bluuebunny/crossref_metadata_2025_split huggingface dataset:
Temperature‐dependent, neurotrophic factor‐elicited, neuronal differentiation in adrenal chromaffin cell line immortalized with temperature‐sensitive SV40 T‐antigen
Authors: Murata Tomiyasu , Hikita Kiyomi , Tsuboi Masaru , Niwa Kentaro , Suzuki Misao , Kaneda Norio 
Published: May 2003

AbstractWe established adrenal medullary cell lines from transgenic mice expressing an oncogene, the temperature‐sensitive simian virus 40 large T‐antigen, under the control of the tyrosine hydroxylase promoter. A clonal cell line, named tsAM5D, conditionally grew at a permissive temperature of 33°C and exhibited the dopaminergic chromaffin cell phenotype as exemplified by the expression pattern of mRNA for catecholamine‐synthesizing enzymes and secretory vesicle‐associated proteins. tsAM5D cells proliferated at the permissive temperature in response to basic fibroblast growth factor (bFGF) and ciliary neurotrophic factor (CNTF). At a non‐permissive temperature of 39°C, bFGF and CNTF acted synergistically to differentiate tsAM5D cells into neuron‐like cells. In addition, tsAM5D cells caused to differentiate by bFGF plus CNTF at 39°C became dependent solely on nerve growth factor for their survival and showed markedly enhanced neurite outgrowth. In the presence of bFGF and CNTF, the morphological change induced by the temperature shift was associated with up‐regulated expression of neuronal marker genes including neuron‐specific enolase, growth‐associated protein‐43, microtubule‐associated protein 2, neurofilament, and p75 neurotrophin receptor, indicating that the cells underwent neuronal differentiation. Thus, we demonstrated that tsAM5D cells could proliferate at permissive 33°C, and also had the capacity to terminally differentiate into neuron‐like cells in response to bFGF and CNTF when the oncogene was inactivated by shifting the temperature to non‐permissive 39°C. These results suggest that tsAM5D cells should be a good tool to allow a detailed study of mechanisms regulating neuronal differentiation.
Gliap − a novel untypical l‐asparaginase localized to rat brain astrocytes
Authors: Dieterich Daniela C. , Landwehr Marco , Reissner Carsten , Smalla Karl‐Heinz , Richter Karin , Wolf Gerald , Böckers Tobias M. , Gundelfinger Eckart D. , Kreutz Michael R. 
Published: May 2003

Abstract l‐asparaginases catalyse the formation of the neuroactive amino acid l‐aspartate by deamination of asparagine. The major pathophysiological significance of l‐asparaginase activity is in its clinical use for the treatment of acute lymphatic leukaemia and neoplasias that require asparagine and obtain it from circulating pools. Here we report the identification and characterization of Gliap, a cytosolic l‐asparaginase, which is the founding member of a new group of l‐asparaginases in mammalia. Structural modelling suggests that Gliap is an atypical mammalian type‐I asparaginase inasmuch as it harbours the active centre of a type‐I glycosylasparaginase but, like plant‐type asparaginases, lacks their auto‐proteolytic site and, in addition, exhibits significant type‐II l‐asparaginase enzymatic activity. Moreover, in contrast to glycosylasparaginases Gliap is enriched in the cytosolic fraction and not in lysosomes. The protein is particularly abundant in liver, testis and brain. In brain Gliap is exclusively expressed in astrocytes and prominently present in structures reminiscent of glial endfeet. These data suggest that Gliap is involved in astroglial production of the neuroactive amino acid l‐aspartate.
An oestrogen membrane receptor participates in estradiol actions for the prevention of amyloid‐β peptide1−40‐induced toxicity in septal‐derived cholinergic SN56 cells
Authors: Marin Raquel , Guerra Borja , Morales Araceli , Díaz Mario , Alonso Rafael 
Published: May 2003

