bluuebunny/crossref_metadata_2025 huggingface dataset:
Peripheral inflammation increases Scya2 expression in sensory ganglia and cytokine and endothelial related gene expression in inflamed tissue
Authors: Yang Hsiu‐Ying T. , Mitchell Kendall , Keller Jason M. , Iadarola Michael J.
Published: September 2007

The sensation of pain (nociception) is a critical factor in host defense during tissue injury and inflammation and is initiated at the site of injury by activation of primary afferent C‐fiber and A‐∂ nerve endings. Inflammation induces tissue alterations that sensitize these nociceptive nerve terminals, contributing to persistent pain. To understand this ‘algesic tissue environment’ and peripheral nervous signaling to the CNS and immune system, we examined cytokine and endothelial‐related gene expression profiles in inflamed rat tissues and corresponding dorsal root ganglia (DRG) by microarray and RT‐PCR following hind paw injection of carrageenan. In inflamed tissue, forty‐two cytokine and endothelial‐related genes exhibited elevated expression. In contrast, in DRG, only Scya2 (chemokine C‐C motif ligand 2) mRNA was up‐regulated, leading to an increase in its gene product monocyte chemoattractant protein‐1. Scya2 mRNA was localized byin situhybridization‐immunocytochemical double‐labeling to a subpopulation of vanilloid receptor‐1 (transient receptor potential vanilloid subtype 1) containing neurons, and its expression was increased by direct transient receptor potential vanilloid subtype 1 stimulation with the vanilloid agonist resiniferatoxin, indicating sensitivity to nociceptive afferent activity. Our results are consistent with the idea that monocyte chemoattractant protein‐1 at the site of peripheral injury and/or in DRG is involved in inflammatory hyperalgesia.
Expression of functional Kir6.1 channels regulates glutamate release at CA3 synapses in generation of epileptic form of seizures
Authors: Soundarapandian Mangala M. , Wu Di , Zhong Xiaofen , Petralia Ronald S. , Peng Lisheng , Tu Weihong , Lu Youming
Published: September 2007

The Kir6.1 channels are a subtype of ATP‐sensitive inwardly rectifying potassium (KATP) channels that play an essential role in coupling the cell’s metabolic events to electrical activity. In this study, we show that functional Kir6.1 channels are located at excitatory pre‐synaptic terminals as a complex with type‐1 Sulfonylurea receptors (SUR1) in the hippocampus. The mutant mice with deficiencies in expressing the Kir6.1 or the SUR1 gene are more vulnerable to generation of epileptic form of seizures, compared to wild‐type controls. Whole‐cell patch clamp recordings demonstrate that genetic deletion of the Kir6.1/SUR1 channels enhances glutamate release at CA3 synapses. Hence, expression of functional Kir6.1/SUR1 channels inhibits seizure responses and possibly acts via limiting excitatory glutamate release.
Developmental changes of glycosphingolipids and expression of glycogenes in mouse brains
Authors: Ngamukote Sathaporn , Yanagisawa Makoto , Ariga Toshio , Ando Susumu , Yu Robert K.
Published: September 2007

Glycosphingolipids (GSLs) and their sialic acid‐containing derivatives, gangliosides, are important cellular components and are abundant in the nervous system. They are known to undergo dramatic changes during brain development. However, knowledge on the mechanisms underlying their qualitative and qualitative changes is still fragmentary. In this investigation, we have provided a detailed study on the developmental changes of the expression patterns of GSLs, GM3, GM1, GD3, GD1a, GD2, GD1b, GT1b, GQ1b, A2B5 antigens (c‐series gangliosides such as GT3 and GQ1c), Chol‐1α (GT1aα and GQ1bα), glucosylceramide, galactosylceramide (O1 antigen), sulfatide (O4 antigen), stage‐specific embryonic antigen‐1 (Lewis x) glycolipids, and human natural killer‐1 glycolipid (sulfoglucuronosyl paragloboside) in developing mouse brains [embryonic day 12 (E12) to adult]. In E12–E14 brains, GD3 was a predominant ganglioside. After E16, the concentrations of GD3 and GM3 markedly decreased, and the concentrations of a‐series gangliosides, such as GD1a, increased. GT3, glucosylceramide, and stage‐specific embryonic antigen‐1 were expressed in embryonic brains. Human natural killer‐1 glycolipid was expressed transiently in embryonic brains. On the other hand, Chol‐1α, galactosylceramide, and sulfatide were exclusively found after birth. To provide a better understanding of the metabolic basis for these changes, we analyzed glycogene expression patterns in the developing brains and found that GSL expression is regulated primarily by glycosyltransferases, and not by glycosidases. In parallel studies using primary neural precursor cells in culture as a tool for studying developmental events, dramatic changes in ganglioside and glycosyltransferase gene expression were also detected in neurons induced to differentiate from neural precursor cells, including the expression of GD3, followed by up‐regulation of complex a‐ and b‐series gangliosides. These changes in cell culture systems resemble that occurring in brain. We conclude that the dramatic changes in GSL pattern and content can serve as useful markers in neural development and that these changes are regulated primarily at the level of glycosyltransferase gene expression.
The effects of oxidative stress on parkin and other E3 ligases
Authors: LaVoie Matthew J. , Cortese Giuseppe P. , Ostaszewski Beth L. , Schlossmacher Michael G.
Published: September 2007

