bluuebunny/crossref_metadata_2025 huggingface dataset:
4‐Aminopyridine‐induced epileptogenesis depends on activation of mitogen‐activated protein kinase ERK
Authors: Merlo Daniela , Cifelli Pierangelo , Cicconi Simona , Tancredi Virginia , Avoli Massimo
Published: December 2005

Extracellular signal‐regulated kinases such as ERK1 [p44 mitogen‐activated protein kinase (MAPK)] and ERK2 (p42 MAPK) are activated in the CNS under physiological and pathological conditions such as ischemia and epilepsy. Here, we studied the activation state of ERK1/2 in rat hippocampal slices during application of the K+channel blocker 4‐aminopyridine (4AP, 50 µm), a procedure that enhances synaptic transmission and leads to the appearance of epileptiform activity. Hippocampal slices superfused with 4AP‐containing medium exhibited a marked activation of ERK1/2 phosphorylation that peaked within about 20 min. These effects were not accompanied by changes in the activation state of c‐Jun N‐terminal kinase (JNK), another member of the MAP kinase superfamily. 4AP‐induced ERK1/2 activation was inhibited by the voltage‐gated Na+channel blocker tetrodotoxin (1 µm). We also found that application of the ERK pathway inhibitors U0126 (50 µm) or PD98059 (100 µm) markedly reduced 4AP‐induced epileptiform synchronization, thus abolishing ictal discharges in the CA3 area. The effects induced by U0126 or PD98059 were not associated with changes in the amplitude and latency of the field potentials recorded in the CA3 area following electrical stimuli delivered in the dentate hylus. These data demonstrate that activation of ERK1/2 accompanies the appearance of epileptiform activity induced by 4AP and suggest a cause‐effect relationship between the ERK pathway and epileptiform synchronization.
Comparison of in vivo and in vitro subcellular localization of estrogen receptors α and β in oligodendrocytes
Authors: Zhang Zhan , Cerghet Mirela , Mullins Chadwick , Williamson Maryann , Bessert Denise , Skoff Robert
Published: December 2005

The existence of estrogen receptors (ERs) in oligodendrocytes (OLGs) in vivo and in vitro is unresolved, as their presence has been reported in some studies and their absence in others. Using molecular and immunocytochemical techniques, we describe the subcellular localization of ERα and ERβ in OLGs in vivo and in vitro. Both ERα and ERβ are detected in an immortalized OLG cell line and in enriched OLG cultures by RT‐PCR and western blot. Immunocytochemistry of OLGs from enriched cultures shows ERα receptors are nuclear, whereas ERβ receptors are cytoplasmic. Confocal and deconvolution microscopy of enriched OLG cultures reveals ERβ immunoreactivity is concentrated in perikarya and veins of OLG membrane sheets; lesser reactivity is present in their plasma membranes and nuclei. In vivo, we readily detect ERα in neurons but not in OLGs, even though we used different fixation procedures and different ERα antibodies. The presence of ERα in cultured OLGs may be due to culture media that contains factors stimulating ERα expression but are reduced in normal brain. In vivo, ERβ immunoreactivity is readily detectable in OLG cytoplasm and in myelin sheaths. Incubation of glial cultures without or with increasing concentrations of 17β–estradiol (E2) shows that E2 significantly accelerates OLG process formation.
Dopamine transporter‐mediated cytotoxicity of β‐carbolinium derivatives related to Parkinson's disease: relationship to transporter‐dependent uptake
Authors: Storch Alexander , Hwang Yu‐I , Gearhart Debra A. , Beach J. Warren , Neafsey Edward J. , Collins Michael A. , Schwarz Johannes
Published: December 2005

Endogenous or exogenous β‐carboline (βC) derivatives structurally related to the selective dopaminergic neurotoxin 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP) and its active metabolite 1‐methyl‐4‐phenylpyridinium (MPP+) may contribute to dopaminergic neurodegeneration in Parkinson's disease (PD). We addressed the importance of the dopamine transporter (DAT) for selective dopaminergic toxicity by testing the differential cytotoxicity and cellular uptake of 12 βCs in human embryonic kidney HEK‐293 cells ectopically expressing the DAT gene. Cell death was measured using [4,5‐Dimethylthiazol‐2‐yl]‐2,5‐diphenyltetrazolium bromide (MTT) and trypan blue exclusion assays, and uptake by a fluorescence‐based uptake assay. All βCs and MPP+ showed general cytotoxicity in parental HEK‐293 cells after 72 h with half‐maximal toxic concentrations (TC50 values) in the upper micromolar range. Besides MPP+, only 2[N]‐methylated compounds showed enhanced cytotoxicity in DAT expressing HEK‐293 cells with 1.3‐ to 4.5‐fold reduction of TC50 values compared with parental cell line. The rank order of selectivity was: MPP+ >> 2[N],9[N]‐dimethyl‐harminium > 2[N]‐methyl‐harminium > 2[N],9[N]‐dimethyl‐harmanium = 2[N]‐methyl‐norharmanium > 2[N]‐methyl‐harmanium > 2[N],9[N]‐dimethyl‐norharminium. Consistently, only 2[N]‐methylated βCs were transported into the cell through the DAT with up to five times greater Km and 12–220 times smaller Vmax values compared with dopamine and MPP+. There was a weak relation of DAT‐mediated selectivity with the affinity of βCs at the DAT (Km), but not with Vmax. Our data suggest that DAT‐mediated cellular uptake of 2[N]‐methylated βCs represents a potential mechanism for selective toxicity towards dopaminergic neurons and may be relevant for the pathogenesis of Parkinson's disease.
Interaction between dopamine and its transporter: role of intracellular sodium ions and membrane potential
Authors: Chen Nianhang , Reith Maarten E. A.
Published: December 2005