AbstractAlthough oestrogen [17β‐estradiol (E2)]‐related neuroprotection has been demonstrated in different models, the involvement of non‐classical oestrogen receptors (ERs) remains unexplored. Using the SN56 cholinergic cell line, we present evidence indicating that an ER associated with the plasma membrane participates in oestrogen‐dependent inhibition of cell death induced by amyloid‐β peptide (Aβ) toxicity. Similarly to E2 alone, a 15‐min exposure to estradiol‐horseradish peroxidase (E‐HRP) significantly reduced Aβ‐induced cell death. This effect was decreased by the ER antagonist ICI 182,780 as well as by MC‐20 antibody directed to a region neighbouring the ligand‐binding domain of ERα. Using confocal microscopy on unpermeabilized SN56 cells exposed to MC‐20 antibody, we identified a protein at the plasma membrane level. Western blot analysis of purified SN56 cell membrane fractions using MC‐20 antibody revealed the presence of one band with the same electrophoretic mobility as intracellular ERα. Using conjugated forms of the steroid, E‐HRP and E2 conjugated to bovine serum albumin‐FITC, we demonstrated by confocal microscopy that SN56 cells contain surface binding sites for E2. Binding of both conjugates was blocked by pre‐incubation with E2 and decreased by either ICI 182,780 or MC‐20 antibody in a concentration‐dependent manner. Thus, a membrane‐related ER that shares some structural homologies with ERα may participate in oestrogen‐mediated neuroprotection.
Concerted stimulation and deactivation of pertussis toxin‐sensitive G proteins by chimeric G protein‐coupled receptor‐regulator of G protein signaling 4 fusion proteins: analysis of the contribution of palmitoylated cysteine residues to the GAP activity of RGS4
Authors: Bahia Daljit S. , Sartania Nana , Ward Richard J. , Cavalli Antonella , Jones Teresa L. Z. , Druey Kirk M. , Milligan Graeme 
Published: May 2003

AbstractAgonists stimulated high‐affinity GTPase activity in membranes of HEK293 cells following coexpression of the α2A‐adrenoceptor and a pertussis toxin‐resistant mutant of Go1α. Enzyme kinetic analysis of Vmax and Km failed to detect regulation of the effect of agonist by a GTPase activating protein. This did occur, however, when cells were also transfected to express RGS4. Both elements of a fusion protein in which the N‐terminus of RGS4 was linked to the C‐terminal tail of the α2A‐adrenoceptor were functional, as it was able to provide concerted stimulation and deactivation of the G protein. By contrast, the α2A‐adrenoceptor‐RGS4 fusion protein stimulated but did not enhance deactivation of a form of Go1α that is resistant to the effects of regulator of G protein signaling (RGS) proteins. Employing this model system, mutation of Asn128 but not Asn88 eliminated detectable GTPase activating protein activity of RGS4 against Go1α. Mutation of all three cysteine residues that are sites of post‐translational acylation in RGS4 also eliminated GTPase activating protein activity but this was not achieved by less concerted mutation of these sites. These studies demonstrate that a fusion protein between a G protein‐coupled receptor and an RGS protein is fully functional in providing both enhanced guanine nucleotide exchange and GTP hydrolysis of a coexpressed G protein. They also provide a direct means to assess, in mammalian cells, the effects of mutation of the RGS protein on function in circumstances in which the spatial relationship and orientation of the RGS to its target G protein is defined and maintained.
Molecular correlates of impaired prefrontal plasticity in response to chronic stress
Authors: Kuipers S. D. , Trentani A. , Den Boer J. A. , Ter Horst G. J. 
Published: May 2003

