bluuebunny/crossref_metadata_2025_split huggingface dataset:
Two rat brain Staufen isoforms differentially bind RNA
Authors: Monshausen Michaela , Putz Ulrich , Rehbein Monika , Schweizer Michaela , DesGroseillers Luc , Kuhl, Dietmar , Richter Dietmar , Kindler Stefan 
Published: March 2003

In neurones, a limited number of mRNAs is found in dendrites, including transcripts encoding the microtubule‐associated protein 2 (MAP2). Recently, we identified a cis‐acting dendritic targeting element (DTE) in MAP2 mRNAs. Here we used the yeast tri‐hybrid system to identify potential trans‐acting RNA‐binding factors of the DTE. A cDNA clone was isolated that encodes a member of a mammalian protein family that is highly homologous to the Drosophila RNA‐binding protein Staufen. Mammalian Staufen appears to be expressed in most tissues and brain areas. Two distinct rat brain Staufen isoforms, rStau+I6 and rStau‐I6, are encoded by alternatively spliced mRNAs. Both isoforms contain four double‐stranded RNA‐binding domains (dsRBD). In the larger rStau+I6 isoform, six additional amino acids are inserted in the second dsRBD. Although both isoforms interacted with the MAP2‐DTE and various additional RNA fragments in an in vitro north‐western assay, rStau‐I6 exhibited a stronger signal of bound radioactively labelled RNAs as compared with rStau+I6. Using an antibody directed against mammalian Staufen, the protein was detected in somata and dendrites of neurones of the adult rat hippocampus and cerebral cortex. Ultrastructural studies revealed that in dendrites, rat Staufen accumulates along microtubules. Thus in neurones, rat Staufen may serve to link RNAs to the dendritic microtubular cytoskeleton and may thereby regulate their subcellular localization.
Inducible nitric oxide synthase expression in brain cortex after acute restraint stress is regulated by nuclear factor κB‐mediated mechanisms
Authors: Madrigal José L. M. , Moro María A. , Lizasoain Ignacio , Lorenzo Pedro , Castrillo Antonio , Boscá Lisardo , Leza Juan C. 
Published: March 2003

The underlying mechanisms by which physical or psychological stress causes neurodegeneration are still unknown. We have demonstrated that the high‐output and long‐lasting synthesizing source of nitric oxide (NO), inducible NO synthase (iNOS), is expressed in brain cortex during stress and that its overexpression accounts for the neurodegenerative changes seen after 3 weeks of repeated stress. Now we have found that acute stress (restraint for 6 h) increases the activity of a calcium‐independent NOS and induces the expression of iNOS in brain cortex in adult male rats. In order to elucidate the possible mechanisms involved in this induction, we studied the role of transcription nuclear factor κB (NF‐κB), which is required for iNOS synthesis. We have observed that an acute restraint stress session stimulates the translocation of the NF‐κB to the nucleus after 4 h and that the administration of the NF‐κB inhibitor pyrrolidine dithiocarbamate [PDTC, 75 and 150 mg/kg intraperitoneally (i.p.)] at the onset of stress inhibits the stress‐induced increase in iNOS expression. Since glutamate release and subsequent NMDA (N‐methyl‐d‐aspartate) receptor activation has been recognized as an early change after exposure to stressful stimuli, and glutamate has been shown to induce iNOS in brain via a NF‐κB‐dependent mechanism, we studied the possible role of excitatory amino acids in the induction of iNOS in our model. Pretreatment with the NMDA receptor antagonist dizocilpine (MK‐801, 0.1 and 0.3 mg/kg i.p.) inhibits the stress‐induced NF‐κB activation as well as the stress‐induced increase in iNOS expression. Taken together, these findings indicate that excitatory amino acids and subsequent activation of NF‐κB account for stress‐induced iNOS expression in cerebral cortex, and support a possible neuroprotective role for specific inhibitors in this situation.
Superoxide dismutase activity in organotypic midbrain–striatum co‐cultures is associated with resistance of dopaminergic neurons to excitotoxicity
Authors: Katsuki Hiroshi , Tomita Michiko , Takenaka Chikako , Shirakawa Hisashi , Shimazu Seiichiro , Ibi Masakazu , Kume Toshiaki , Kaneko Shuji , Akaike Akinori 
Published: March 2003