The present study addresses the effect of intracellular Na+and membrane potential on the binding of dopamine (DA) to the dopamine transporter (DAT). Perforation of plasma membranes of DAT‐expressing cells with gramicidin diminished DA uptake and decreased the potency (increases Ki) of DA in inhibiting the binding of cocaine analog [3H]2β‐carbomethoxy‐3β‐(4‐fluorophenyl)tropane (CFT). It also compromised the ability of external Na+to reduce DA Ki. No substantial effect on DA Kiwas observed upon gramicidin treatment in Na+‐free buffer, membrane depolarization with high [K+]o, or elevation of [Na+]iwith monensin under non‐depolarizing conditions. Elevation of DA Kiwas greater at more positive potentials when [Na+]iwas raised to a similar level, or at higher [Na+]iwhen the membrane was depolarized to a similar level. In cells expressing D313N DAT, DA Kiwas significantly higher but less sensitive to gramicidin than that in wild‐type (WT) cells. In contrast, DA Kiin cell‐free membranes was insensitive to Na+, gramicidin, and D313N mutation. The data suggest that (i) intracellular Na+plays a role in affecting the external access to DA binding sites at DAT on depolarized plasma membranes of cells, and (ii) access to DA binding sites in cell‐free membranes may occur from the intracellular side of the membrane. Unlike DA binding, CFT binding to both cells and membranes was sensitive to Na+and D313N mutation but insensitive to gramicidin, consistent with exclusively external access to sites that are different from but conformationally linked to those for DA.
Effects of 6‐hydroxydopamine on primary cultures of substantia nigra: specific damage to dopamine neurons and the impact of glial cell line‐derived neurotrophic factor
Authors: Ding Yun Min , Jaumotte Juliann D. , Signore Armando P. , Zigmond Michael J.
Published: December 2005

6‐Hydroxydopamine (6‐OHDA)‐induced loss of dopamine (DA) neurons has served to produce an animal model of DA neuron loss in Parkinson's disease. We report here the use of 6‐OHDA to produce an in vitro model of this phenomena using dissociated cultures prepared from neonatal rat mesencephalon. Cultures were exposed to 6‐OHDA (40–100 μm, 15 min) in an antioxidant medium, and DA and GABA neurons evaluated by immunocytochemistry. 6‐OHDA induced morphological and biochemical signs of cell death in DA neurons within 3 h, followed by loss of tyrosine hydroxylase immunoreactive neurons within 2 days. In substantia nigra (SN) cultures, DA neurons were much more affected by 6‐OHDA than were GABA neurons. In contrast, DA neurons from the ventral tegmental area were only lost at higher, non‐specific concentrations of 6‐OHDA. The effects of 6‐OHDA on nigral DA neurons were blocked by inhibitors of high affinity DA transport and by z‐DEVD‐fmk (150 μm), a caspase inhibitor. Glial cell line‐derived neurotrophic factor (GDNF) treatment reduced TUNEL labeling 3 h after 6‐OHDA exposure, but did not prevent loss of DA neurons at 48 h. Thus, 6‐OHDA can selectively destroy DA neurons in post‐natal cultures of SN, acting at least in part by initiating caspase‐dependent apoptosis, and this effect can be attenuated early but not late by GDNF.
Cellular localization and functional expression of P‐glycoprotein in rat astrocyte cultures
Authors: Ronaldson Patrick T. , Bendayan Moise , Gingras Diane , Piquette‐Miller Micheline , Bendayan Reina
Published: December 2005

We investigated the cellular/subcellular localization and functional expression of P‐glycoprotein, an ATP‐dependent membrane‐associated efflux transporter, in astrocytes, a brain parenchyma compartment that is poorly characterized for the expression of membrane drug transporters. Analyses were carried out on primary cultures of astrocytes isolated from the cerebral cortex of neonatal Wistar rats and CTX TNA2, an immortalized rat astrocyte cell line. Both cell cultures display morphological features typical of type I astrocytes. RT‐PCR analysis revealed mdr1a and mdr1b mRNA in primary cultures of astrocytes and in CTX TNA2 cells. Western blot analysis using the P‐glycoprotein monoclonal C219 antibody detected a single band of appropriate size in both cell systems. Immunocytochemical analysis using the monoclonal antibodies C219 and MRK16 labeled P‐glycoprotein along the plasma membrane, caveolae, coated vesicles and nuclear envelope. Immunoprecipitation studies using the caveolin‐1 polyclonal H‐97 antibody demonstrated that P‐glycoprotein is physically associated with caveolin‐1 in both cell culture systems. The accumulation of [3H]digoxin (an established P‐glycoprotein substrate) by the astrocyte cultures was significantly enhanced in the presence of standard P‐glycoprotein inhibitors and an ATP depleting agent. These results demonstrate the cellular/subcellular location and functional expression of P‐glycoprotein in rat astrocytes and suggest that this glial compartment may play an important role in the regulation of drug transport in the CNS.
Theory for the impact of basal turnover on dopamine clearance kinetics in the rat striatum after medial forebrain bundle stimulation and pressure ejection
Authors: Michael Adrian C. , Borland Laura M. , J. Mitala Joseph , Willoughby Bridget M. , Motzko Christina M.
Published: December 2005

