bluuebunny/crossref_metadata_2025_split huggingface dataset:
Modulation of electrically evoked acetylcholine release in cultured rat septal neurones
Authors: Ehret Andreas , Haaf Andreas , Jeltsch Hélène , Heimrich Bernd , Feuerstein Thomas J. , Jackisch Rolf 
Published: March 2003

The electrically evoked release of acetylcholine and its modulation via auto‐ and heteroreceptors were studied in primary cell cultures prepared from embryonic rat septum (ED 17). Cultures were grown for 1, 2 or 3 weeks on circular, poly D‐lysine‐coated glass coverslips. They developed a dense network of non‐neuronal and neuronal cells, only some of which were immunopositive for choline acetyltransferase. To measure acetylcholine release, the cells on the coverslips were pre‐incubated with [3H]choline (0.1 µmol/L), superfused with modified Krebs–Henseleit buffer at 25°C and electrically stimulated twice for 2 min (S1, S2; 3 Hz, 0.5 ms, 90–100 mA). The electrically evoked overflow of [3H] from the cells consisted of ≈ 80% of authentic [3H]Ach, was largely Ca2+‐dependent and tetrodotoxin sensitive, and hence represents an action potential‐evoked, exocytotic release of acetylcholine. Using pairs of selective agonists and antagonists added before S2, muscarinic autoreceptors, as well as inhibitory adenosine A1‐ and opioid µ‐receptors, could be detected, whereas δ‐opioid receptors were not found. Evoked [3H] overflow from cultures grown for 1 week, although Ca2+ dependent and tetrodotoxin sensitive, was insensitive to the muscarinic agonist oxotremorine, whereas the effect of oxotremorine on cells grown for 3 weeks was even more pronounced than that in 2‐week‐old cultures. In conclusion, similar to observations on rat septal tissue in vivo, acetylcholine release from septal cholinergic neurones grown in vitro is inhibited via muscarinic, adenosine A1 and µ‐opioid receptors. This in vitro model may prove useful in the exploration of regulatory mechanisms underlying the expression of release modulating receptors on septal cholinergic neurones.
Sustained extracellular signal‐regulated kinase activation by 6‐hydroxydopamine: implications for Parkinson's disease
Authors: Kulich Scott M. , Chu Charleen T. 
Published: March 2003

Although the toxin 6‐hydroxydopamine (6‐OHDA) is utilized extensively in animal models of Parkinson's disease, the underlying mechanism of its toxic effects on dopaminergic neurons is not completely understood. We examined the effects of 6‐OHDA on the CNS‐derived tyrosine hydroxylase expressing B65 cell line, with particular attention to the regulation of the extracellular signal‐regulated protein kinases (ERK). 6‐OHDA elicited a dose‐dependent cytotoxicity in B65 cells. Toxic doses of 6‐OHDA also elicited a biphasic pattern of ERK phosphorylation with a prominent sustained phase, a pattern that differed from that observed with hydrogen peroxide (H2O2) treatment. 6‐OHDA‐elicited ERK phosphorylation was blocked by PD98059, an inhibitor of the upstream mitogen activated protein kinase kinase (MEK) that phosphorylates and activates ERK. PD98059 also conferred protection against 6‐OHDA cytotoxicity, but did not affect H2O2 toxicity in B65 cells. These results suggest that ERK activation plays a direct mechanistic role in 6‐OHDA toxicity, rather than representing a protective compensatory response, and raise the possibility that abnormal patterns of ERK activation may contribute to dopaminergic neuronal cell death.
Brain ischemia and reperfusion activates the eukaryotic initiation factor 2α kinase, PERK
Authors: Kumar Rita , Azam Salman , Sullivan Jonathan M. , Owen Cheri , Cavener Douglas R. , Zhang Peichuan , Ron David , Harding Heather P. , Chen Jane‐Jane , Han Anping , White Blaine C. , Krause Gary S. , DeGracia Donald J. 
Published: March 2003