We have previously demonstrated that dopaminergic neurons in midbrain–striatum slice co‐cultures are more resistant to NMDA cytotoxicity than the same neuronal population in single midbrain slice cultures. Here, we show that dopaminergic neurons in midbrain–striatum co‐cultures also exhibit resistance to the cytotoxicity of nitric oxide donors, 2,2′‐(hydroxynitrosohydrazono)bis‐ethanamine (NOC‐18) and 3‐morpholinosydnonimine (SIN‐1). The cytotoxicity of NMDA (30 µm) in single cultures was significantly attenuated by the nitric oxide synthase (NOS) inhibitor Nω‐nitro‐l‐arginine (100 µm), whereas the toxicity in co‐cultures was not. The levels of tyrosine residue nitration of tyrosine hydroxylase, a hallmark of the occurence of peroxynitrite anion in dopaminergic neurons, were lower in co‐cultures than those in single cultures. Single cultures and co‐cultures did not show appreciable differences in the number or distribution of NOS‐containing neurons as assessed by NADPH diaphorase histochemistry. On the other hand, midbrain slices cultured with striatal slices showed higher levels of superoxide dismutase (SOD) activity as well as increased protein levels of Cu,Zn‐SOD, than midbrain slices cultured alone. These results suggested that the generation of NO is involved in NMDA cytotoxicity on dopaminergic neurons, and that increased activity of SOD in co‐cultures renders dopaminergic neurons resistant to NMDA cytotoxicity by preventing the formation of peroxynitrite.
Ca2+ changes induced by different presynaptic nicotinic receptors in separate populations of individual striatal nerve terminals
Authors: Nayak Sanjay V. , Dougherty John J. , McIntosh J. Michael , Nichols Robert A. 
Published: March 2003

Presynaptic nicotinic acetylcholine receptors likely play a modulatory role in the nerve terminal. Using laser‐scanning confocal microscopy, we have characterized physiological responses obtained on activation of presynaptic nicotinic receptors by measuring calcium changes in individual nerve terminals (synaptosomes) isolated from the rat corpus striatum. Nicotine (500 nm) induced Ca2+ changes in a subset (10–25%) of synaptosomes. The Ca2+ responses were dependent on extracellular Ca2+ and desensitized very slowly (several minutes) on prolonged exposure to agonist. The nicotine‐induced Ca2+ responses were dose‐dependent and were completely blocked by dihydro‐β‐erythroidine (5 µm), differentially affected by mecamylamine (10 µm) and α‐conotoxin MII (100 nm), and not affected by α‐bungarotoxin (500 nm). Immunocytochemical studies using well‐characterized monoclonal antibodies revealed the presence of the α4 and α3/α5 nicotinic subunits. The nicotine‐induced responses were unaffected by prior depolarization or by a mixture of Ca2+ channel toxins including ω‐conotoxin MVIIC (500 nm), ω‐conotoxin GVIA (500 nm) and agatoxin TK (200 nm). Our results indicate that nicotinic receptors present on striatal nerve terminals induce Ca2+ entry largely without involving voltage‐gated Ca2+ channels, most likely by direct permeation via the receptor channel itself. In addition, at least two subpopulations of presynaptic nicotinic receptors reside on separate terminals in the striatum, suggesting distinct modulatory roles.
Hypoxia regulates glutamate metabolism and membrane transport in rat PC12 cells
Authors: Kobayashi Shuichi , Millhorn David E. 
Published: March 2003

