bluuebunny/crossref_metadata_2025_split huggingface dataset:
Neuroserpin regulates neurite outgrowth in nerve growth factor‐treated PC12 cells
Authors: Parmar Parmjeet K. , Coates Leigh C. , Pearson John F. , Hill Rena M. , Birch Nigel P. 
Published: March 2003

AbstractNeuroserpin is a serine protease inhibitor widely expressed in the developing and adult nervous systems and implicated in the regulation of proteases involved in processes such as synaptic plasticity, neuronal migration and axogenesis. We have analysed the effect of neuroserpin on growth factor‐induced neurite outgrowth in PC12 cells. We show that small changes in neuroserpin expression result in changes to the number of cells extending neurites and total neurite length following NGF treatment. Increased expression of neuroserpin resulted in a decrease in the number of cells extending neurites and a reduction in total free neurite length whereas reduced levels of neuroserpin led to a small increase in the number of neurite extending cells and a significant increase in total free neurite length compared to the parent cell line. Neuroserpin also altered the response of PC12 cells to bFGF and EGF treatment. Neuroserpin was localised to dense cored secretory vesicles in PC12 cells but was unable to complex with its likely enzyme target, tissue plasminogen activator at the acidic pH found in these vesicles. These data suggest that modulation of neuroserpin levels at the extending neurite growth cone may play an important role in regulating axonal growth.
Altered chemokine expression in the spinal cord and brain contributes to differential interleukin‐1β‐induced neutrophil recruitment
Authors: Campbell Sandra J. , Wilcockson David C. , Butchart Angus G. , Perry V. Hugh , Anthony Daniel C. 
Published: March 2003

AbstractThe pattern of neutrophil recruitment that accompanies inflammation in the CNS depends on the site of injury and the stage of development. The adult brain parenchyma is refractory to neutrophil recruitment and associated damage as compared to the spinal cord or juvenile brain. Using quantitative Taqman RT–PCR and enzyme‐liked immunosorbent assay (ELISA), we compared mRNA and protein expression of the rat neutrophil chemoattractant chemokines (CINC) in spinal cord and brain of adult and juvenile rats to identify possible association with the observed differences in neutrophil recruitment. Interleukin‐1β (IL‐1β) injection resulted in up‐regulated chemokine expression in both brain and spinal cord. CINC‐3 mRNA was elevated above CINC‐1 and CINC‐2α, with expression levels for each higher in spinal cord than in brain. By ELISA, IL‐1β induced greater CINC‐1 and CINC‐2α expression compared to CINC‐3, with higher protein levels in spinal cord than in brain. In the juvenile brain, significantly higher levels of CINC‐2α protein were observed in response to IL‐1β injection than in the adult brain following an equivalent challenge. Correspondingly, neutrophil recruitment was observed in the juvenile brain and adult spinal cord, but not in the adult brain. No expression of CINC‐2β mRNA was detected. Thus differential chemokine induction may contribute to variations in neutrophil recruitment in during development and between the different CNS compartments.
Identity of adenylyl cyclase isoform determines the G protein mediating chronic opioid‐induced adenylyl cyclase supersensitivity
Authors: Ammer Hermann , Christ Thomas E. 
Published: March 2003

AbstractTo determine the intracellular signal transduction pathway responsible for the development of tolerance/dependence, the ability of Gzα to substitute for pertussis toxin (PTX)‐sensitive G proteins in mediating adenylyl cyclase (AC) supersensitivity was examined in the presence of defined AC isoforms. In transiently µ‐opioid receptor (OR) transfected COS‐7 cells (endogenous inhibitory G proteins: Giα2, Giα3 and Gzα), neither acute (1 µmol/L) nor chronic morphine treatment (1 µmol/L; 18 h) influenced intracellular cAMP production. Coexpression of the µ‐OR together with AC type V and VI fully restored the ability of morphine to acutely inhibit cAMP generation. Chronic morphine treatment further resulted in the development of tolerance/dependence, as assessed by desensitization of the acute inhibitory opioid effect (tolerance) as well as the induction of AC supersensitivity after drug withdrawal (dependence). Specific direction of µ‐OR signalling via Gzα by both PTX treatment and Gzα over‐expression had no effect on chronic morphine regulation of AC type V, but completely abolished the development of tolerance/dependence with AC type VI. Similar results were obtained in stably µ‐OR‐expressing HEK293 cells transiently cotransfected with Gzα and either AC type V or VI. Coprecipitation studies further verified that Gzα specifically binds to AC type V but not type VI. Taken together, these results demonstrate that in principle each of the OR‐activated G proteins per se is able to mediate AC supersensitivity. However, they also indicate that it is the molecular nature of AC isoform that selects and determines the OR‐activated G protein mediating tolerance/dependence.
TorsinA and heat shock proteins act as molecular chaperones: suppression of α‐synuclein aggregation
Authors: McLean Pamela J. , Kawamata Hibiki , Shariff Saadat , Hewett Jeffrey , Sharma Nutan , Ueda Kenji , Breakefield Xandra O. , Hyman Bradley T. 
Published: March 2003