Although microdialysis measurements suggest that extracellular dopamine concentrations in the rat striatum are in the low nanomolar range, some recent voltammetry studies suggest that the concentration may be considerably higher, perhaps in the micromolar range. The presence of such high dopamine levels in the extracellular space has to be rationalized with the rapid, linear clearance of extracellular dopamine observed after electrical stimulation of the medial forebrain bundle. Kinetic analysis of dopamine clearance after evoked release suggests that the basal extracellular dopamine concentration is below theKMof dopamine uptake, which is near 0.2 µm. However, dopamine clearance after pressure ejection of dopamine into the rat striatum is slow and non‐linear, which may alternatively be a sign that basal dopamine release is only slightly slower than the maximal velocity of dopamine uptake,Vmax. A high basal extracellular dopamine concentration would exist if basal dopamine release were only slightly slower than theVmaxof uptake. This report introduces a new kinetic analysis of dopamine uptake that sheds light on the possible source of the different clearance rates observed following evoked dopamine release and dopamine pressure ejection. Furthermore, the analysis rationalizes the rapid dopamine clearance after evoked release with the possibility that basal extracellular dopamine levels are above theKMof the transporter.
Familial Alzheimer's disease mutations inhibit γ‐secretase‐mediated liberation of β‐amyloid precursor protein carboxy‐terminal fragment
Authors: Wiley Jesse C. , Hudson Mark , Kanning Kevin C. , Schecterson Leslyanne C. , Bothwell Mark
Published: December 2005

Cleavage of the β‐secretase processed β‐amyloid precursor protein by γ‐secretase leads to the extracellular release of Aβ42, the more amyloidogenic form of the β‐amyloid peptide, which subsequently forms the amyloid‐plaques diagnostic of Alzheimer's disease. Mutations in β‐amyloid precursor protein (APP), presenilin‐1 and presenilin‐2 associated with familial Alzheimer's disease (FAD) increase release of Aβ42, suggesting that FAD may directly result from increased γ‐secretase activity. Here, we show that familial Alzheimer's disease mutations clustered near the sites of γ‐secretase cleavage actually decrease γ‐secretase‐mediated release of the intracellular fragment of APP (CTFγ). Concordantly, presenilin‐1 mutations that result in Alzheimer's disease also decrease the release of CTFγ. Mutagenesis of the epsilon cleavage site in APP mimicked the effects of the FAD mutations, both decreasing CTFγ release and increasing Aβ42 production, suggesting that perturbation of this site may account for the observed decrement in γ‐secretase‐mediated proteolysis of APP. As CTFγ has been implicated in transcriptional activation, these data indicate that decreased signaling and transcriptional regulation resulting from FAD mutations in β‐amyloid precursor protein and presenilin‐1 may contribute to the pathology of Alzheimer's disease.
Acute anoxia induces tau dephosphorylation in rat brain slices and its possible underlying mechanisms
Authors: Liu Rong , Pei Jin‐Jing , Wang Xiao‐Chuan , Zhou Xin‐Wen , Tian Qing , Winblad Bengt , Wang Jian‐Zhi
Published: December 2005

Abnormal phosphorylation of microtubule‐associated protein tau plays a critical role in Alzheimer's disease (AD), together with a distinct decrease of energy metabolism in the affected brain regions. To explore the effect of acute energy crisis on tau phosphorylation and the underlying mechanisms, we incubated rat brain slices in artificial cerebrospinal fluid (aCSF) at 37°C with or without an oxygen supply, or in aCSF with low glucose concentrations. Then, the levels of total, phosphorylated and unphosphorylated tau, as well as the activities and levels of protein phosphatase (PP)‐1, PP‐2A, glycogen synthase kinase 3 (GSK‐3), extracellular signal‐regulated protein kinase (ERK) and C‐jun amino terminal kinase (JNK), were measured. It was found, unexpectedly, that tau was significantly dephosphorylated at Ser396/Ser404 (PHF‐1), Ser422 (R145), Ser199/Ser202 (Tau‐1), Thr181 (AT270), Ser202/Thr205 (AT8) and Thr231 (AT180) by acute anoxia for 30 min or 120 min. The activity of PP‐2A and the level of dephosphorylated PP‐2A catalytic subunit at tyrosine 307 (Tyr307) were simultaneously increased. The active forms of ERK1/2 and JNK1/2 were decreased under anoxic incubation. The PP‐2A inhibitor, okadaic acid (OA, 0.75 µm), completely prevented tau from acute anoxia‐induced dephosphorylation and restored the active forms of ERK1/2 and JNK1/2 to the control level. The activities and protein levels of GSK‐3 and PP‐1 showed no change during acute anoxia. These data suggest that acute anoxia induces tau dephosphorylation, and that PP‐2A may play a key role in tau dephosphorylation induced by acute anoxia.
In vivo evidence of CHIP up‐regulation attenuating tau aggregation
Authors: Sahara Naruhiko , Murayama Miyuki , Mizoroki Tatsuya , Urushitani Makoto , Imai Yuzuru , Takahashi Ryosuke , Murata Shigeo , Tanaka Keiji , Takashima Akihiko
Published: December 2005

