bluuebunny/crossref_metadata_2025_split huggingface dataset:
Pro‐apoptotic protein kinase Cδ is associated with intranuclear inclusions in a transgenic model of Huntington's disease
Authors: Zemskov Evgeny A. , Jana Nihar R. , Kurosawa Masaru , Miyazaki Haruko , Sakamoto Naoaki , Nekooki Munenori , Nukina Nobuyuki 
Published: September 2003

AbstractIn order to investigate any effect of truncated mutant huntingtin (tNhtt) aggregation on protein kinase C (PKC) signaling in Huntington's disease (HD), we studied a possible association of PKC isoforms with the aggregates using cellular and transgenic models of HD. In this report we describe an association of mutant tNhtt with at least three PKC isoforms (α, δ, ζ), as revealed by co‐immunoprecipitation assays and immunocytochemistry in a cellular model of HD (Neuro2a cells expressing tNhtt‐150Q‐EGFP), as well as a specific association of PKCδ with intranuclear aggregates in a transgenic model (R6/2 mice). Immunoblot analysis of isolated nuclear fractions shows an elevation of nuclear PKCδ in transgenic brain tissue. The observed elevation has a strong similarity with the apoptotic translocation of PKCδ detected in experiments with the mouse neuroblastoma Neuro2a cells. Using a Neuro2a cell line expressing tNhtt with the nuclear localization signal, we demonstrate the association of PKCδ with intranuclear aggregates and present evidence that accumulation of PKCδ in cell nuclei does not depend on mutant htt nuclear translocation. Our results suggest that the association of PKCδ with intranuclear htt‐aggregates may affect its apoptotic function in a transgenic model of HD.
Glutamate‐induced declustering of post‐synaptic adaptor protein Cupidin (Homer 2/vesl‐2) in cultured cerebellar granule cells
Authors: Shiraishi Yoko , Mizutani Akihiro , Yuasa Shigeki , Mikoshiba Katsuhiko , Furuichi Teiichi 
Published: September 2003

AbstractCupidin (Homer 2/vesl‐2) is a post‐synaptic adaptor protein that associates with glutamate receptor complexes and the actin cytoskeleton. We analyzed the developmental and activity‐dependent localization of Cupidin in mouse cerebellar granule cells. Cupidin is predominantly localized to granule cell post‐synapses connecting with mossy fiber terminals in developing post‐natal cerebellum, but is diminished in adult cerebellum. In cultured granule cells 7 days in vitro, Cupidin was present as synaptic and extra‐synaptic punctate clusters that largely co‐localized with the actin‐cytoskeletal binding partners F‐actin and drebrin, as well as a post‐synaptic scaffold protein PSD‐95. Upon stimulation with glutamate, Cupidin clusters were rapidly dissociated without protein degradation, and by short‐term but not sustained stimulation they were recovered after post‐incubation without glutamate. The glutamate‐induced declustering of Cupidin preceded that of F‐actin and drebrin, was elicited by NMDA receptor‐mediated Ca2+ influx, and was followed by a downstream pathway including MAPK/ERK and protein tyrosine kinase. Specific isoforms with post‐translational modification were reduced depending on Ca2+‐dependent protein phosphatase activity. In cultured hippocampal neurons, Homer family members Homer 1, Cupidin/Homer 2 and Homer 3 showed similar glutamate‐induced declustering. We suggest that Cupidin acts as a mobile adaptor protein that changes the distribution states, clustered versus declustered, in response to synaptic activity.
Regulation of microglial inflammatory response by histone deacetylase inhibitors
Authors: Suuronen Tiina , Huuskonen Jari , Pihlaja Rea , Kyrylenko Sergiy , Salminen Antero 
Published: September 2003