AbstractTorsinA, a protein with homology to yeast heat shock protein104, has previously been demonstrated to colocalize with α‐synuclein in Lewy bodies, the pathological hallmark of Parkinson's disease. Heat shock proteins are a family of chaperones that are both constitutively expressed and induced by stressors, and that serve essential functions for protein refolding and/or degradation. Here, we demonstrate that, like torsinA, specific molecular chaperone heat shock proteins colocalize with α‐synuclein in Lewy bodies. In addition, using a cellular model of α‐synuclein aggregation, we demonstrate that torsinA and specific heat shock protein molecular chaperones colocalize with α‐synuclein immunopositive inclusions. Further, overexpression of torsinA and specific heat shock proteins suppress α‐synuclein aggregation in this cellular model, whereas mutant torsinA has no effect. These data suggest that torsinA has chaperone‐like activity and that the disease‐associated GAG deletion mutant has a loss‐of‐function phenotype. Moreover, these data support a role for chaperone proteins, including torsinA and heat shock proteins, in cellular responses to neurodegenerative inclusions.
Proteasomal inhibition by misfolded mutant superoxide dismutase 1 induces selective motor neuron death in familial amyotrophic lateral sclerosis
Authors: Urushitani Makoto , Kurisu Junko , Tsukita Kayoko , Takahashi Ryosuke 
Published: March 2003

AbstractAccumulating evidence indicates that abnormal conformation of mutant superoxide dismutase 1 (SOD1) is an essential feature underlying the pathogenesis of mutant SOD1‐linked familial amyotrophic lateral sclerosis (ALS). Here we investigated the role of ubiquitin‐proteasome pathway in the mutant SOD1‐related cell death and the effect of oxidative stress on the misfolding of mutant SOD1. Transient overexpression of ubiquitin with human SOD1 (wild‐type, ala4val, gly85arg, gly93ala) in Neuro2A cells decreased the amount of mutant SOD1, but not of wild‐type, while only mutants were co‐immunoprecipitated with poly‐ubiquitin. Proteasome inhibition by lactacystin augmented accumulation of mutant SOD1 in the non‐ionic detergent‐insoluble fraction. The spinal cord lysates from mutant SOD1 transgenic mice showed multiple carbonylated proteins, including mutant SOD1 with SDS‐resistant dimer formation. Furthermore, the treatment of hSOD1‐expressing cells with hydrogen peroxide promoted the oligomerization, and detergent‐insolubility of mutant SOD1 alone, and the oxidized mutant SOD1 proteins were more heavily poly‐ubiquitinated. In Neuro2A cells stably expressing human SOD1 protein, the proteasome function measured by chymotrypsin‐like activity, was decreased over time without a quantitative alteration of the 20S proteasomal component. Finally, primary motor neurons from the mouse embryonic spinal cord were more vulnerable to lactacystin than non‐motor neurons. These results indicate that the sustained expression of mutant SOD1 leads to proteasomal inhibition and motor neuronal death, which in part explains the pathogenesis of mutant SOD1‐linked ALS.
Norepinephrine alters the expression of genes involved in neuronal sprouting and differentiation: relevance for major depression and antidepressant mechanisms
Authors: Laifenfeld Daphna , Klein Ehud , Ben‐Shachar Dorit 
Published: March 2003