The carboxyl terminus of heat‐shock cognate (Hsc)70‐interacting protein (CHIP) is a ubiquitin E3 ligase that can collaborate with molecular chaperones to facilitate protein folding and prevent protein aggregation. Previous studies showed that, together with heat‐shock protein (Hsp)70, CHIP can regulate tau ubiquitination and degradation in a cell culture system. Ubiquitinated tau is one component in neurofibrillary tangles (NFTs), which are a major histopathological feature of Alzheimer's disease (AD). However, the precise sequence of events leading to NFT formation and the mechanisms involved remain unclear. To confirm CHIP's role in suppressing NFT formation in vivo, we performed a quantitative analysis of CHIP in human and mouse brains. We found increased levels of CHIP and Hsp70 in AD compared with normal controls. CHIP levels in both AD and controls corresponded directly to Hsp90 levels, but not to Hsp70 or Hsc70 levels. In AD samples, CHIP was inversely proportional to sarkosyl‐insoluble tau accumulation. In a JNPL3 mouse brain tauopathy model, CHIP was widely distributed but weakly expressed in spinal cord, which was the most prominent region for tau inclusions and neuronal loss. Protein levels of CHIP in cerebellar regions of JNPL3 mice were significantly higher than in non‐transgenic littermates. Human tau was more highly expressed in this region of mouse brains, but only moderate levels of sarkosyl‐insoluble tau were detected. This was confirmed when increased insoluble tau accumulation was found in mice lacking CHIP. These findings suggest that increases in CHIP may protect against NFT formation in the early stages of AD. If confirmed, this would indicate that the quality‐control machinery in a neuron might play an important role in retarding the pathogenesis of tauopathies.
Differences in gene expression profile among SH‐SY5Y neuroblastoma subclones with different neurite outgrowth responses to nerve growth factor
Authors: Oe Tomoya , Sasayama Takeshi , Nagashima Takeyuki , Muramoto Masakazu , Yamazaki Takao , Morikawa Noriyuki , Okitsu Osamau , Nishimura Shintaro , Aoki Toshiaki , Katayama Yoshiki , Kita Yasuhiro
Published: December 2005

Nerve growth factor (NGF) plays a key role in the differentiation of neurons. In this study, we established three NGF‐induced neurite‐positive (NIN+) subclones that showed high responsiveness to NGF‐induced neurite outgrowth and three NGF‐induced neurite‐negative (NIN–) subclones that abolished NGF‐induced neurite outgrowth from parental SH‐SY5Y cells, and analyzed differences in the NGF signaling cascade. The NIN+ subclones showed enhanced responsiveness to FK506‐mediated neurite outgrowth as well. To clarify the mechanism behind the high frequency of NGF‐induced neurite outgrowth, we investigated differences in NGF signaling cascade among subclones. Expression levels of the NGF receptor TrkA, and NGF‐induced increases in mRNAs for the immediate‐early genes (IEGs) c‐fosand NGF inducible (NGFI) genesNGFI‐A,NGFI‐BandNGFI‐C, were identical among subclones. Microarray analysis revealed that the NIN+ cell line showed a very different gene expression profile to the NIN– cell line, particularly in terms of axonal vesicle‐related genes and growth cone guidance‐related genes. Thus, the difference in NGF signaling cascade between the NIN+ and NIN– cell lines was demonstrated by the difference in gene expression profile. These differentially expressed genes might play a key role in neurite outgrowth of SH‐SY5Y cells in a region downstream from the site of induction of IEGs, or in a novel NGF signaling cascade.
Angiotensin II induces peroxisome proliferator‐activated receptor gamma in PC12W cells via angiotensin type 2 receptor activation
Authors: Zhao Yi , Foryst‐Ludwig Anna , Bruemmer Dennis , Culman Juraj , Bader Michael , Unger Thomas , Kintscher Ulrich
Published: December 2005

The angiotensin type 2 (AT2) receptor has been previously demonstrated to exert neuroprotective actions possibly by inducing neuronal cell differentiation involving neurite outgrowth. The nuclear hormone receptor peroxisome proliferator‐activated receptor gamma (PPARγ) is an important transcriptional regulator of cell differentiation. The aim of the present study was to clarify whether PPARγ is involved in AT2‐receptor‐mediated morphological neuronal cell differentiation. To investigate AT2‐receptor‐mediated morphological neuronal cell differentiation, rat pheochromocytoma cells (PC12W cells) expressing AT2 but not AT1 receptors, were stimulated with angiotensin II (Ang II, 100 nmol/L) ± the PPARγ antagonists GW9662 (3 µmol/L) and bisphenol A diglycidyl ether (BADGE, 1 µmol/L), and neurite outgrowth of these cells was assessed. Ang II induced neurite outgrowth by 19 ± 1.6‐fold (p < 0.01). Antagonizing PPARγ activity by GW9662 or BADGE potently blocked Ang II‐induced neurite outgrowth (Ang II + GW9662: 6.6 ± 1.5‐fold,p < 0.05; Ang II + BADGE: 1.3 ± 0.7‐fold,p < 0.01). AT2 receptor activation by Ang II markedly induced mRNA and protein expression of the PPARγ2 isoform and enhanced ligand‐induced PPARγ activity in transactivation assays. In conclusion, the present study demonstrates that Ang II induces PPARγ expression and ligand‐mediated PPARγ activity via AT2 receptor activation, which appears to be a crucial process in AT2 receptor mediated neurite outgrowth. AT2 receptor/PPARγ‐dependent neurite outgrowth may play an important role during neuroprotective processes.
PERK is responsible for the increased phosphorylation of eIF2α and the severe inhibition of protein synthesis after transient global brain ischemia
Authors: Owen Cheri R. , Kumar Rita , Zhang Peichuan , McGrath Barbara C. , Cavener Douglas R. , Krause Gary S.
Published: December 2005

Reperfusion after global brain ischemia results initially in a widespread suppression of protein synthesis in neurons that is due to inhibition of translation initiation as a result of the phosphorylation of the α‐subunit of eukaryotic initiation factor 2 (eIF2). To address the role of the eIF2α kinase RNA‐dependent protein kinase‐like endoplasmic reticulum kinase (PERK) in the reperfused brain, transgenic mice with a targeted disruption of the Perk gene were subjected to 20 min of forebrain ischemia followed by 10 min of reperfusion. In wild‐type mice, phosphorylated eIF2α was detected in the non‐ischemic brain and its levels were elevated threefold after 10 min of reperfusion. Conversely, there was no phosphorylated eIF2α detected in the non‐ischemic transgenic mice and there was no sizeable rise in phosphorylated eIF2α levels in the forebrain after ischemia and reperfusion. Moreover, there was a substantial rescue of protein translation in the reperfused transgenic mice. Neither group showed any change in total eIF2α, phosphorylated eukaryotic elongation factor 2 or total eukaryotic elongation factor 2 levels. These data demonstrate that PERK is responsible for the large increase in phosphorylated eIF2α and the suppression of translation early in reperfusion after transient global brain ischemia.
Lithium enhances secretion from large dense‐core vesicles in nerve growth factor‐differentiated PC12 cells
Authors: Umbach Joy A. , Zhao Ying , Gundersen Cameron B.
Published: December 2005