AbstractThe activation of microglial cells is involved in the pathogenesis of a variety of neurodegenerative diseases, stroke and traumatic brain injuries. Recent studies suggest that protein acetylation can affect the extent of inflammatory responses. Our aim was to elucidate whether histone deacetylase inhibitors, inducers of protein hyperacetylation, regulate the inflammatory response in neural models of inflammation in vitro and whether neurone–glia interactions affect this regulation. Interestingly, we observed that histone deacetylase inhibitors, such as trichostatin A (TSA) and suberoylanilide hydroxamic acid, strongly potentiated the lipopolysaccharide (LPS)‐induced inflammatory response in murine N9 and rat primary microglial cells as well in neural co‐cultures and hippocampal slice cultures. TSA clearly potentiated the LPS‐induced expression of interleukin (IL)‐6 and inducible nitric oxide synthase mRNAs, as well as the secretion of cytokines IL‐6, tumour necrosis factor‐α and macrophage inflammatory protein (MIP)‐2, and nitric oxide (NO). Co‐culture and slice culture experiments showed that the presence of astrocytes and neurones did not stimulate or prevent the pro‐inflammatory potentiation induced by histone deacetylase inhibitor in microglial cells. The potentiation of cytokine and NO responses was blocked by the nuclear factor kappa B (NF‐κB) inhibitors caffeic acid phenethyl ester and helenalin, demonstrating that the NF‐κB signalling pathway is involved. The DNA‐binding activity of the NF‐κB complex was strongly increased by LPS treatment but not enhanced by TSA. This suggests that potentiation of the inflammatory response is not dependent on the level of cytoplasmic NF‐κB activation or DNA‐binding activity but that site of action may be at the level of transcriptional regulation. Our results suggest that environmental stresses, ageing, diet and diseases that regulate protein acetylation status may also affect the inflammatory response.
Regulatory roles of presenilin‐1 and nicastrin in neuronal differentiation during in vitro neurogenesis
Authors: Sarkar Saumyendra N. , Das Hriday K. 
Published: September 2003

AbstractPresenilin (PS) in association with nicastrin (NICA) forms a γ‐secretase complex that plays a crucial role in facilitating intramembranous processing of Notch, a signaling receptor that is essential for neuronal fate specification and differentiation. Loss of function studies have implicated a role for PS1 in regulating neuronal differentiation in association with the down‐regulation of Notch signaling during neurogenesis. By using a system for stable, as well as tetracycline‐inducible expression of interfering RNAs (RNAi), we studied the functions of PS1 during neuronal differentiation in the murine pluripotent p19 embryonic carcinoma cell line. After retinoic acid (RA) treatment and in the absence of doxycycline, neuronal progenitor cells in the p19 clone were found to extend their processes towards the neighboring colony to form network‐like connections, as revealed by neuron‐specific microtubule‐associated protein 2 staining and laser scanning confocal microscopy. However, doxycycline‐induced expression of PS1 small interfering RNA (siRNA) in the p19 clone resulted in a severe defect in the formation of network‐like connections. Expression of the NICA and Notch down‐stream effector genes Hes1 and Hes5 was unaffected in p19 cells expressing doxycycline‐induced PS1 siRNA. In contrast to PS1, constitutive inactivation of NICA by siRNA in p19 cells resulted in premature and partial differentiation without RA treatment. In these NICA siRNA‐expressing p19 cells the expression of the Notch1 down‐stream effector Hes1 gene was substantially reduced. After RA treatment the NICA siRNA clone failed to differentiate completely into networks of neurons. These results taken together provide direct evidence that PS1 and NICA may participate in neuronal differentiation during neurogenesis in vitro.
Pharmacokinetics of systemically administered tyrosine: a comparison of serum, brain tissue and in vivo microdialysate levels in the rat
Authors: Bongiovanni Rodolfo , Yamamoto Bryan K. , Simpson Catherine , Jaskiw George E. 
Published: September 2003

AbstractTyrosine uptake has been reported to differ across brain regions. However, such studies have typically been conducted over brief intervals and in anesthetized rats; anesthesia itself affects amino acid transport across the blood–brain barrier. To address these concerns, serum, brain tissue and in vivo microdialysate tyrosine levels were compared for 0–3 h after administration of tyrosine [0.138–1.10 mmol/kg intraperitoneally (i.p.)] to groups of awake rats. Serum and brain tissue tyrosine levels increased linearly with respect to dose. Basal tissue tyrosine levels varied significantly across brain regions [medial prefrontal cortex (MPFC), striatum, hypothalamus, and cerebellum], but the rate of tyrosine uptake was similar for hypothalamus, striatum and MPFC. For brain regions in which tyrosine levels in both microdialysate and tissue were assayed, namely MPFC and striatum, there was a high degree of correlation between tyrosine levels in tissue and in microdialysate. Increasing brain tyrosine levels had no effect on DA levels in MPFC microdialysate. We conclude that (i) regional differences in the response of dopamine neurons to systemic tyrosine administration cannot be attributed to pharmacokinetic factors; (ii) in vivo microdialysate provides an excellent index over time and across a wide range of tyrosine doses, of brain tissue tyrosine levels; and (iii) increases in brain tyrosine levels do not affect basal DA release in the MPFC.
Differential influence of increased polyol pathway on protein kinase C expressions between endoneurial and epineurial tissues in diabetic mice
Authors: Yamagishi Shin‐Ichiro , Uehara Kenji , Otsuki Saori , Yagihashi Soroku 
Published: September 2003