We investigated the effect of hypoxia on glutamate metabolism and uptake in rat pheochromocytoma (PC12) cells. Various key enzymes relevant to glutamate production, metabolism and transport were coordinately regulated by hypoxia. PC12 cells express two glutamate‐metabolizing enzymes, glutamine synthetase (GS) and glutamate decarboxylase (GAD), as well as the glutamate‐producing enzyme, phosphate‐activated glutaminase (PAG). Exposure to hypoxia (1% O2) for 6 h or longer increased expression of GS mRNA and protein and enhanced GS enzymatic activity. In contrast, hypoxia caused a significant decrease in expression of PAG mRNA and protein, and also decreased PAG activity. In addition, hypoxia led to an increase in GAD65 and GAD67 protein levels and GAD enzymatic activity. PC12 cells express three Na+‐dependent glutamate transporters; EAAC1, GLT‐1 and GLAST. Hypoxia increased EAAC1 and GLT‐1 protein levels, but had no effect on GLAST. Chronic hypoxia significantly enhanced the Na+‐dependent component of glutamate transport. Furthermore, chronic hypoxia decreased cellular content of glutamate, but increased that of glutamine. Taken together, the hypoxia‐induced changes in enzymes related to glutamate metabolism and transport are consistent with a decrease in the extracellular concentration of glutamate. This may have a role in protecting PC12 cells from the cytotoxic effects of glutamate during chronic hypoxia.
Enhancement of 3,4‐methylenedioxymethamphetamine neurotoxicity by the energy inhibitor malonate
Authors: Nixdorf W. L. , Burrows K. B. , Gudelsky G. A. , Yamamoto B. K. 
Published: March 2003

The acute and long‐term effects of the local perfusion of 3,4‐methylenedioxymethamphetamine (MDMA) and the interaction with the mitochondrial inhibitor malonate (MAL) were examined in the rat striatum. MDMA, MAL or the combination of MAL with MDMA was reverse dialyzed into the striatum for 8 h via a microdialysis probe while extracellular dopamine (DA) and serotonin (5‐HT) were measured. One week later, tissue immediately surrounding the probe was assayed for DA and 5‐HT tissue content. Local perfusion of MDMA increased DA and 5‐HT release but did not produce long‐term depletion of DA or 5‐HT in tissue. Malonate also increased both DA and 5‐HT release but, in contrast to MDMA, produced only long‐term depletion of DA. The combined perfusion of MDMA/MAL synergistically increased the release of DA and 5‐HT and produced long‐term depletion of both DA and 5‐HT in tissue. These results support the conclusion that DA, compared with 5‐HT, neurons are more susceptible to mitochondrial inhibition. Moreover, MDMA, which does not normally produce DA depletion in the rat, exacerbated MAL‐induced DA depletions. The effect of MDMA in combination with MAL to produce 5‐HT depletion suggests a role for bio‐energetic stress in MDMA‐induced toxicity to 5‐HT neurons. Overall, these results highlight the importance of energy balance to the function of DA and 5‐HT neurons and to the toxic effects of MDMA.
[35S]GTPγS autoradiography reveals a wide distribution of Gi/o‐linked ADP receptors in the nervous system: close similarities with the platelet P2YADP receptor
Authors: Laitinen Jarmo T. , Uri Asko , Raidaru Gerda , Miettinen Riitta 
Published: March 2003