AbstractDisturbed adaptations at the molecular and cellular levels following stress could represent compromised neural plasticity that contributes to the pathophysiology of stress‐induced disorders. Evidence illustrates atrophy and cell death of stress‐vulnerable neurones in the prefrontal cortex. Reduced plasticity may be realized through the destabilized function of selective proteins involved in organizing the neuronal skeleton and translating neurotrophic signals. To elucidate the mechanisms underlying these effects, rats were exposed to chronic footshock stress. Patterns of c‐fos, phospho‐extracellular‐regulated protein kinases 1/2 (ERK1/2), calcineurin and phospho‐cyclic‐AMP response‐element binding protein (CREB) expression were subsequently investigated. The results indicate chronic stress‐induced impairments in prefrontal and cingulate signal transduction cascades underlying neuronal plasticity. The medial prefrontal cortex, demonstrated functional hyperactivity and dendritic phospho‐ERK1/2 hyperphosphorylation, while reduced c‐fos and calcineurin immunoreactivity occurred in the cingulate cortex. Significantly reduced phospho‐CREB expression in both cortical regions, considering its implication in brain‐derived neurotrophic factor (BDNF) transcription, suggests reduced synaptic plasticity. This data confirms the damaging effect of stress on cortical activity, on a molecular level. Due to the association of these markers in the regulation of BDNF signalling, these findings suggest a central role for intracellular neurotrophin transduction members in the pathways underlying cellular actions of stress in the brain.
The same cellular signaling pathways mediate survival in sensory neurons that switch their trophic requirements during development
Authors: Salvarezza Susana B. , López Hector S. , Mascó Daniel H. 
Published: May 2003

AbstractA distinct subpopulation of rat dorsal root sensory (DRG) neurons, termed P‐neurons, switch their trophic requirements for survival during development from nerve growth factor (NGF) at embryonic stages to basic fibroblast growth factor (bFGF) just after birth. We investigated in cultured P‐neurons the intracellular signaling pathways mediating survival before and after this switch. The NGF‐induced survival was completely blocked by either wortmannin (100 nm) or PD98059 (25–50 nm), which selectively inhibit the phosphatidylinositol 3‐kinase‐AKT (PI3 kinase‐AKT) and mitogen‐activated kinase kinase extracellular regulated kinase (MEK‐ERKs) pathways, respectively. NGF activated AKT and ERKs in single embryonic P‐neurons, as assayed by immunofluorescence of phorphorylated proteins. In concordance with the survival assays, wortmannin and PD98059 blocked AKT and ERKs activation, respectively. Following the trophic switch, bFGF used the same signaling pathways to promote survival of post‐natal P‐neurons, as either wortmannin or PD98059 blocked its effect. Also, bFGF activated AKT and ERKs in single P‐neurons, and this activation was blocked by the same inhibitors. These results strongly suggest that both pathways concurrently mediate the action of NGF and bFGF during embryonic and post‐natal periods, respectively. Thus, we report the novel result that the switch in trophic requirements occurs with conservation of the signaling pathways mediating survival.
Chronic nicotine administration does not increase nicotinic receptors labeled by [125I]epibatidine in adrenal gland, superior cervical ganglia, pineal or retina
Authors: Dávila‐García Martha I. , Musachio John L. , Kellar Kenneth J. 
Published: May 2003

AbstractNeuronal nicotinic acetylcholine receptors (nAChRs) were measured in CNS and peripheral tissues following continuous exposure to saline or nicotine hydrogen tartrate (3.3 or 10 mg/kg/day) for 14 days via osmotic pumps. Initially, binding of [3H](–)nicotine, [3H]cytisine and [3H]epibatidine to nAChRs was compared to determine the suitability of each for these kinds of studies. The predominant nAChR labeled by agonists in the cerebral cortex is an α4β2 subtype, whereas the predominant nicotinic receptors in the adrenal gland, superior cervical ganglia and pineal gland contain an α3 subunit, and they do not bind either [3H](–)nicotine or [3H]cytisine with high affinity. In retina some nAChRs bind all three ligands with high affinity, and others appear to bind only [3H]epibatidine. Thus, only [3H]epibatidine had high enough affinity to be useful for measuring the nAChRs in all of the tissues. The receptors from nicotine‐treated rats were then measured using [125I]epibatidine, which has binding characteristics very similar to [3H]epibatidine. Treatment with the two doses of nicotine hydrogen tartrate increased binding sites in the cerebral cortex by 40% and 70%, respectively. In contrast, no significant changes in the density of receptor binding sites were found in the adrenal gland, superior cervical ganglia, pineal gland or retina. These data indicate that chronic administration of nicotine even at high doses does not increase all nicotinic receptor subtypes, and that receptors containing α3 subunits may be particularly resistant to this nicotine‐induced change.
MCP‐1 (CCL2) protects human neurons and astrocytes from NMDA or HIV‐tat‐induced apoptosis
Authors: Eugenin E. A. , D'Aversa T. G. , Lopez L. , Calderon T. M. , Berman J. W. 
Published: May 2003