Reperfusion after global brain ischemia results initially in a widespread suppression of protein synthesis in neurons, which persists in vulnerable neurons, that is caused by the inhibition of translation initiation as a result of the phosphorylation of the α‐subunit of eukaryotic initiation factor 2 (eIF2α). To identify kinases responsible for eIF2α phosphorylation [eIF2α(P)] during brain reperfusion, we induced ischemia by bilateral carotid artery occlusion followed by post‐ischemic assessment of brain eIF2α(P) in mice with homozygous functional knockouts in the genes encoding the heme‐regulated eIF2α kinase (HRI), or the amino acid‐regulated eIF2α kinase (GCN2). A 10‐fold increase in eIF2α(P) was observed in reperfused wild‐type mice and in the HRI–/– or GCN2–/– mice. However, in all reperfused groups, the RNA‐dependent protein kinase (PKR)‐like endoplasmic reticulum eIF2α kinase (PERK) exhibited an isoform mobility shift on SDS–PAGE, consistent with the activation of the kinase. These data indicate that neither HRI nor GCN2 are required for the large increase in post‐ischemic brain eIF2α(P), and in conjunction with our previous report that eIF2α(P) is produced in the brain of reperfused PKR–/– mice, provides evidence that PERK is the kinase responsible for eIF2α phosphorylation in the early post‐ischemic brain.
Interleukin‐1β up‐regulates expression of neurofilament light in human neuronal cells
Authors: Rempel Hans , Kusdra Leonard , Pulliam Lynn 
Published: March 2003

Elevated expression of interleukin‐1 (IL‐1β), a pro‐inflammatory cytokine secreted by activated microglia, is a pathogenic marker of numerous neurodegenerative processes including Alzheimer's disease (AD). We have characterized a link between IL‐1β and the 68‐kDa neurofilament light (NF‐L) protein, which is a major component of the neuronal cytoskeleton. Using human brain aggregate cultures, we found that IL‐1β treatment significantly increased NF‐L expression in primary neurons. Analysis of mRNA levels demonstrated elevated NF‐L expression within 72 h while imaging of neurons by immunofluorescent staining for NF‐L confirmed IL‐1β‐induced NF‐L protein expression. These observations suggest a potential inflammatory‐induced mechanism for deregulation of an important cytoskeletal protein, NF‐L, possibly leading to neuronal dysfunction.
l‐DOPA and glia‐conditioned medium have additive effects on tyrosine hydroxylase expression in human catecholamine‐rich neuroblastoma NB69 cells
Authors: Rodríguez‐Martín E. , Canals S. , Casarejos M. J. , de Bernardo S. , Handler A. , Mena M. A. 
Published: March 2003

The aim of this study was to investigate the effect of l‐DOPA and glia‐conditioned medium (GCM) on cell viability, tyrosine hydroxylase (TH) expression, dopamine (DA) metabolism and glutathione (GSH) levels of NB69 cells. l‐DOPA (200 µm) induced differentiation of NB69 cells of more than 4 weeks in vitro, as shown by phase‐contrast microscopy and TH immunocytochemistry, and decreased replication, as shown by 5‐bromodeoxyuridine immunostaining. l‐DOPA did not increase the number of necrotic or apoptotic cells, as shown by morphological features, Trypan Blue, lactate dehydrogenase activity, bis‐benzimide staining and TUNEL assay. Furthermore, l‐DOPA (200 µm) increased Bcl‐xL protein expression. Incubation of cells with l‐DOPA (50, 100, 200 µm) for 24 h resulted in an increase in TH protein levels (174, 196 and 212% versus control). Neither carbidopa, an inhibitor of l‐aromatic amino acid decarboxylase enzyme, nor l‐buthionine sulfoximine, which inhibits GSH synthesis, or ascorbic acid, an antioxidant, blocked the l‐DOPA‐induced effect on TH protein expression. l‐DOPA (0, 50, 100 and 200 µm) plus GCM further increased the amount of TH protein (346, 446, 472 and 424%). l‐DOPA (200 µm) increased TH protein levels to 132, 191 and 245% of controls after incubation for 24, 48 and 72 h. DA metabolism in NB69 cells was increased in cultures treated with either l‐DOPA (200–300 µm) or GCM and these two agents had a synergistic effect on DA metabolism. In addition, l‐DOPA (200 µm) or/and GCM‐treated cells increased their GSH extracellular levels (223, 257, 300% of controls) after 48 h of treatment. The l‐DOPA‐induced increase of TH protein expression in NB69 cells was independent of DA production, free radicals and GSH up‐regulation.
Semaphorin4F interacts with the synapse‐associated protein SAP90/PSD‐95
Authors: Schultze Wiebke , Eulenburg Volker , Lessmann Volkmar , Herrmann Lutz , Dittmar Thomas , Gundelfinger Eckart D. , Heumann Rolf , Erdmann Kai S. 
Published: March 2003