AbstractRecent research into depression has focused on the involvement of long‐term intracellular processes, leading to abnormal neuronal plasticity in brains of depressed patients, and reversed by antidepressant treatment. Given a suggested decrease in noradrenergic transmission in depression, and an antidepressant induced increase in norepinephrine (NE) level, a possible role for NE in mediating alterations in neuronal morphology and plasticity was examined. Human neuroblastoma SH‐SY5Y cells treated with 10−5 m NE presented an elongated granule‐rich cell‐body and increased number of neurites, when compared with non‐treated cells. Moreover, cell survival was enhanced in the presence of NE, while proliferation was inhibited. The above effects suggest a role for NE in cell differentiation. Indeed similar effects on cell survival and neurite outgrowth were induced in SH‐SY5Y cells by retinoic acid (RA), an established differentiating agent. Finally, NE treatment resulted in a progressive decrease in the pluripotent marker Oct4 and an increase in the neuronal growth cone marker, growth‐associated‐protein 43 (GAP‐43). Alongside these effects, NE‐treated cells presented alterations in the expression of 44 genes as observed in a neurobiology cDNA microarray. Among the altered genes, an increase in the expression level of two neurite‐outgrowth promoting genes, neural cell adhesion molecule L1 and laminin, was confirmed by RT‐PCR. Taken together, the results support a role for NE in processes of synaptic connectivity, and may point to a role for this neurotransmitter in mediating the suggested neuronal plasticity in depression and in antidepressant treatment.
Progesterone enhances ethanol‐induced modulation of mesocortical dopamine neurons: antagonism by finasteride
Authors: Dazzi Laura , Serra Mariangela , Seu Emanuele , Cherchi Giulia , Pisu M. Giuseppina , Purdy Robert H. , Biggio Giovanni 
Published: March 2003

AbstractThe effect of endogenous 3α‐hydroxy‐5α‐pregnan‐20‐one (3α,5α‐TH PROG) on the modulation of mesocortical dopamine extracellular concentration by ethanol was investigated by microdialysis in rats. Intraperitoneal injection of progesterone (5 mg/kg, once a day for 5 days) increased the cortical content of 3α,5α‐TH PROG and potentiated the biphasic effect of acute intraperitoneal administration of ethanol on dopamine content. A dose of ethanol (0.25 g/kg) that was ineffective in naïve rats induced a 55% increase in dopamine extracellular concentration in rats pretreated with progesterone. This increase was similar to that induced by a higher dose (0.5 g/kg) of ethanol in naïve rats. Administration of ethanol at 0.5 g/kg to progesterone‐pretreated rats inhibited dopamine content by an extent similar to that observed with an even higher dose (1 g/kg) in naïve rats. The administration of the 5α‐reductase inhibitor finasteride (25 mg/kg, subcutaneous), together with progesterone, prevented the effects of the latter, both on the cortical concentration of 3α,5α‐TH PROG and on the modulation by ethanol of dopamine content. These data suggest that 3α,5α‐TH PROG contributes to the action of ethanol on the mesocortical dopaminergic system. They also suggest that physiological fluctuations in the brain concentrations of neuroactive steroids associated with the oestrous cycle, menopause, pregnancy and stress may alter the response of mesocortical dopaminergic neurons to ethanol.
Quantitative relationship between inhibition of respiratory complexes and formation of reactive oxygen species in isolated nerve terminals
Authors: Sipos Ildikó , Tretter Laszlo , Adam‐Vizi Vera 
Published: March 2003

AbstractIn this study reactive oxygen species (ROS) generated in the respiratory chain were measured and the quantitative relationship between inhibition of the respiratory chain complexes and ROS formation was investigated in isolated nerve terminals. We addressed to what extent complex I, III and IV,respectively, should be inhibited to cause ROS generation. For inhibition of complex I, III and IV, rotenone, antimycin and cyanide were used, respectively, and ROS formation was followed by measuring the activity of aconitase enzyme. ROS formation was not detected until complex III was inhibited by up to 71 ± 4%, above that threshold inhibition, decrease in aconitase activity indicated an enhanced ROS generation. Similarly, threshold inhibition of complex IV caused anaccelerated ROS production. By contrast, inactivation of complex I to a small extent (16 ± 2%) resulted in a significant increase in ROS formation, and no clear threshold inhibition could be determined. However, the magnitude of ROS generated at complex I when it is completely inhibited is smaller than that observed when complex III or complex IV was fully inactivated. Our findings may add a novel aspect to the pathology of Parkinson's disease, showing that a moderate level of complex I inhibition characteristic in Parkinson's disease leads to significant ROS formation. The amount of ROS generated by complex I inhibition is sufficient to inhibit in situ the activity of endogenous aconitase.
Degraded myelin‐associated glycoprotein (dMAG) formation from pure human brain myelin‐associated glycoprotein (MAG) is not mediated by calpain or cathepsin L‐like activities
Authors: Päiväläinen Satu , Suokas Marko , Lahti Outi , Heape Anthony M. 
Published: March 2003