AbstractTo explore the relationship between polyol pathway and protein kinase C (PKC), we examined PKC activities and expressions of PKC isoforms separately in endoneurial and vessel‐rich epineurial tissues in diabetic mice transgenic for human aldose reductase (Tg). Tg and littermate control mice (Lm) were made diabetic by streptozotocin at 8 weeks of age and treated orally with aldose reductase inhibitor (ARI) (fidarestat 3–5 mg/kg/day) or placebo for 12 weeks. At the end, compared with non‐diabetic state, sorbitol contents were increased 6.4‐fold in endoneurium and 5.1‐fold in epineurium in diabetic Tg, whereas the increase was detected only in endoneurium in diabetic Lm. Endoneurial PKC activity was significantly reduced in diabetic Tg. By contrast, epineurial PKC activity was increased in both diabetic Lm and diabetic Tg and there was no significant difference between the two groups. These changes were all corrected by ARI treatment. Consistent with the changes of PKC activities, diabetic Tg showed decreased expression of PKCα in endoneurium, whereas there was an increased expression of PKCβII in epineurium in both diabetic Tg and diabetic Lm. These findings suggest the presence of dichotomous metabolic pathway between neural and vascular tissues in the polyol‐PKC‐related pathogenesis of diabetic neuropathy.
Substance P protects spiral ganglion neurons from apoptosis via PKC‐Ca2+‐MAPK/ERK pathways
Authors: Lallemend F. , Lefebvre P. P. , Hans G. , Rigo J. M. , Van De Water T. R. , Moonen G. , Malgrange B. 
Published: September 2003

AbstractIn the current study, we have investigated the ability of substance P (SP) to protect 3‐day‐old (P3) rat spiral ganglion neurons (SGNs) from trophic factor deprivation (TFD)‐induced cell death. The presence of SP high affinity neurokinin‐1 receptor (NK1) transcripts was detected in the spiral ganglion and the NK1 protein localized to SGNs both ex vivo and in vitro. Treatment with SP increased cytoplasmic Ca2+ in SGNs, further arguing for the presence of functional NK1 on these neurons. Both SP and the agonist [Sar9,Met(O2)11]‐SP significantly decreased SGN cell death induced by TFD, with no effect on neurite outgrowth. The survival promoting effect of SP was blocked by the NK1 antagonist, WIN51708. Both pan‐caspase inhibitor BOC‐D‐FMK and SP treatments markedly reduced activation of caspases and DNA fragmentation in trophic factor deprived‐neurons. The neuroprotective action of SP was antagonised by specific inhibitors of second messengers, including 1.2‐bis‐(O‐aminophenoxy)‐ethane‐N,N,N′,N′‐tetraacetic acid (BAPTA‐AM) to chelate cytosolic Ca2+, the protein kinase C (PKC) inhibitors bisindolylmaleimide I, Gö6976 and LY333531 and the MAPK/ERK inhibitor U0126. In contrast, nifedipine, a specific inhibitor of l‐type Ca2+ channel, and LY294002, a phosphatidylinositol‐3‐OH kinase (PI3K) inhibitor, had no effect on SP trophic support of SGNs. Moreover, activation of endogenous PKC by 4β‐phorbol 12‐myristate 13‐acetate (PMA) also reduced the loss of trophic factor‐deprived SGNs. Thus, NK1 expressed by SGNs transmit a survival‐promoting regulatory signal during TFD‐induced SGN cell death via pathways involving PKC activation, Ca2+ signalling and MAPK/ERK activation, which can be accounted for by an inhibition of caspase activation.
Opposed effects of lithium on the MEK‐ERK pathway in neural cells: inhibition in astrocytes and stimulation in neurons by GSK3 independent mechanisms
Authors: Pardo Raúl , Andreolotti Alberto G. , Ramos Belén , Picatoste Fernando , Claro Enrique 
Published: September 2003