No Gi‐linked P2Y receptors have been cloned to date but the presence of such receptors is thought to be restricted to platelets and certain clonal cell lines. Using the functional approach of [35S]guanosine 5′‐[γ‐thio]‐triphosphate autoradiography, we uncovered the widespread presence of such receptors in the CNS. Under conditions in which the prominent signal due to tonic adenosine receptor activity is masked, ADP and ATP stimulated G‐protein activity in multiple grey and white matter regions. Localization in the grey matter suggests inhibitory auto‐/heteroreceptor function. In the white matter, activated G proteins appeared as ‘hot spots’ (presumed oligodendrocyte progenitors) with scattered distribution along the main fibre tracts. Responses to ATP were diminished under conditions that inhibited degradation, suggesting that prior conversion to ADP explained agonist action. Uracil nucleotides were ineffective but 2‐methylthio‐ADP activated G proteins ≈ 500‐fold more potently than ADP, although both were similarly degraded. Throughout the brain, ADP‐dependent G‐protein activity was reversed by 2‐hexylthio‐AdoOC(O)Asp2, a non‐phosphate ATP analogue, whereas selective P2Y1 receptor antagonists proved ineffective. A similar receptor was also disclosed from the adrenal medulla. These data witness a hitherto unrecognized abundance of Gi/o‐linked ADP receptors in the nervous system. Biochemical and pharmacological behaviour suggests striking similarities to the elusive platelet P2YADP receptor.
CEP‐1347/KT‐7515, an inhibitor of SAPK/JNK pathway activation, promotes survival and blocks multiple events associated with Aβ‐induced cortical neuron apoptosis
Authors: Bozyczko‐Coyne Donna , O'Kane Teresa M. , Wu Zhi‐Liang , Dobrzanski Pawel , Murthy Seetha , Vaught Jeffry L. , Scott Richard W. 
Published: March 2003

Although the mechanism of neuronal death in Alzheimer's disease (AD) has yet to be elucidated, a putative role for c‐junin this process has emerged. Thus, it was of interest to delineate signal transduction pathway(s) which regulate the transcriptional activity of c‐jun, and relate these to alternate gene inductions and biochemical processes associated with beta‐amyloid (Aβ) treatment. In this regard, the survival promoting activity of CEP‐1347, an inhibitor of the stress‐activated/c‐junN‐terminal (SAPK/JNK) kinase pathway, was evaluated against Aβ‐induced cortical neuron deathin vitro. Moreover, CEP‐1347 was used as a pharmacologic probe to associate multiple biochemical events with Aβ‐induced activation of the SAPK/JNK pathway. CEP‐1347 promoted survival and blocked Aβ‐induced activation of JNK kinase (MKK4, also known as MEK‐4, JNKK and SEK1) as well as other downstream events associated with JNK pathway activation. CEP‐1347 also blocked Aβ‐induction of cyclinD1andDP5genes and blocked Aβ‐induced increases in cytoplasmic cytochrome c, caspase 3‐like activity and calpain activation. The critical time window for cell death blockade by CEP‐1347 resided within the peak of Aβ‐induced MKK4 activation, thus defining this point as the most upstream event correlated to its survival‐promoting activity. Together, these data link the SAPK/JNK pathway and multiple biochemical events associated with Aβ‐induced neuronal death and further delineate the point of CEP‐1347 interception within this signal transduction cascade.
Testosterone‐mediated neuroprotection through the androgen receptor in human primary neurons
Authors: Hammond Jennifer , Le Quynh , Goodyer Cynthia , Gelfand Morrie , Trifiro Mark , LeBlanc Andrea 
Published: March 2003

Estrogen is an active neuroprotectant and is presently investigated as a potential therapy against Alzheimer's disease for women. To determine if male hormones could also be neuroprotective, we investigated the effect of testosterone, methyltestosterone, and epitestosterone at physiological concentrations on primary cultures of human neurons induced to undergo apoptosis by serum deprivation. Serum deprivation significantly induces neuronal apoptosis in a protracted fashion. As expected, physiological concentrations of 17‐β‐estradiol and transcriptionally inactive 17‐α‐estradiol protect neurons against apoptosis. Similar to 17‐β‐estradiol, physiological concentrations of testosterone are also neuroprotective. Androgen receptors are present at 8 ± 2 fmol/mg protein in the neuron cultures. The non‐aromatizable androgen, mibolerone, is also neuroprotective and aromatase inhibitor, 4‐androsten‐4‐OL‐3,17‐dione, does not prevent testosterone‐mediated neuroprotection. In contrast, anti‐androgen, flutamide, eliminates testosterone‐mediated neuroprotection. Testosterone analog, methyltestosterone, showed androgen receptor‐dependent neuroprotection that was delayed in time indicating that a metabolite may be the active agent. The endogenous anti‐androgen, epitestosterone, also showed a slight neuroprotective effect but not through the androgen receptor. These results indicate that androgens induce neuroprotection directly through the androgen receptor. These data suggest that androgens may also be of therapeutic value against Alzheimer's disease in aging males.
Electrical stimulation of cerebellar fastigial nucleus protects rat brain, in vitro, from staurosporine‐induced apoptosis
Authors: Zhou Ping , Qian Liping , Glickstein Sara B. , Golanov Eugene V. , Pickel Virginia M. , Reis Donald J. 
Published: March 2003