AbstractAcquired immunodeficiency syndrome (AIDS)‐associated dementia is often characterized by chronic inflammation, with infected macrophage infiltration of the CNS resulting in the production of human immunodeficiency virus type 1 (HIV‐1) products, including tat, and neurotoxins that contribute to neuronal loss. In addition to their established role in leukocyte recruitment and activation, we identified an additional role for chemokines in the CNS. Monocyte chemoattractant protein‐1 (MCP‐1 or CCL2) and regulated upon activation normal T cell expressed and secreted (RANTES) were found to protect mixed cultures of human neurons and astrocytes from tat or NMDA‐induced apoptosis. Neuronal and astrocytic apoptosis in these cultures was significantly inhibited by co‐treatment with MCP‐1 or RANTES but not IP‐10. The protective effect of RANTES was blocked by antibodies to MCP‐1, indicating that RANTES protection is mediated by the induction of MCP‐1. The NMDA blocker, MK801, also abolished the toxic effects of both tat and NMDA. Tat or NMDA treatment of mixed cultures for 24 h resulted in increased extracellular glutamate ([Glu]e) and NMDA receptor 1 (NMDAR1) expression, potential contributors to apoptosis. Co‐treatment with MCP‐1 inhibited tat and NMDA‐induced increases in [Glu]e and NMDAR1, and also reduced the levels and number of neurons containing intracellular tat. These data indicate that MCP‐1 may play a novel role as a protective agent against the toxic effects of glutamate and tat.
Heme oxygenase‐1 and heme oxygenase‐2 have distinct roles in the proliferation and survival of olfactory receptor neurons mediated by cGMP and bilirubin, respectively
Authors: Chen Jijun , Tu Yajun , Moon Cheil , Nagata Eiichiro , Ronnett Gabriele V. 
Published: May 2003

AbstractHeme oxygenase (HO) is implicated in protection against oxidative stress, proliferation and apoptosis in many cell types, including neurons. We utilized olfactory receptor neurons (ORNs) as a model to define the roles of HO‐1 and HO‐2 in neuronal development and survival, and to determine the mediators of these effects. The olfactory system is a useful model as ORNs display neurogenesis post‐natally and do not contain nitric oxide synthase (NOS) activity, which could confound results. HO isoforms were expressed in ORNs during embryogenesis and post‐natally. Mice null for either HO‐1 or HO‐2 displayed decreased proliferation of neuronal precursors. However, apoptosis was increased only in HO‐2 null mice. Cyclic GMP immunostaining was reduced in ORNs in both genotypes, providing direct evidence that HO mediates cGMP production in vivo. Bilirubin immunostaining was reduced only in HO‐2 null mice. These roles for HO‐1 and HO‐2 were confirmed using detergent ablation of the epithelium to observe increased neurogenesis of ORNs after target disruption in HO null mice. Primary cultures of ORNs revealed that proliferative and survival effects of HO were mediated through cGMP and bilirubin, respectively. These results support a role for HO, the CO‐cGMP signaling system and bilirubin in neurodevelopment and in response to injury.
In vivo neuroprotective role of NMDA receptors against kainate‐induced excitotoxicity in murine hippocampal pyramidal neurons
Authors: Ogita Kiyokazu , Okuda Hiroaki , Yamamoto Yasuhiro , Nishiyama Norito , Yoneda Yukio 
Published: May 2003