AbstractThe myelin‐associated glycoprotein (MAG) is a transmembrane cell adhesion molecule participating in myelin formation and maintenance. Calcium‐activated/‐dependent proteolysis of myelin‐associated glycoprotein by calpain and cathepsin L‐like activities has already been detected in purified myelin fractions, producing a soluble fragment, called degraded (d)MAG, characterized by the loss of the transmembrane and cytoplasmic domains. Here, we demonstrate and analyze dMAG formation from pure human brain myelin‐associated glycoprotein. The activity never exhibited the high rate previously reported in human myelin fractions. Degradation is time‐, temperature‐, buffer‐ and structure‐dependent, is inhibited at 4°C and by denaturation of the sample, and is mediated by a trans‐acting factor. There is no strict pH dependency of the proteolysis. Degradation was inhibited by excess aprotinin, but not by 1–10 µg/mL aprotinin and was not eliminated by the use of an aprotinin‐sepharose matrix during the purification. dMAG formation was not enhanced by calcium, nor inhibited by a wide variety of protease inhibitors, including specific calpain and cathepsin L inhibitors. Therefore, while cysteine proteases may be present in human myelin membrane fractions, they are not involved in dMAG formation from highly purified human brain myelin‐associated glycoprotein preparations.
Transients in acetylcholine receptor site density and degradation during reinnervation of mouse sternomastoid muscle
Authors: Szabo Maria , Salpeter Edwin E. , Randall William , Salpeter Miriam M. 
Published: March 2003

AbstractThe degradation rates of acetylcholine receptors (AchRs) were evaluated at the neuromuscular junction during and just after reinnervation of denervated muscles. When mouse sternomastoid muscles are denervated by multiple nerve crush, reinnervation begins 2–4 days later and is complete by day 7–9 after the last crush. In fully innervated muscles, the AChR degradation rate is stable and slow (t1/2 ≈ 10 days), whereas after denervation the newly inserted receptors degrade rapidly (t1/2 ≈ 1.2 days). The composite profile of degradation, which a mixture of the stable and the rapid receptors would give, is not observed during reinnervation. Instead, the receptors inserted between 2.5 and 7.5 days after the last crush all have an intermediate degradation rate of t1/2 ≈ 3.7 days with standard error ± 0.3 days. The total receptor site density at the endplate was evaluated during denervation and during reinnervation. As predicted theoretically, the site density increased substantially, but temporarily, after denervation. An analogous deleterious substantial decrease in density would be expected during reinnervation, without the intermediate receptor. This decrease is not observed, however, because of a large insertion rate at intermediate times (3000 ± 700 receptor complexes per µm2 per day). The endplate density of receptors thus remains relatively constant.
Far‐upstream elements are dispensable for tissue‐specific proenkephalin expression using a Cre‐mediated knock‐in strategy
Authors: Le Yunzheng , Gagneten Sara , Larson Teresa , Santha Edit , Dobi Albert , Agoston Denes v. , Sauer Brian 
Published: March 2003

AbstractSeveral cis‐regulatory DNA elements are present in the 5′ upstream regulatory region of the enkephalin gene (ENK) promoter. To determine their role in conferring organ‐specificity of ENK expression in mice and to circumvent the position effects from random gene insertion that are known to often frustrate such analysis in transgenic mice, we used a Cre‐mediated gene knock‐in strategy to target reporter constructs to a ‘safe haven’loxP‐tagged locus in the hypoxanthine phosphoribosyltransferase (HPRT) gene. Here we report reliable and reproducible reporter gene expression under the control of the 5′ upstream regulatory region of the mouse ENK gene in gene‐modified mice using this Cre‐mediated knock‐in strategy. Comparison of two 5′ENK regulatory regions (one with and the other without known cis‐regulatory DNA elements) in the resulting adult mice showed that conserved far‐upstream cis‐regulatory DNA elements are dispensable for correct organ‐specific gene expression. Thus the proximal 1.4 kb of the murine ENK promoter region is sufficient for organ‐specificity of ENK gene expression when targeted to a safe‐haven genomic locus. These results suggest that conservation of the far‐upstream DNA elements serves more subtle roles, such as the developmental or cell‐specific expression of the ENK gene.
Nicotine trapping causes the persistent desensitization of α4β2 nicotinic receptors expressed in oocytes
Authors: Jia Li , Flotildes Karen , Li Maureen , Cohen Bruce N. 
Published: March 2003

