but only delays the neuronal death in the hippocampal CA1 region after transient cerebral ischemia by reducing lipid peroxidation and increasing SOD1.

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Deletion of aquaporin-4 renders retinal glial cells more susceptible to osmotic stress.

Publication Date: 2010 Oct PMID: 20544823
Authors: Pannicke, T. - Wurm, A. - Iandiev, I. - Hollborn, M. - Linnertz, R. - Binder, D. K. - Kohen, L. - Wiedemann, P. - Steinhauser, C. - Reichenbach, A. - Bringmann, A.
Journal: J Neurosci Res

The glial water channel aquaporin-4 (AQP4) is implicated in the control of ion and osmohomeostasis in the sensory retina. Using retinal slices from AQP4-deficient and wild-type mice, we investigated whether AQP4 is involved in the regulation of glial cell volume under altered osmotic conditions. Superfusion of retinal slices with a hypoosmolar solution induced a rapid swelling of glial somata in tissues from AQP4 null mice but not from wild-type mice. The swelling was mediated by oxidative stress, inflammatory lipid mediators, and sodium influx into the cells and was prevented by activation of glutamatergic and purinergic receptors. Distinct inflammatory proteins, including interleukin-1 beta, interleukin-6, and inducible nitric oxide synthase, were up-regulated in the retina of AQP4 null mice compared with control, whereas cyclooxygenase-2 was down-regulated. The data suggest that water flux through AQP4 is involved in the rapid volume regulation of retinal glial (Muller) cells in response to osmotic stress and that deletion of AQP4 results in an inflammatory response of the retinal tissue. Possible implications of the data for understanding the pathophysiology of neuromyelitis optica, a human disease that has been suggested to involve serum antibodies to AQP4, are discussed.

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Modulation of the spinal excitability by muscle metabosensitive afferent fibers.

Publication Date: 2010 Sep PMID: 20544822
Authors: Laurin, J. - Dousset, E. - Decherchi, P.
Journal: J Neurosci Res

The aim of this study was to identify the effect of chemical activation of muscle metabosensitive afferent fibers from groups III and IV on Hoffmann (H-) reflex modulation in the vastus medialis muscle. The experiment was conducted in rats and was divided into two experiments. The first experiment consisted of recording the metabosensitive afferent activity from femoral nerve in rats in response to KCl intraarterial injections in nontreated adults and adults treated neonatally with capsaicin. Thus, the dose-response curve was determined. The second experiment consisted of eliciting the H- and M-waves before and after KCl injection in nontreated adult animals and those treated neonatally with capsaicin. Thus, the H(max)/M(max) ratio was measured. Results indicated that, 1) in nontreated animals, afferent fibers peak discharge was found after 10 mM KCl injection; 2) no significant increase in afferent discharge rate was found in capsaicin-treated animal after KCl injections, confirming that capsaicin is an excitotoxic agent that had destroyed the thin metabosensitive nerve fibers; 3) in nontreated animals, H(max)/M(max) ratio was significantly attenuated after a 10 mM KCl injection activating metabosensitive afferent fibers; and 4) in capsaicin-treated animals, no significant change in H(max)/M(max) ratio was observed after the KCl injection. These results reinforce the hypothesis that the spinal reflex response was influenced by metabosensitive muscle fibers and provide direct evidence that activation of these fibers could partially explain the reported decrease in H-reflex when metabolites are released in muscle.

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Altered sensitivity to excitotoxic cell death and glutamate receptor expression between two commonly studied mouse strains.