Semaphorins are a family of secreted and membrane‐associated proteins involved in growth cone guidance during development. Here, we describe the interaction of Semaphorin4F (Sema4F) with the post‐synaptic density protein SAP90/PSD‐95. Using the yeast two‐hybrid system and coprecipitation assays we were able to show an interaction between the extreme C‐terminus of Sema4F and the PDZ domains of SAP90/PSD‐95. Heterologous coexpression of a chimeric EphrinB1/Semaphorin4F protein with SAP90/PSD‐95 in COS cells leads to translocation of SAP90/PSD‐95 from the cytosol to the membrane. Deletion analysis shows that this translocation activity of Sema4F is completely dependent on the presence of the last three C‐terminal amino acids. In addition, Sema4F immunoreactivity is present in synaptosome fractions and enriched in post‐synaptic density fractions. Consistently, in cultured hippocampal neurons, we demonstrate punctate colocalization of Sema4F and SAP90/PSD‐95 in dendrites, furthermore we found colocalization of Sema4F with synapsin1 suggesting a synaptic localization. Our data implicate a new functional context for semaphorins at glutamatergic synapses.
The role of 5‐HT1B receptors in the regulation of serotonin cell firing and release in the rat brain
Authors: Adell Albert , Celada Pau , Artigas Francesc 
Published: March 2003

The release of 5‐HT in terminal areas of the rodent brain is regulated by 5‐HT1B receptors. Here we examined the role of 5‐HT1B receptors in the control of 5‐HT output and firing in the dorsal raphe nucleus (DR), median raphe nucleus (MnR) and forebrain of the rat in vivo. The local perfusion (30–300 µm) of the selective 5‐HT1B receptor agonist CP‐93,129 to freely moving rats decreased 5‐HT release in the DR and more markedly in the MnR. Likewise, 300 µm CP‐93,129 reduced 5‐HT output in substantia nigra pars reticulata, ventral pallidum, lateral habenula and the suprachiasmatic nucleus. The effect of CP‐93,129 was prevented by SB‐224289, but not by WAY‐100635, selective 5‐HT1B and 5‐HT1A receptor antagonists, respectively. SB‐224289 did not alter dialysate 5‐HT in any raphe nuclei. The intravenous administration of the brain‐penetrant selective 5‐HT1B receptor agonist CP‐94,253 (0.5–2.0 mg/kg) to anesthetized rats decreased dialysate 5‐HT in dorsal hippocampus and globus pallidus, increased it in MnR and left it unaltered in the DR and medial prefrontal cortex. SB‐224289, at a dose known to block 5‐HT1B autoreceptor‐mediated effects (5 mg/kg), did not prevent the effect of CP‐94,253 on MnR 5‐HT. The intravenous administration of CP‐94,253 (0.05–1.6 mg/kg) to anesthetized rats increased the firing rate of MnR, but not DR‐5‐HT neurons. The local perfusion of CP‐94,253 in the MnR showed a biphasic effect, with 5‐HT reductions at 0.3–3 µm and increase at 300 µm. These results suggest that 5‐HT cell firing and release in midbrain raphe nuclei (particularly in the MnR) are under control of 5‐HT1B receptors. The activation of 5‐HT1B autoreceptors (possibly located on 5‐HT nerve endings and/or varicosities within DR and MnR) reduces 5‐HT release. The effects of higher concentrations of 5‐HT1B receptor agonists seem more compatible with the activation of 5‐HT1B heteroreceptors on inhibitory neurons.
Oxidative inactivation of calcineurin by Cu,Zn superoxide dismutase G93A, a mutant typical of familial amyotrophic lateral sclerosis
Authors: Ferri Alberto , Gabbianelli Roberta , Casciati Arianna , Celsi Fulvio , Rotilio Giuseppe , Carrì Maria Teresa 
Published: March 2003