AbstractTo determine whether prolonged nicotine exposure persistently inactivates rat α4β2 nicotinic receptors expressed in Xenopus oocytes, we measured the voltage‐clamped α4β2 response to acetylcholine (ACh) before and 24 h after, 1‐h or 12‐h incubations in 10 µm nicotine. A 12‐h incubation in 10 µm nicotine depressed the α4β2 ACh response for 24 h without affecting total or surface α4β2 expression. To determine whether oocyte‐mediated nicotine release caused this depression, we co‐incubated an α4β2‐expressing oocyte with an un‐injected one (pre‐incubated in 10 µm nicotine for 12 h) for 24 h and measured the change in the α4β2 ACh response. The response decreased by the same factor after the co‐incubation as it did after a 12‐h incubation in 10 µm nicotine and a 24‐h incubation in nicotine‐free media. Thus, oocyte‐mediated nicotine release caused the persistent desensitization we observed after a 12‐h incubation in 10 µm nicotine. Consistent with this result, measurements of [3H]nicotine release show that oocytes release enough nicotine into the wash media to desensitize α4β2 receptors and that prolonged incubation in 300 µm ACh (which cannot readily cross the membrane or accumulate in acidic vesicles) did not persistently depress the α4β2 response.
BDNF heightens the sensitivity of motor neurons to excitotoxic insults through activation of TrkB
Authors: Hu Peter , Kalb Robert G. 
Published: March 2003

AbstractThe survival promoting and neuroprotective actions of brain‐derived neurotrophic factor (BDNF) are well known but under certain circumstances this growth factor can also exacerbate excitotoxic insults to neurons. Prior exploration of the receptor through which BDNF exerts this action on motor neurons deflects attention away from p75. Here we investigated the possibility that BDNF acts through the receptor tyrosine kinase, TrkB, to confer on motor neurons sensitivity to excitotoxic challenge. We blocked BDNF activation of TrkB using a dominant negative TrkB mutant or a TrkB function blocking antibody, and found that this protected motor neurons against excitotoxic insult in cultures of mixed spinal cord neurons. Addition of a function blocking antibody to BDNF to mixed spinal cord neuron cultures is also neuroprotective indicating that endogenously produced BDNF participates in vulnerability to excitotoxicity. We next examined the intracellular signaling cascades that are engaged upon TrkB activation. Previously we found that inhibition of the phosphatidylinositide‐3′‐kinase (PI3′K) pathway blocks BDNF‐induced excitotoxic sensitivity. Here we show that expression of a constitutively active catalytic subunit of PI3′K, p110, confers excitotoxic sensitivity (ES) upon motor neurons not incubated with BDNF. Parallel studies with purified motor neurons confirm that these events are likely to be occuring specifically within motor neurons. The abrogation of BDNF's capacity to accentuate excitotoxic insults may make it a more attractive neuroprotective agent.
Analysis of phospholipid molecular species in brains from patients with infantile and juvenile neuronal‐ceroid lipofuscinosis using liquid chromatography‐electrospray ionization mass spectrometry
Authors: Käkelä Reijo , Somerharju Pentti , Tyynelä Jaana 
Published: March 2003

AbstractPhospholipids (PL) in cerebral cortex from patients with infantile (INCL or CLN1) and juvenile (JNCL or CLN3) forms of neuronal ceroid‐lipofuscinosis (NCL) and controls were analysed by normal phase HPLC and on‐line electrospray ionization ion‐trap mass spectrometric detection (LC‐ESI‐MS). The method provided quantitative data on numerous molecular species of different PL classes, which are not achieved by using the conventional chromatographic methods. Compared with the controls, the INCL brains contained proportionally more phosphatidylcholine (PC), and less phosphatidylethanolamine (PE) and phosphatidylserine (PS). Different molecular species of PC, PE, PS, phosphatidylinositol and sphingomyelin were quantified using multiple internal PL standards that differed in fatty acyl chain length and thus allowed correction for chain length dependency of instrument response. In INCL cortex, which had lost 65% of the normal PL content, the proportions of polyunsaturated molecular species, especially the PS and PE that contained docosahexaenoic acid (22:6n‐3), were dramatically decreased. The membranes may have adapted to this alteration by increasing the proportions of PL molecules substituted with monounsaturated and short‐chain fatty acids. Lysobisphosphatidic acid was highly elevated in the INCL brain and consisted mostly of polyunsaturated species. It is possible that changes in the composition of PL membranes accelerate progression of INCL by altering signalling and membrane trafficking in neurons.