AbstractActivation of NMDA receptors has been shown to induce either neuronal cell death or neuroprotection against excitotoxicity in cultured cerebellar granule neurons in vitro. We have investigated the effects of pretreatment with NMDA on kainate‐induced neuronal cell death in mouse hippocampus in vivo. The systemic administration of kainate (30 mg/kg), but not NMDA (100 mg/kg), induced severe damage in pyramidal neurons of the hippocampal CA1 and CA3 subfields 3–7 days later, without affecting granule neurons in the dentate gyrus. An immunohistochemical study using an anti‐single‐stranded DNA antibody and TdT‐mediated dUTP nick end labeling analysis both revealed that kainate, but not NMDA, induced DNA fragmentation in the CA1 and CA3 pyramidal neurons 1–3 days after administration. Kainate‐induced neuronal loss was completely prevented by the systemic administration of NMDA (100 mg/kg) 1 h to 1 day previously. No pyramidal neuron was seen with fragmented DNA in the hippocampus of animals injected with kainate 1 day after NMDA treatment. The neuroprotection mediated by NMDA was prevented by the non‐competitive NMDA receptor antagonist MK‐801. Taken together these results indicate that in vivo activation of NMDA receptors is capable of protecting against kainate‐induced neuronal damage through blockade of DNA fragmentation in murine hippocampus.
Two modes of exocytosis from synaptosomes are differentially regulated by protein phosphatase types 2A and 2B
Authors: Baldwin Monique L. , Rostas John A. P. , Sim Alistair T. R. 
Published: May 2003

AbstractThe inhibitors okadaic acid (OA), fostriecin (FOS) and cyclosporin A (CsA), were used to investigate the roles of protein phosphatases in regulating exocytosis in rat brain synaptosomes by measuring glutamate release and the release of the styryl dye FM 2‐10. Depolarization was induced by 30 mm KCl, or 0.3 mm or 1 mm 4‐aminopyridine (4AP). OA and FOS produced a similar partial inhibition of KCl‐ and 0.3 mm 4AP‐ evoked exocytosis in both assays, but had little effect upon exocytosis evoked by 1 mm 4AP. In contrast, CsA had no effect upon KCl‐ and 0.3 mm 4AP‐evoked exocytosis, but significantly enhanced glutamate release but not FM 2‐10 dye release evoked by 1 mm 4AP. None of the phosphatase inhibitors changed calcium signals from FURA‐2‐loaded synaptosomes either before or after depolarization. Pretreatment with 100 nm phorbol 12‐myristate 13‐acetate abolished the inhibitory effect of OA on exocytosis induced by 0.3 mm 4AP. Taken together, these results show that exocytosis from synaptosomes has a phosphatase‐sensitive and phosphatase‐insensitive component, and that there are two modes of phosphatase‐sensitive exocytosis that can be elicited by different depolarization conditions. Moreover, these two modes are differentially sensitive to phosphatase 2A and 2B.
Pentylenetetrazole decreases metabolic glutamate turnover in rat brain
Authors: Eloqayli Haytham , Dahl Carl Bredo , Götestam K. Gunnar , Unsgård Geirmund , Hadidi Hakam , Sonnewald Ursula 
Published: May 2003