Calcineurin is a serine/threonine phosphatase involved in a wide range of cellular responses to calcium mobilizing signals. Previous evidence supports the notion of the existence of a redox regulation of this enzyme, which might be relevant for neurodegenerative processes, where an imbalance between generation and removal of reactive oxygen species could occur. In a recent work, we have observed that calcineurin activity is depressed in two models for familial amyotrophic lateral sclerosis (FALS) associated with mutations of the antioxidant enzyme Cu,Zn superoxide dismutase (SOD1), namely in neuroblastoma cells expressing either SOD1 mutant G93A or mutant H46R and in brain areas from G93A transgenic mice.In this work we report that while wild‐type SOD1 has a protective effect, calcineurin is oxidatively inactivated by mutant SOD1s in vitro; this inactivation is mediated by reactive oxygen species and can be reverted by addition of reducing agents. Furthermore, we show that calcineurin is sensitive to oxidation only when it is in an ‘open’, calcium‐activated conformation, and that G93A‐SOD1 must have its redox‐active copper site available to substrates in order to exert its pro‐oxidant properties on calcineurin. These findings demonstrate that both wild‐type and mutant SOD1s can interfere directly with calcineurin activity and further support the possibility of a relevant role for calcineurin‐regulated biochemical pathways in the pathogenesis of FALS.
The degenerative effect of a single intranigral injection of LPS on the dopaminergic system is prevented by dexamethasone, and not mimicked by rh‐TNF‐α, IL‐1β and IFN‐γ
Authors: Castaño A. , Herrera A. J. , Cano J. , Machado A. 
Published: March 2003

AbstractIt is becoming widely accepted that the inflammatory response is involved in neurodegenerative disease. In this context, we have developed an animal model of dopaminergic system degeneration by the intranigral injection of lipopolysaccharide (LPS), a potent inductor of inflammation. To address the importance of the inflammatory response in the LPS‐induced degeneration of nigral dopaminergic neurones, we carried out two different kinds of studies: (i) the possible protective effect of an anti‐inflammatory compound, and (ii) the effect of the intranigral injection of inflammatory cytokines (TNF‐α, IL‐1β and IFN‐γ) on dopaminergic neurones viability. Present results show that dexamethasone, a potent anti‐inflammatory drug that interferes with many of the features characterizing pro‐inflammatory glial activation, prevented the loss of catecholamine content, Tyrosine hydroxylase (TH) activity and TH immunostaining induced by LPS‐injection and also the bulk activation of microglia/macrophages. Surprisingly, injection of the pro‐inflammatory cytokines failed to reproduce the LPS effect. Taken together, our results suggest that inflammatory response is implicated in LPS‐induced neurodegeneration. This damage may be due, at least in part, to a cascade of events independent of that described for TNF‐α/IL‐1β/IFN‐γ.
p75 neurotrophin receptor is required for constitutive and NGF‐induced survival signalling in PC12 cells and rat hippocampal neurones
Authors: Bui Nguyen Truc , König Hans‐Georg , Culmsee Carsten , Bauerbach Elke , Poppe Monika , Krieglstein Josef , Prehn Jochen H. M. 
Published: March 2003