Considerable attention has been focused on the therapeutic role of lithium (Li) in bipolar disorders. Although no consensus has emerged, Li presumably influences the behavior of neurons that regulate mood and behavior. Using PC12 cells to study cellular and molecular actions of Li, we previously reported that Li modulates the expression of proteins associated with large dense‐core vesicles (LDCVs; organelles typically containing monoamines, neuropeptides and other cargo proteins). The current investigation indicates that this enhanced expression of LDCV proteins correlates with an altered secretory phenotype in Li‐treated cells. Immunoblotting detects significant increases in the cellular content and secretion of the LDCV cargo proteins chromogranin B and secretogranin II. Amperometry reveals an increase of spike number elicited by K+‐depolarization of Li‐treated cells but no change of spike amplitude or kinetics. Electron microscopy reveals no significant change in LDCV number per unit area in Li‐treated cells. However, there is a significant increase (about 15%) in the diameter of LDCVs after Li. Thus, Li induces changes in the properties of LDCVs that culminate in augmented regulated secretion in nerve growth factor‐differentiated PC12 cells. These results extend our understanding of Li‐dependent changes of cellular function that may be germane to the therapeutic action of Li.
Deficits in dopamine clearance and locomotion in hypoinsulinemic rats unmask novel modulation of dopamine transporters by amphetamine
Authors: Owens W. Anthony , Sevak Rajkumar J. , Galici Ruggero , Chang Xiaoying , Javors Martin A. , Galli Aurelio , France Charles P. , Daws Lynette C.
Published: December 2005

Insulin affects brain reward pathways and there is converging evidence that this occurs through insulin regulation of the dopamine (DA) transporter (DAT). In rats made hypoinsulinemic by fasting, synaptosomal DA uptake is reduced. Interestingly, [3H]DA uptake is increased in hypoinsulinemic rats with a history of amphetamine self‐administration. The possibility that amphetamine and insulin act in concert to regulate DAT activity prompted this study. Here we show that [3H]DA uptake, measuredin vitroand clearance of exogenously applied DAin vivo, is significantly reduced in rats made hypoinsulinemic by a single injection of streptozotocin. Strikingly, amphetamine (1.78 mg/kg, given every other day for 8 days) restored DA clearance in streptozotocin‐treated rats but was without effect on DA clearance in saline‐treated rats. Basal locomotor activity of streptozotocin‐treated rats was lower compared to control rats; however, in streptozotocin‐treated rats, hyperlocomotion induced by amphetamine increased over successive amphetamine injections. In saline‐treated rats the locomotor stimulant effect of amphetamine remained stable across the four amphetamine injections. These results provide exciting new evidence that actions of amphetamine on DA neurotransmission are insulin‐dependent and further suggest that exposure to amphetamine may cause long‐lasting changes in DAT function.
Role of NADPH oxidase in the brain injury of intracerebral hemorrhage
Authors: Tang Jiping , Liu Jun , Zhou Changman , Ostanin Dmitry , Grisham Matthew B. , Neil Granger D. , Zhang John H.
Published: December 2005

The major risk factors for intracerebral hemorrhage (ICH) are hypertension and aging. A fundamental mechanism for hypertension‐ and aging‐induced vascular injury is oxidative stress. We hypothesize that oxidative stress has a crucial role in ICH. To test our hypothesis, we used bacterial collagenase to produce ICH in wild‐type C57BL/6 andgp91phoxknockout (gp91phoxKO) mice (deficient in gp91phoxsubunit of the superoxide‐producing enzyme NADPH oxidase). All animals were studied at 20–35 weeks of age, resembling an older patient population. We found that collagenase produced less bleeding ingp91phoxKO mice than wild‐type mice. Total oxidative product was lower ingp91phoxKO mice than in wild‐type mice, both under basal conditions and after ICH. Consistent with the ICH volume, brain edema formation, neurological deficit and a high mortality rate was noted in wild‐type but not ingp91phoxKO mice. This ICH‐induced brain injury in wild‐type mice is associated with enhanced expression of thegp91phoxsubunit of NADPH oxidase. In conclusion, the oxidative stress resulting from activation of NADPH oxidase contributes to ICH induced by collagenase and promotes brain injury.
Molecular and functional characterization of an Na+‐independent choline transporter in rat astrocytes
Authors: Inazu Masato , Takeda Hiroshi , Matsumiya Teruhiko
Published: December 2005