AbstractSeizures were induced in rats by intraperitoneal injection of pentylenetetrazole (PTZ, 70 mg/kg), followed, 30 min later, by injection of [1‐13C]glucose and [1,2‐13C]acetate. Analyses of extracts from cortex, subcortex and cerebellum were performed using 13C magnetic resonance spectroscopy and HPLC. It could be shown that PTZ affected different brain regions differently. The total amounts of glutamate, glutamine, GABA, aspartate and taurine were decreased in the cerebellum and unchanged in the other brain regions. GABAergic neurones in the cortex and subcortex were not affected, whereas those in the cerebellum showed a pronounced decrease of GABA synthesis. However, glutamatergic neurones in all brain regions showed a decrease in glutamate labelling and in addition a decreased turnover in cerebellum. It could be shown that this decrease was in the metabolic pool of glutamate whereas release of glutamate was unaffected since glutamine labelling from glutamate was unchanged. Aspartate turnover was also decreased in all brain regions. Changes in astrocyte metabolism were not detected, indicating that PTZ had no effect on astrocyte metabolism in the early postictal stage.
Proteasome activation and nNOS down‐regulation in neuroblastoma cells expressing a Cu,Zn superoxide dismutase mutant involved in familial ALS
Authors: Aquilano Katia , Rotilio Giuseppe , Ciriolo Maria Rosa 
Published: May 2003

AbstractReactive oxygen and nitrogen species have emerged as predominant effectors of neurodegeneration. We demonstrated that expression of the fully active G93A Cu,Zn superoxide dismutase mutant in neuroblastoma cells is associated with an increased level of oxidatively modified proteins, in terms of carbonylated residues. A parallel increase in proteasome activity was detected and this was mandatory in order to assure cell viability. In fact, proteasome inhibition by lactacystin or MG132 resulted in programmed cell death. Nitrosative stress was not involved in the oxidative unbalance, as a decrease in neuronal nitric oxide production and down‐regulation of neuronal nitric oxide synthase (nNOS) level were detected. The nNOS down‐regulation was correlated to increased proteolytic degradation by proteasome, because comparable levels of nNOS were detected in G93A and parental cells upon treatment with lactacystin. The altered rate of proteolysis observed in G93A cells was specific for nNOS as Cu,Zn superoxide dismutase (Cu,Zn SOD) degradation by proteasome was influenced neither by its mutation nor by increased proteasome activity. Treatment with the antioxidant 5,5′‐dimethyl‐1‐pyrroline N‐oxide resulted in inhibition of protein oxidation and decrease in proteasome activity to the basal levels. Overall these results confirm the pro‐oxidant activity of G93A Cu,Zn SOD mutant and, at the same time, suggest a cross‐talk between reactive oxygen and nitrogen species via the proteasome pathway.
Chronic hypoxia potentiates capacitative Ca2+entry in type‐I cortical astrocytes
Authors: Smith I. F. , Plant L. D. , Boyle J. P. , Skinner R. A. , Pearson H. A. , Peers C. 
Published: May 2003

AbstractProlonged hypoxia exerts profound effects on cell function, and has been associated with increased production of amyloid β peptides (AβPs) of Alzheimer's disease. Here, we have investigated the effects of chronic hypoxia (2.5% O2, 24 h) on capacitative Ca2+entry (CCE) in primary cultures of rat type‐I cortical astrocytes, and compared results with those obtained in astrocytes exposed to AβPs. Chronic hypoxia caused a marked enhancement of CCE that was observed after intracellular Ca2+stores were depleted by bradykinin application or by exposure to thapsigargin (1 µm). Exposure of cells for 24 h to 1 µmAβP(1−40)did not alter CCE. Enhancement of CCE was not attributable to cell hyperpolarization, as chronically hypoxic cells were significantly depolarized as compared with controls. Mitochondrial inhibition [by FCCP (10 µm) and oligomycin (2.5 µg/mL)] suppressed CCE in all three cell groups, but more importantly there were no significant differences in the magnitude of CCE in the three astrocyte groups under these conditions. Similarly, the antioxidants melatonin and Trolox abolished the enhancement of CCE in hypoxic cells. Our results indicate that chronic hypoxia augments CCE in cortical type‐I astrocytes, a finding which is not mimicked by AβP(1−40)and appears to be dependent on altered mitochondrial function.