Electrical stimulation of the cerebellar fastigial nucleus (FN) elicits a prolonged (∼10 days) and substantial (50–80%) protection against ischemic and excitotoxic injuries. The mechanism(s) of protection are unknown. We investigated whether FN stimulation directly protects brain cells against apoptotic cell death in an in vitro rat brain slice culture model. Rats were electrically stimulated in FN or, as control, the cerebellar dentate nucleus (DN). Coronal slices through the forebrain were explanted, exposed to staurosporine, harvested, and analyzed for caspase‐3 activity by a fluorescence assay. FN, but not DN, stimulation significantly reduced staurosporine‐induced caspase‐3 activity by 39 ± 7% at 3 h, 31 ± 3% at 6 h and 26 ± 4% at 10 h of incubation. Immunocytochemistry revealed FN‐specific reductions in activated caspase‐3 mainly in glial‐like cells throughout the forebrain. FN stimulation also results in a 56.5% reduction in cytochrome c release upon staurosporine incubation. We conclude that neuroprotection elicited from FN stimulation can directly modify the sensitivity of brain cells to apoptotic stimuli and thereby suppress staurosporine induced apoptosis in adult rat brain slices. This model indicates that neuroprotection can be studied in vitro and provides new insight into the potential role of glial cells in ischemic protection of neurons induced by FN stimulation.
Repression of myelin proteolipid protein gene expression is mediated through both general and cell type‐specific negative regulatory elements in nonexpressing cells
Authors: Li Shenyang , Dobretsova Anna , Kokorina Natalia A. , Wight Patricia A. 
Published: March 2003

AbstractThe myelin proteolipid protein gene (Plp) is expressed primarily in oligodendrocytes. Yet how the gene remains repressed in nonexpressing cells has not been defined, and potentially could cause adverse effects in an organism if the mechanism for repression was impaired. Previous studies suggest that the first intron contains element(s), which suppress expression in nonexpressing cells, although the identity of these elements within the 8 kb intron was not characterized. Here we report the localization of multiple negative regulatory elements that repress Plp gene expression in nonexpressing cells (+/+ Li). Two of these elements (regions) correspond to those used by Plp expressing cells (N20.1), whilst another acts in a cell type‐specific manner (i.e. operational in +/+ Li liver cells, but not N20.1 cells). By gel‐shift and DNase I footprinting analyses, the factor(s) that bind to the cell type‐specific negative regulatory region appear to be far more abundant in +/+ Li cells than in N20.1 cells. Thus, Plp gene repression is mediated through the combinatorial action of both ‘general’ and cell type‐specific negative regulatory elements. Additionally, repression in +/+ Li cells cannot be overcome via an antisilencer/enhancer element, which previously has been shown to function in N20.1 cells.
Glial cell line‐derived neurotrophic factor up‐regulates the expression of tyrosine hydroxylase gene in human neuroblastoma cell lines
Authors: Xiao Hengyi , Hirata, Yoko , Isobe Ken‐ichi , Kiuchi Kazutoshi 
Published: March 2003