In this study, we examined the molecular and functional characterization of choline uptake into cultured rat cortical astrocytes. Choline uptake into astrocytes showed little dependence on extracellular Na+. Na+‐independent choline uptake was saturable and mediated by a single transport system, with an apparent Michaelis–Menten constant (Km) of 35.7 ± 4.1 µmand a maximal velocity (Vmax) of 49.1 ± 2.0 pmol/mg protein/min. Choline uptake was significantly decreased by acidification of the extracellular medium and by membrane depolarization. Na+‐independent choline uptake was inhibited by unlabeled choline, acetylcholine and the choline analogue hemicholinium‐3. The prototypical organic cation tetrahexylammonium (TEA), and othern‐tetraalkylammonium compounds such as tetrabutylammonium (TBA) and tetrahexylammonium (THA), inhibited Na+‐independent choline uptake, and their inhibitory potencies were in the order THA > TBA > TEA. Various organic cations, such as 1‐methyl‐4‐tetrahydropyridinium (MPP+), clonidine, quinine, quinidine, guanidine,N‐methylnicotinamide, cimetidine, desipramine, diphenhydramine and verapamil, also interacted with the Na+‐independent choline transport system. Corticosterone and 17β‐estradiol, known inhibitors of organic cation transporter 3 (OCT3), did not cause any significant inhibition. However, decynium22, which inhibits OCTs, markedly inhibited Na+‐independent choline uptake. RT‐PCR demonstrated that astrocytes expressed low levels of OCT1, OCT2 and OCT3 mRNA, but the functional characteristics of choline uptake are very different from the known properties of these OCTs. The high‐affinity Na+‐dependent choline transporter, CHT1, is not expressed in astrocytes as evidenced by RT‐PCR. Furthermore, mRNA for choline transporter‐like protein 1 (CTL1), and its splice variants CTL1a and CTL1b, was expressed in rat astrocytes, and the inhibition of CTL1 expression by RNA interference completely inhibited Na+‐independent choline uptake. We conclude that rat astrocytes express an intermediate‐affinity Na+‐independent choline transport system. This system seems to occur through a CTL1 and is responsible for the uptake of choline and organic cations in these cells.
Phosphorylation of tau at serine 416 by Ca2+/calmodulin‐dependent protein kinase II in neuronal soma in brain
Authors: Yamamoto Hideyuki , Hiragami Yukari , Murayama Miyuki , Ishizuka Koko , Kawahara Masahiro , Takashima Akihiko
Published: December 2005

It is well known that tau is a good in vitro substrate for Ca2+/calmodulin‐dependent protein kinase II (CaM kinase II). However, it is not clear at present whether CaM kinase II phosphorylates tau in vivo or not. Serine 416, numbered according to the longest human tau isoform, has been reported to be one of the major phosphorylation sites by CaM kinase II in vitro. In this study, we produced a specific antibody against tau phosphorylated at serine 416 (PS416‐tau). Immunoblot analysis revealed that the antibody reacted with tau in the rat brain extract which was prepared in the presence of protein phosphatase inhibitors. Developmental study indicated that serine 416 was strongly phosphorylated at early developmental stages in rat brain. We examined the localization of PS416‐tau in primary cultured hippocampal neurons and the immortalized GnRH neurons (GT1‐7 cells), which were stably transfected with CaM kinase IIα cDNA. Immunostaining of these cells indicated that tau was phosphorylated mainly in neuronal soma. Interestingly, tau in neuronal soma in Alzheimer's disease (AD) brain was strongly immunostained by the antibody. These results suggest that CaM kinase II is involved in the accumulation of tau in neuronal soma in AD brain.
Lovastatin inhibits the growth and survival pathway of phosphoinositide 3‐kinase/protein kinase B in immortalized rat brain neuroblasts
Authors: Cerezo‐Guisado Maria Isabel , Garcia‐Marin Luis Jesus , Lorenzo Maria Jesus , Bragado Maria Julia
Published: December 2005

We previously showed that lovastatin, an HMG‐CoA reductase inhibitor, suppresses cell growth by inducing apoptosis in rat brain neuroblasts. Our aim was to study intracellular signalling induced by lovastatin in neuroblasts. Lovastatin significantly decreases the phosphoinositide 3‐kinase (PI3‐K) activity in a concentration‐dependent manner. Expression of p85 subunit and its association with phosphotyrosine‐containing proteins are unaffected by lovastatin. Lovastatin decreases protein kinase B (PKB)/Akt phosphorylation, and its downstream effectors, p70S6K and the eukaryotic initiation factor 4E (eIF4E) regulatory protein 1, 4E‐BP1, in a concentration‐dependent manner, and reduces p70S6K expression. Lovastatin effects are fully prevented with mevalonate. Only the highest dose of PI3‐K inhibitors that significantly reduce PI3‐K kinase activity induces apoptosis in neuroblasts but to a lower degree than lovastatin. In summary, this work shows that treatment of brain neuroblasts with lovastatin leads to an inhibition of the main pathway that controls cell growth and survival, PI3‐K/PKB and the subsequent blockade of downstream proteins implicated in the regulation of protein synthesis. This work suggests that inactivation of the antiapoptotic PI3‐K appears insufficient to induce the degree of neuroblasts apoptosis provoked by lovastatin, which must necessarily involve other intracellular pathways. These findings might contribute to elucidate the molecular mechanisms of some statins effects in the central nervous system.
Species Review of Amphibian Extinction Risks in Madagascar: Conclusions from the Global Amphibian Assessment
Authors: ANDREONE FRANCO , CADLE JOHN E. , COX NEIL , GLAW FRANK , NUSSBAUM RONALD A. , RAXWORTHY CHRISTOPHER J. , STUART SIMON N. , VALLAN DENIS , VENCES MIGUEL
Published: December 2005