AbstractWe have previously shown that nerve growth factor (NGF)‐induced activation of nuclear factor‐κB increased neuronal expression of Bcl‐xL, an anti‐apoptotic Bcl‐2 family protein. In the present study we determined the role of the p75 neurotrophin receptor in constitutive and NGF‐induced survival signalling. Treatment of rat pheochromocytoma (PC12) cells with a blocking anti‐rat p75 antibody or inhibition of p75 expression by antisense oligonucleotides reduced constitutive and NGF‐induced bcl‐xL expression. Treatment with the blocking anti‐p75 antibody also inhibited NGF‐induced activation of the survival kinase Akt. Inhibition of phosphatidylinositol‐3‐kinase (PI3 kinase) activity or overexpression of a dominant‐negative mutant of Akt kinase inhibited NGF‐induced nuclear factor‐κB activation. Activation of Akt kinase by NGF was also observed in PC12nnr5 cells and cultured rat hippocampal neurones which both lack significant TrkA expression. Treatment of hippocampal neurones with the blocking anti‐p75 antibody inhibited constitutive and NGF‐induced Bcl‐xL expression, activation of Akt, and blocked the protective effect of NGF against excitotoxic and apoptotic injury. Our data suggest that the p75 neurotrophin receptor mediates constitutive and NGF‐induced survival signalling in PC12 cells and hippocampal neurones, and that these effects are mediated via the PI3‐kinase pathway.
Reduced number of unmyelinated sensory axons in peripherin null mice
Authors: Larivière R. C. ,   M. D. Nguyen,, Ribeiro‐da‐Silva A. , Julien J.‐P. 
Published: March 2003

AbstractPeripherin is a type III intermediate filament (IF) abundantly expressed in developing neurons, but in the adult, it is primarily found in neurons extending to the peripheral nervous system. It has been suggested that peripherin may play a role in axonal elongation and/or cytoskeletal stabilization during development and regeneration. To further clarify the function of peripherin, we generated and characterized mice with a targeted disruption of the peripherin gene. The peripherin null mice were viable, reproduced normally and did not exhibit overt phenotypes. Microscopic analysis revealed no gross morphological defects in the ventral and dorsal roots, spinal cord, retina and gut, but protein analyses showed increased levels of the type IV IF α‐internexin in ventral roots of peripherin null mice. Whereas the number and caliber of myelinated motor and sensory axons in the L5 roots remained unchanged in peripherin knockout mice, there was a substantial reduction (∼ 34%) in the number of L5 unmyelinated sensory fibers that correlated with a decreased binding of the lectin IB4. These results demonstrate a requirement of peripherin for the proper development of a subset of sensory neurons.
NF‐κB‐mediated up‐regulation of Bcl‐XSand Bax contributes to cytochromecrelease in cyanide‐induced apoptosis
Authors: Shou Yan , Li Nianyu , Li Li , Borowitz Joseph L. , Isom Gary E. 
Published: March 2003