SummaryLithium is widely used in the treatment of bipolar disorder, but despite its proven therapeutic efficacy, the molecular mechanisms of action are not fully understood. The present study was undertaken to explore lithium effects of the MEK/ERK cascade of protein kinases in astrocytes and neurons. In asynchronously proliferating rat cortical astrocytes, lithium decreased time‐ and dose‐dependently the phosphorylation of MEK and ERK, with 1 mm concentrations achieving 60 and 50% inhibition of ERK and MEK, respectively, after a 7‐day exposure. Lithium also inhibited [3H]thymidine incorporation into DNA and induced a G2/M cell cycle arrest. In serum‐deprived, quiescent astrocytes, pre‐exposure to lithium resulted in the inhibition of cell cycle re‐entry as stimulated by the mitogen endothelin‐1: under this experimental setting, lithium did not affect the rapid, peak phosphorylation of MEK taking place after 3–5 min, but was effective in inhibiting the long‐term, sustained phosphorylation of MEK. Lithium inhibition of the astrocyte MEK/ERK pathway was independent of inositol depletion. Further, compound SB216763 inhibited Tau phosphorylation at Ser396 and stabilized cytosolic β‐catenin, consistent with the inhibition of glycogen synthase kinase‐3β (GSK‐3β), but failed to reproduce lithium effects on MEK and ERK phosphorylation and cell cycle arrest. In cerebellar granule neurons, millimolar concentrations of lithium enhanced MEK and ERK phosphorylation in a concentration‐dependent manner, again through an inositol and GSK‐3β independent mechanism. These opposing effects in astrocytes and neurons make lithium treatment a promising strategy to favour neural repair and reduce reactive gliosis after traumatic injury.
Activation of stimulatory heterotrimeric G proteins increases glutathione and protects neuronal cells against oxidative stress
Authors: Lewerenz Jan , Letz Julia , Methner Axel 
Published: September 2003

AbstractOxidative glutamate toxicity in the neuronal cell line HT22 is a model for cell death by oxidative stress, where an excess of extracellular glutamate inhibits import of cystine, a building block of the antioxidant glutathione. The subsequent decrease in glutathione then leads to the accumulation of reactive oxygen species (ROS) and programmed cell death. We used pharmacological compounds known to interact with heterotrimeric G‐protein signalling and studied their effects on cell survival, morphology, and intracellular events that ultimately lead to cell death. Cholera toxin and phorbol esters were most effective and prevented cell death through independent pathways. Treating HT22 cells with cholera toxin attenuated the glutamate‐induced accumulation of ROS and calcium influx. This was, at least in part, caused by an increase in glutathione due to improved uptake of cystine mediated by the induction of the glutamate/cystine‐antiporter subunit xCT or, additionally, by the up‐regulation of the antiapoptotic protein Bcl‐2. Gs activation also protected HT22 cells from hydrogen peroxide or inhibition of glutathione synthesis by buthionine sulfoximine, and immature cortical neurones from oxidative glutamate toxicity. Thus, this pathway might be more generally implicated in protection from neuronal death by oxidative stress.
Alteration in calcium channel properties is responsible for the neurotoxic action of a familial frontotemporal dementia tau mutation
Authors: Furukawa Katsutoshi , Wang Yue , Yao Pamela J. , Fu Weiming , Mattson Mark P. , Itoyama Yasuto , Onodera Hiroshi , D'Souza Ian , Poorkaj Parvone H. , Bird Thomas D. , Schellenberg Gerard D. 
Published: September 2003

AbstractTau, a microtubule binding protein, is not only a major component of neurofibrillary tangles in Alzheimer's disease, but also a causative gene for hereditary frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP‐17). We show here that an FTDP‐17 tau mutation (V337M) in SH‐SY5Y cells reduces microtubule polymerization, increases voltage‐dependent calcium current (ICa) density, and decreases ICa rundown. The reduced rundown of ICa by V337M was significantly inhibited by nifedipine (L‐type Ca channel blocker), whereas ω‐conotoxin GVIA (N‐type Ca channel blocker) showed smaller effects, indicating that tau mutations affect L‐type calcium channel activity. The depolarization‐induced increase in intracellular calcium was also significantly augmented by the V337M tau mutation. Treatment with a microtubule polymerizing agent (taxol), an adenylyl cyclase inhibitor, or a protein kinase A (PKA) inhibitor, counteracted the effects of mutant tau on ICa. Taxol also attenuated the Ca2+ response to depolarization in cells expressing mutant tau. Apoptosis in SH‐SY5Y cells induced by serum deprivation was exacerbated by the V337M mutation, and nifedipine, taxol, and a PKA inhibitor significantly protected cells against apoptosis. Our results indicate that a tau mutation which decreases its microtubule‐binding ability augments calcium influx by depolymerizing microtubules and activating adenylyl cyclase and PKA.
Flow cytometric analysis of mitochondria from CA1 and CA3 regions of rat hippocampus reveals differences in permeability transition pore activation
Authors: Mattiasson Gustav , Friberg Hans , Hansson Magnus , Elmér Eskil , Wieloch Tadeusz 
Published: September 2003