Abstract:  We assessed the extinction risks of Malagasy amphibians by evaluating their distribution, occurrence in protected areas, population trends, habitat quality, and prevalence in commercial trade. We estimated and mapped the distribution of each of the 220 described Malagasy species and applied, for the first time, the IUCN Red List categories and criteria to all species described at the time of the assessment. Nine species were categorized as critically endangered, 21 as endangered, and 25 as vulnerable. The most threatened species occur on the High Plateau and/or have been subjected to overcollection for the pet trade, but restricted extent of occurrence and ongoing habitat destruction were identified as the most important factors influencing extinction threats. The two areas with the majority of threatened species were the northern Tsaratanana‐Marojejy‐Masoala highlands and the southeastern Anosy Mountains. The current system of protected areas includes 82% of the threatened amphibian species. Of the critically endangered species, 6 did not occur in any protected area. For conservation of these species we recommend the creation of a reserve for the species of the Mantella aurantiaca group, the inclusion of two Scaphiophryne species in the Convention on the International Trade in Endangered Species Appendix II, and the suspension of commercial collecting for Mantella cowani. Field surveys during the last 15 years reveal no pervasive extinction of Malagasy amphibians resulting from disease or other agents, as has been reported in some other areas of the world.
Co‐Management Approach to Marine Conservation in Mohéli, Comoros Islands
Authors: GRANEK ELISE F. , BROWN MARK A.
Published: December 2005

Abstract:  Many developing countries experience habitat degradation and unsustainable natural resource exploitation, with biodiversity and habitat conservation efforts often impeded by political instability and limited funding. Challenges in previous conservation efforts coupled with the current rate of marine habitat degradation and species declines warrant consideration of an innovative conservation approach. Co‐management of protected areas addresses biological, cultural, economic, and political concerns and empowers communities through collaboration and integration in conservation efforts. It provides flexibility for adaptive practices to address underlying socioeconomic factors affecting conservation efforts and may compensate for limited or missing scientific data. The ecosystems of the Comoros Islands in the West Indian Ocean, a biodiversity hotspot with high endemism and diverse tropical marine habitats, are adversely affected by existing ecological, socioeconomic, and political conditions. Mohéli Marine Park was designed to address threats to the marine environment and is a model for co‐management practices. We conducted a year‐long evaluation of the park implementation process, including community and fisher participation. After 3 years of operation with 80% local community control, the park maintains a small staff to monitor sea turtle nesting beaches, reef health, fisheries, and uninhabited islets and to guide ecotourists and educate visitors. Our analysis revealed successes and shortcomings of the co‐management approach. Successes included local communities empowered to participate in natural resource management, increased local involvement in conservation initiatives, and use of traditional knowledge when scientific information was unavailable. The Comoros example also illustrates that co‐management is not immune to social issues, inadequate government law enforcement, or political instability and is an incomplete substitute for sound science. Lessons learned are applicable elsewhere and offer a template for effective scientific research and monitoring, policy making, and management of protected areas in developing nations.
Science and Society: Marine Reserve Design for the California Channel Islands
Authors: DAVIS GARY E.
Published: December 2005

Abstract:  We explored the interaction of science and society in attempts to restore impaired marine ecosystems in Channel Islands National Park and National Marine Sanctuary, California. Deteriorating resource conditions triggered a community's desire to change public policy. Channel Islands National Park, one of 40 marine protected areas in the U.S. National Park System, was proclaimed a national monument in 1938 and expanded substantially in 1980 by an act of Congress. Collapse of marine life populations and loss of 80% of the giant kelp (Macrocystis pyrifera) forests in the park between 1980 and 1998 showed that habitat and water quality protection alone had not secured sustainable ocean ecosystems or fisheries. The failed fishery management strategies and practices prompted formal community and agency requests in 1998 for a network of reserves protected from direct fishing impacts to serve as marine recovery areas. A 2‐year attempt to build a community consensus based on science for a reserve network successfully identified recovery goals for fisheries, biodiversity, education, economics, and heritage values. Nevertheless, the community group failed to garner unanimous support for a specific reserve network to achieve those common goals. The group submitted a recommendation, supported by 14 of 16 members, to state and federal authorities in 2001 for action in their respective jurisdictions. California adopted the half of the network in state waters in 2003. This process exposed the socioeconomic factors involved in the design of marine protected areas that can be negotiated successfully among groups of people and factors determined by nature that cannot be negotiated. Understanding the differences among the factors was crucial in reaching consensus and changing public policy.
Knowledge Management Using Information Technology: Determinants of Short‐Term Impact on Firm Value*
Authors: Sabherwal Rajiv , Sabherwal Sanjiv
Published: December 2005

The importance of knowledge management (KM) processes for organizational performance is now well recognized. Seeking to better understand the short‐term impact of KM on firm value, this article focuses on public announcements of information technology (IT)‐based KM efforts, and uses cumulative abnormal return (CAR) associated with an announcement as the dependent variable. This article employs a contingency approach, arguing that the KM announcement would have a positive short‐term impact on firm value in some conditions but not in others. Thus, it pursues the following research question: What are the effects of contextual factors on the CAR associated with the announcement of an IT‐based KM effort? Specific hypotheses are proposed based on information‐processing theory, organizational learning theory, the knowledge‐based theory of the firm, and the theory of knowledge creation. These hypotheses link CARs to alignment between industry innovativeness and the KM process, alignment between firm efficiency and the KM process, firm‐specific instability, and firm diversification. The empirical study utilizes secondary data on 89 KM announcements from 1995 to 2002. The results largely support the hypotheses. Overall, this article provides empirical support for the theory‐based arguments, and helps develop a contingency framework of the effectiveness of KM efforts.
Discount Pricing Policies and the Coordination of Decentralized Distribution Systems*
Authors: Wang Qinan
Published: December 2005

A challenge of supply chain management is to align the objectives, and hence coordinate the activities, of independent supply chain members. In this study, we approach this problem in a simple way by extending traditional quantity discounts that are based solely on buyers' individual order size to discount policies that are based on both buyers' individual order size and their annual volume. We show that discount policies are able to achieve nearly optimal system profit and, hence, provide effective coordination, for a decentralized two‐echelon distribution system, whereby a supplier sells a product to a group of heterogeneous and independent retailers each facing a downward‐sloping demand curve of its retail price. When buyers are heterogeneous, a critical issue of coordination is to motivate different customers to increase their demand and lot size according to their potential so as to improve profits. We show that market heterogeneity presents an effective discriminating factor for the supplier to segment customers in the design of a coordination mechanism.
Regression Analysis of Doubly Censored Failure Time Data with Frailty
Authors: Kim Yang‐Jin
Published: December 2005