AbstractCyanide induces apoptosis through cytochromecactivated caspase cascade in primary cultured cortical neurons. The underlying mechanism for cytochromecrelease from mitochondria after cyanide treatment is still unclear. In this study, the roles of endogenous Bcl‐2 proteins in cyanide‐induced apoptosis were investigated. After cyanide (100–500 µm) treatment for 24 h, two pro‐apoptotic Bcl‐2 proteins, Bcl‐XSand Bax were up‐regulated as shown by western blot and RT‐PCR analysis. The expression levels of two antiapoptotic Bcl‐2 proteins, Bcl‐2 and Bcl‐XL, remained unchanged after cyanide treatment, whereas the mRNA levels of Bcl‐XSand Bax began to increase within 2 h and their protein levels increased 6 h after treatment. NF‐κB, a redox‐sensitive transcription factor activated after cyanide treatment, is responsible for the up‐regulation of Bcl‐XSand Bax. SN50, which is a synthetic peptide that blocks translocation of NF‐κB from cytosol to nucleus, inhibited the up‐regulation of Bcl‐XSand Bax. Similar results were obtained using a specific κB decoy DNA. NMDA receptor activation and reactive oxygen species (ROS) generation are upstream events of NF‐κB activation, as blockade of these two events by MK801,l‐NAME or PBN inhibited cyanide‐induced up‐regulation of Bcl‐XSand Bax. Up‐regulation of pro‐apoptotic Bcl‐XSand Bax contributed to cyanide‐induced cytochromecrelease, because SN50 and a specific Bax antisense oligodeoxynucleotide significantly reduced release of cytochromecfrom mitochondria as shown by western blot analysis. It was concluded that NF‐κB‐mediated up‐regulation of Bcl‐XSand Bax is involved in regulating cytochromecrelease in cyanide‐induced apoptosis.
PSD‐95 eliminates Src‐induced potentiation of NR1/NR2A‐subtype NMDA receptor channels and reduces high‐affinity zinc inhibition
Authors: Yamada Yasue , Iwamoto Takashi , Watanabe Yoshifumi , Sobue Kenji , Inui Makoto 
Published: March 2003

AbstractThe channel activity of NMDA receptors is regulated by phosphorylation by protein kinases and by interaction with other proteins. Recombinant NR1/NR2A subtype NMDA receptor channels are potentiated by the protein tyrosine kinase Src, an effect which is mediated by a reduction in the high‐affinity, voltage‐independent Zn2+ inhibition. However, it has been reported that Src‐induced potentiation of NMDA receptor currents in hippocampus neurons is not mediated by a reduction in Zn2+ inhibition. The post‐synaptic density protein PSD‐95 interacts with the C‐terminus of NR2 subunits of the NMDA receptor. Here we demonstrate that PSD‐95 eliminates the Src‐induced potentiation of NR1/NR2A channels expressed in oocytes and reduces the sensitivity of the channels to Zn2+. Our results reveal that the absence of Src‐induced potentiation of PSD‐95‐coupled NR1/NR2A channels is not to due to the reduced sensitivity of these channels to Zn2+. These results indicate that PSD‐95 functionally modulates NR1/NR2A channels and explain why Src‐induced potentiation of NMDA receptor currents in hippocampus neurons is not mediated by a reduction in Zn2+ inhibition.
Involvement of NF‐Y and Sp1 in basal and cAMP‐stimulated transcriptional activation of the tryptophan hydroxylase (TPH) gene in the pineal gland
Authors: Côté F. , Schussler N. , Boularand S. , Peirotes A. , Thévenot E. , Mallet J. , Vodjdani G. 
Published: March 2003

AbstractThe expression of the tryptophan hydroxylase (TPH) gene, encoding the rate‐limiting enzyme of serotonin biosynthesis, is tightly regulated both at the transcriptional and at the post‐transcriptional levels. In the pineal gland, transcription of the gene is activated in response to an intracellular circadian increase of the cAMP concentration. We have previously shown that transcription of a 2.1‐kb fragment of the human TPH promoter is induced by cAMP, although it lacks the canonical cAMP responsive element, CRE. The minimal promoter (−73/+29) has only weak transcriptional activity but is responsive to cAMP. It contains an inverted CCAAT box, which was demonstrated to be involved in this response. Here, we have extended our investigation to the functional features of the inverted CCAAT box in the −252/+29 TPH promoter, which has a higher basal activity. We show that an additional cis‐acting sequence, the adjacent GC‐rich region, cooperates with the inverted CCAAT box for the full activation of basal transcription, and that both elements are essential for the full cAMP response. We also show that in pinealocytes, NF‐Y and Sp1 transactivators bind the inverted CCAAT box and GC‐rich‐region, respectively. These factors participate in a novel pathway for the cAMP‐mediated response of the TPH promoter, which is independent of the canonical CRE‐mediated response.