AbstractThe role of glial cell line‐derived neurotrophic factor (GDNF) in the survival of dopaminergic neurons has been well documented, but its effect on dopamine biosynthesis remains to be elucidated. In this study, the effect of GDNF on the gene expression of tyrosine hydroxylase (TH), the rate‐limiting enzyme of dopamine biosynthesis, was investigated. We found that GDNF elevated the expression of the TH gene at both mRNA and protein levels in TGW cells, a human neuroblastoma cell line. GDNF significantly enhances the transcription rate of the TH gene as actinomycin D prevented the induction of TH mRNA and GDNF increased the activity of the TH promoter. In addition, GDNF exerts a relatively weak but significant effect on the stability of TH mRNA, because GDNF delayed the degradation of TH mRNA and strengthened a special TH mRNA/protein interaction known to be relevant with TH mRNA stability. By comparing several human neurogenic cell lines, we found that GDNF‐induced TH expression was only observed in the cells possessing Ret protein and coincided with the expression levels. Taken together, these results indicate that GDNF up‐regulates the expression of the TH gene by promoting the transcription of the TH gene and the stability of TH mRNA with the Ret receptor dependency in some neuroblastoma cell lines. Thus, GDNF exerts its neurotrophic role not only in promoting cells survival, but also in affecting dopamine biosynthesis.
X‐Linked inhibitor of apoptosis protein is involved in mutant SOD1‐mediated neuronal degeneration
Authors: Ishigaki Shinsuke , Liang Yideng , Yamamoto Masahiko , Niwa Jun‐ichi , Ando Yoshio , Yoshihara Tsuyoshi , Takeuchi Hideyuki , Doyu Manabu , Sobue Gen 
Published: March 2003

AbstractMutations in the superoxide dismutase 1 (SOD1) gene cause the degeneration of motor neurons in familial amyotrophic lateral sclerosis (FALS). An apoptotic process including caspase‐1 and ‐3 has been shown to participate in the pathogenesis of FALS transgenic (Tg) mouse model. Here we report that IAP proteins, potent inhibitors of apoptosis, are involved in the FALS Tg mouse pathologic process. The levels of X‐linked inhibitor of apoptosis protein (XIAP) mRNA and protein were significantly decreased in the spinal cord of symptomatic G93A‐SOD1 Tg mice compared with littermates. In contrast, the levels of cIAP‐1 mRNA and protein were increased in symptomatic G93A‐SOD1 Tg mice, whereas the levels of cIAP‐2 mRNA and protein were unchanged. In situ hybridization showed that the expression of XIAP was remarkably reduced in the motor neurons of Tg mice, and the expression of cIAP‐1 was strongly increased in the reactive astrocytes of Tg mice. Overexpression of XIAP markedly inhibited the cell death and caspase‐3 activity in the neuro2a cells expressing mutant SOD1. Deletional mutant analysis revealed that the N‐terminal domain of XIAP, the BIR1‐2 domains, was essential for this inhibitory activity. These results suggest that XIAP plays a role in the apoptotic mechanism in the progression of disease in mutant SOD1 Tg mice and holds therapeutic possibilities for FALS.
Gene expression profile after intense second messenger activation in cortical primary neurones
Authors: Mayer Peter , Ammon Susanne , Braun Holger , Tischmeyer Helga , Riechert Uta , Kahl Evelyn , Höllt Volker 
Published: March 2003

AbstractNumerous stimuli induce immediate early gene (IEG) expression in neurones, but a comprehensive overview of the late‐response genes is lacking. Therefore we aimed to identify changes in the neuronal gene expression profile following intense stimulation. Forskolin and 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA), direct activators of intracellular second messengers, were applied to primary cultured cortical neurones. The gene expression profiles were analyzed on Affymetrix DNA chips which cover around 8000 rat genes. Out of these, 95 genes (1.2%) were increased at least three‐fold, and 43 genes (0.5%) were at least three‐fold decreased. The gene chip results were verified by testing 15 of the altered genes by quantitative real‐time PCR. The majority of the up‐regulated genes were transcription factors, neurotrophic factors or (putative) neuropeptides. Furthermore, there were marked changes in intracellular signal processing enzymes and in postsynaptic structural proteins (e.g. vesl, arc, narp), which have been implicated in synaptic plasticity. Notably, classical players in neurotransmission or plasticity such as glutamate and GABA receptors or voltage‐gated ion channels were not increased. It is likely that the increased production of components of intracellular signalling and of postsynaptic proteins is involved in neuronal plasticity.