SummaryIn doubly censored failure time data, the survival time of interest is defined as the elapsed time between an initial event and a subsequent event, and the occurrences of both events cannot be observed exactly. Instead, only right‐ or interval‐censored observations on the occurrence times are available. For the analysis of such data, a number of methods have been proposed under the assumption that the survival time of interest is independent of the occurrence time of the initial event. This article investigates a different situation where the independence may not be true with the focus on regression analysis of doubly censored data. Cox frailty models are applied to describe the effects of covariates and an EM algorithm is developed for estimation. Simulation studies are performed to investigate finite sample properties of the proposed method and an illustrative example from an acquired immune deficiency syndrome (AIDS) cohort study is provided.
An Alternative Chemical Route for the Synthesis and Thermal Stability of Chemically Enriched Hydroxyapatite
Authors: Choi Daiwon , Kumta Prashant N.
Published: December 2005

Hydroxyapatite (HAp: Ca10(PO4)6(OH)2) was synthesized by aqueous precipitation using CaCl2 and Na3PO4 with NaOH added to ensure completion of the reaction at room temperature. The HAp powder prepared using stoichiometric amounts of NaOH was stable even at 1200°C, but the HAp prepared with sub‐stoichiometric amounts of NaOH resulted in its transformation into β‐tricalcium phosphate at 600°C. The reaction pH, X‐ray diffraction, thermal analysis, scanning electron microscopy, Fourier transform infrared analyses and inductively coupled plasma‐optical emission spectroscopy were used to characterize the phase purity, thermal stability, morphology, and chemical composition of the synthesized HAp powder.
Raman Microspectroscopic Characterization of Amorphous Silica Plastic Behavior
Authors: Perriot A. , Vandembroucq D. , Barthel E. , Martinez V. , Grosvalet L. , Martinet Ch. , Champagnon B.
Published: December 2005

Raman microspectroscopy was used to characterize amorphous silica plastic behavior. Using a correlation between Raman spectrum and density, a map of the local residual indentation‐induced densification was obtained. The existence of a densification‐induced hardening was also evidenced through a diamond–anvil cell experiment. Such observations are not accounted for by the previously proposed hardening‐free pressure‐dependent yield criterion based on indentation curves. These results open the way toward a more accurate description of a constitutive law for amorphous silica.
The Internationalization of Chinese Firms: A Case for Theoretical Extension?
Authors: Child John , Rodrigues Suzana B.
Published: December 2005

This paper examines the patterns of, and motives for, internationalization by prominent market-seeking Chinese firms. Case studies of these firms indicate that they are seeking technological and brand assets to create a competitive position in international markets. While mainstream theory tends to assume that firms internationalize to exploit competitive advantages, Chinese firms are generally making such investments in order to address competitive disadvantages. They are engaging in ‘inward’ internationalization by means of original equipment manufacture (OEM) and joint venture partnerships, and ‘outward’ internationalization by means of acquisition and organic expansion abroad. Each of these routes offers certain benefits coupled with its own challenges or risks. The paper concludes that the Chinese case offers an opportunity to extend present theorizing in four primary areas concerning the latecomer perspective and catch-up strategies, institutional analysis with reference to the role of government, the relations between entrepreneurs and institutions, and the liability of foreignness.
Placing narrative correspondence in the geographer's toolbox: Insights from care research in New Zealand
Authors: Milligan Christine
Published: December 2005

Abstract:  To date, narrative correspondence has been relatively neglected as a geographical research method. This is particularly true within the subdiscipline of health geography. Drawing on the literature on narrative approaches, as well as a qualitative study of informal carers’ experiences of transitions in the place of care for older people in New Zealand, this paper highlights the valuable insights this approach can offer into the interrelationship between people, place and care. More specifically, it explores how caring identities and care related behaviour are constructed and reconstructed over time and space.
Active Microbial Sulfur Disproportionation in the Mesoproterozoic
Authors: Johnston David T. , Wing Boswell A. , Farquhar James , Kaufman Alan J. , Strauss Harald , Lyons Timothy W. , Kah Linda C. , Canfield Donald E.
Published: December 2005

The environmental expression of sulfur compound disproportionation has been placed between 640 and 1050 million years ago (Ma) and linked to increases in atmospheric oxygen. These arguments have their basis in temporal changes in the magnitude of
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S/
32
S fractionations between sulfate and sulfide. Here, we present a Proterozoic seawater sulfate isotope record that includes the less abundant sulfur isotope
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S. These measurements imply that sulfur compound disproportionation was an active part of the sulfur cycle by 1300 Ma and that progressive Earth surface oxygenation may have characterized the Mesoproterozoic.
Divergent Immunoglobulin G Subclass Activity Through Selective Fc Receptor Binding
Authors: Nimmerjahn Falk , Ravetch Jeffrey V.
Published: December 2005

Subclasses of immunoglobulin G (IgG) display substantial differences in their ability to mediate effector responses, contributing to variable activity of antibodies against microbes and tumors. We demonstrate that the mechanism underlying this long-standing observation of subclass dominance in function is provided by the differential affinities of IgG subclasses for specific activating IgG Fc receptors compared with their affinities for the inhibitory IgG Fc receptor. The significant differences in the ratios of activating-to-inhibitory receptor binding predicted the in vivo activity. We suggest that these highly predictable functions assigned by Fc binding will be an important consideration in the design of therapeutic antibodies and vaccines.