Publication Date: 2010 Sep PMID: 20544821
Authors: Finn, R. - Kovacs, A. D. - Pearce, D. A.
Journal: J Neurosci Res

Alterations in glutamatergic synapse function have been implicated in the pathogenesis of many different neurological disorders, including ischemia, epilepsy, Parkinson's disease, Alzheimer's disease, and Huntington's disease. While studying glutamate receptor function in juvenile Batten disease on the C57BL/6J and 129S6/S(v)E(v) mouse backgrounds, we noticed differences unlikely to be due to mutation difference alone. We report here that primary cerebellar granule cell cultures from C57BL/6J mice are more sensitive to N-methyl-D-aspartate (NMDA)-mediated cell death. Moreover, sensitivity to AMPA-mediated excitotoxicity is more variable and is dependent on the treatment conditions and age of the cultures. Glutamate receptor surface expression levels examined in vitro by in situ ELISA and in vivo by Western blot in surface cross-linked cerebellar samples indicated that these differences in sensitivity likely are due to strain-dependent differences in cell surface receptor expression levels. We propose that differences in glutamate receptor expression and in excitotoxic vulnerability should be taken into consideration in the context of characterizing disease models on the C57BL/6J and 129S6/S(v)E(v) mouse backgrounds.

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Chemical inducers and transcriptional markers of oligodendrocyte differentiation.

Publication Date: 2010 Sep PMID: 20544820
Authors: Joubert, L. - Foucault, I. - Sagot, Y. - Bernasconi, L. - Duval, F. - Alliod, C. - Frossard, M. J. - Pescini Gobert, R. - Curchod, M. L. - Salvat, C. - Nichols, A. - Pouly, S. - Rommel, C. - Roach, A. - Hooft van Huijsduijnen, R.
Journal: J Neurosci Res

Oligodendrocytes generate and maintain myelin, which is essential for axonal function and protection of the mammalian central nervous system. To advance our molecular understanding of differentiation by these cells, we screened libraries of pharmacologically active compounds and identified inducers of differentiation of Oli-neu, a stable cell line of mouse oligodendrocyte precursors (OPCs). We identified four broad classes of inducers, namely, forskolin/cAMP (protein kinase A activators), steroids (glucocorticoids and retinoic acid), ErbB2 inhibitors, and nucleoside analogs, and confirmed the activity of these compounds on rat primary oligodendrocyte precursors and mixed cortical cultures. We also analyzed transcriptional responses in the chemically induced mouse and rat OPC differentiation processes and compared these with earlier studies. We confirm the view that ErbB2 is a natural signaling component that is required for OPC proliferation, whereas ErbB2 inhibition or genetic knockdown results in OPC differentiation.

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Low levels of mutant ubiquitin are degraded by the proteasome in vivo.

Publication Date: 2010 Aug 15 PMID: 20336771
Authors: van Tijn, P. - Verhage, M. C. - Hobo, B. - van Leeuwen, F. W. - Fischer, D. F.
Journal: J Neurosci Res

The ubiquitin-proteasome system fulfills a pivotal role in regulating intracellular protein turnover. Impairment of this system is implicated in the pathogenesis of neurodegenerative diseases characterized by ubiquitin- containing proteinaceous deposits. UBB(+1), a mutant ubiquitin, is one of the proteins accumulating in the neuropathological hallmarks of tauopathies, including Alzheimer's disease, and polyglutamine diseases. In vitro, UBB(+1) properties shift from a proteasomal ubiquitin-fusion degradation substrate at low expression levels to a proteasome inhibitor at high expression levels. Here we report on a novel transgenic mouse line (line 6663) expressing low levels of neuronal UBB(+1). In these mice, UBB(+1) protein is scarcely detectable in the neuronal cell population. Accumulation of UBB(+1) commences only after intracranial infusion of the proteasome inhibitors lactacystin or MG262, showing that, at these low expression levels, the UBB(+1) protein is a substrate for proteasomal degradation in vivo. In addition, accumulation of the protein serves as a reporter for proteasome inhibition. These findings strengthen our proposition that, in healthy brain, UBB(+1) is continuously degraded and disease-related UBB(+1) accumulation serves as an endogenous marker for proteasomal dysfunction. This novel transgenic line can give more insight into the intrinsic properties of UBB(+1) and its role in neurodegenerative disease.

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