AbstractMitochondria are important in the pathophysiology of several neurodegenerative diseases, and mitochondrial production of reactive oxygen species (ROS), membrane depolarization, permeability changes and release of apoptogenic proteins are involved in these processes. Following brain insults, cell death often occurs in discrete regions of the brain, such as the subregions of the hippocampus. To analyse mitochondrial structure and function in such subregions, only small amounts of mitochondria are available. We developed a protocol for flow cytometric analysis of very small samples of isolated brain mitochondria, and analysed mitochondrial swelling and formation of ROS in mitochondria from the CA1 and CA3 regions of the hippocampus. Calcium‐induced mitochondrial swelling was measured, and fluorescent probes were used to selectively stain mitochondria (nonyl acridine orange), to measure membrane potential (tetramethylrhodamine‐methyl‐ester, 1,1′,3,3,3′,3′‐hexamethylindodicarbocyanine‐iodide) and to measure production of ROS (2′,7′‐dichlorodihydrofluorescein‐diacetate). We found that formation of ROS and mitochondrial permeability transition pore activation were higher in mitochondria from the CA1 than from the CA3 region, and propose that differences in mitochondrial properties partly underlie the selective vulnerability of the CA1 region to brain insults. We also conclude that flow cytometry is a useful tool to analyse the role of mitochondria in cell death processes.
Modulation of mitochondrial membrane potential and reactive oxygen species production by copper in astrocytes
Authors: Gyulkhandanyan Armen V. , Feeney Chris J. , Pennefather Peter S. 
Published: September 2003

AbstractIn monolayers of cultured rat astrocytes a number of agents that induce oxidative stress act synergistically with exposure to copper leading to rapid depolarization of the mitochondrial membrane potential (Ψm) and increased reactive oxygen species (ROS) production. Copper sensitized astrocytes to the action of menadione, an intracellular generator of superoxide anion radical, exogenous hydrogen peroxide (H2O2) and rotenone, an inhibitor of mitochondrial electron transport chain complex I. However, significant differences were observed in the ability to modulate the copper‐enhanced oxidative stress depending on which stressor was used. The inhibitor of mitochondrial permeability transition cyclosporin A attenuated the effect of copper and rotenone, but had no protective action in the case of H2O2/copper and menadione/copper combinations. The H2O2 scavenger pyruvate was effective at protecting mitochondria against damage associated with the combined exposure to H2O2/copper and menadione/copper but not to the rotenone/copper combination. The antioxidant Trolox was ineffective at protecting against any of these actions and indeed had a damaging effect when combined with copper. The membrane‐permeable copper chelator neocuproine combined with sensitizing concentrations of menadione caused a decrease in Ψm, mimicking the action of copper. Penicillamine, a membrane‐impermeable copper chelator, was effective at reducing copper sensitization. Endogenous copper, mobilized during periods of oxidative stress, may play a role in the pathophysiology of brain injury. Our results suggest that this might be particularly dangerous in dysfunctional conditions in which the mitochondrial electron transport chain is compromised.
Prion protein expression modulates neuronal copper content
Authors: Brown David R. 
Published: September 2003

AbstractThe prion protein is a copper (Cu)‐binding protein. The abnormal isoform of this protein is associated with the transmissible spongiform encephalopathies or prion diseases. In prion diseases, the prion protein loses its Cu binding capacity. The effect of prion protein expression on the Cu content of the brain was investigated. Transgenic mice, either overexpressing the prion protein or expressing a mutant form lacking the Cu‐binding region of the protein, were compared with wild‐type mice and mice in which expression of the protein was knocked out. Age‐dependent differences in Cu content of the brain were detected. Also, synaptosomal fractions from the brains of the mice showed different Cu content depending on the expression of the prion protein. Mice expressing prion protein, but without the Cu‐binding domain showed reduced Cu content. Mice overexpressing the prion protein showed little difference in Cu in the brain compared with wild type but also the prion protein expressed by the mice showed a reduction in the level of Cu bound. These results confirm that prion protein expression modulates the Cu level found at the synapse and this effect is dependent on its Cu binding capacity. Loss of normal Cu binding by the prion protein altered age‐related increases in metals in the brain. This may explain why many forms of human prion disease do not develop until late in life.