Autosomal recessive mutations within the Parkin gene are associated with degeneration of the substantia nigra and locus coeruleus and an inherited form of Parkinson’s disease (PD). As loss‐of‐function mutations in parkin are responsible for a familial variant of PD, conditions that affect wild‐type parkin are likely to be associated with increased risk of idiopathic disease. Previous studies uncovered a unique vulnerability of the parkin protein to dopamine (DA)‐induced aggregation and inactivation. In this study, we compared several proteins that share structural elements or ubiquitinating activity with parkin. We report that oxidative stress in several cell lines and primary neurons induces the aggregation of parkin into high molecular weight species, at least a portion of which are self‐associated homo‐multimers. While parkin was preferentially affected by excess DA, each of the E3 proteins tested were made more insoluble by oxidative stress, and they varied in degree of susceptibility (e.g. parkin > HHARI ≅ CHIP > c‐Cbl > E6AP). These conditions of oxidative stress were also associated with decreased parkin E3 ligase activity. Similar to recently conducted studies on α‐synuclein processing, both macroautophagy and the proteasome participate in parkin degradation, with the proteasome playing the predominant role for normal parkin turnover and macroautophagy being more important in the degradation of aggregated parkin. These data further highlight the selective vulnerability of parkin to DA‐induced modifications, demonstrating for the first time the ability of both endogenous and ectopically expressed parkin to transition into an insoluble state in part through self‐association and oligomer formation.
Monoamine oxidase‐A modulates apoptotic cell death induced by staurosporine in human neuroblastoma cells
Authors: Fitzgerald Julia C. , Ufer Christoph , De Girolamo Luigi A. , Kuhn Hartmut , Billett E. Ellen
Published: September 2007

Monoamine oxidases (MAOs) are mitochondrial enzymes which control the levels of neurotransmitters in the brain and dietary amines in peripheral tissues via oxidative deamination. MAO has also been implicated in cell signalling. In this study, we describe the MAO‐A isoform as functional in apoptosis induced by staurosporine (STS) in human dopaminergic neuroblastoma cells (SH‐SY5Y). Increased levels of MAO‐A activity were induced by STS, accompanied by increased MAO‐A protein and activation of the initiator of the intrinsic pathway, caspase 9, and the executioner caspase 3. MAO‐A mRNA levels were unaffected by STS, suggesting that changes in MAO‐A protein are due to post‐transcriptional events. Two unrelated MAO‐A inhibitors reduced caspase activation. STS treatment resulted in sustained activation of the mitogen‐activated protein kinase pathway enzymes extracellular regulated kinase, c‐jun terminal kinase and p38, and depletion of the anti‐apoptotic protein Bcl‐2. These changes were significantly reversed by MAO inhibition. Production of reactive oxygen species was increased following STS exposure, which was blocked by both MAO inhibition and the antioxidant N‐acetylcysteine. Therefore our data provide evidence that MAO‐A, through its production of reactive oxygen species as a by‐product of its catalytic activity on the mitochondrial surface, is recruited by the cell to enhance apoptotic signalling.
Metabotropic actions of kainate receptors in the CNS
Authors: Rodríguez‐Moreno Antonio , Sihra Talvinder S.
Published: September 2007

Kainate receptors (KARs), together with NMDA and α‐amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionate receptors (AMPA), are typically described as ionotropic glutamate receptors. Although ionotropic functions for KARs are beginning to be characterized in multiple brain regions, both, in the pre‐ and post‐synaptic compartments of the synapse, there is accumulating evidence that KARs mediate some of their effects without invoking ion‐fluxes. Thus, since 1998, when the first metabotropic action of KARs was described in the modulation of GABA release in hippocampal interneurons, there have been increasing reports that some of the functions of KARs involve the participation of intracellular signalling cascades and depend on G protein activation. These surprising observations, attesting metabotropic actions of KARs, akin to those usually attributed to seven transmembrane region G protein‐coupled receptors, make the physiological classification and description of glutamate receptors more complex. In the present review, we describe the metabotropic roles of KARs in the CNS and discuss the intriguing properties of this receptor which, structurally shows all the facets of a typical ionotropic receptor, but appears to express a metabotropic remit at some key synapses.
Which clinical and experimental data link temporal lobe epilepsy with depression?
Authors: Kondziella Daniel , Alvestad Silje , Vaaler Arne , Sonnewald Ursula
Published: September 2007

The association of temporal lobe epilepsy with depression and other neuropsychiatric disorders has been known since the early beginnings of neurology and psychiatry. However, only recently havein vivoandex vivotechniques such as Positron Emission Tomography, Magnetic Resonance Imaging and Magnetic Resonance Spectroscopy in combination with refined animal models and behavioral tests made it possible to identify an emerging pattern of common pathophysiological mechanisms. We now have growing evidence that in both disorders altered interaction of serotonergic and noradrenergic neurons with glutamatergic systems is associated with abnormal neuronal circuits and hyperexcitability. Neuronal hyperexcitability can possibly evoke seizure activity as well as disturbed emotions. Moreover, decreased synaptic levels of neurotransmitters and high glucocorticoid levels influence intracellular signaling pathways such as cAMP, causing disturbances of brain‐derived and other neurotrophic factors. These may be associated with hippocampal atrophy seen on Magnetic Resonance Imaging and memory impairment as well as altered fear processing and transient hypertrophy of the amygdala. Positron Emission Tomography studies additionally suggest hypometabolism of glucose in temporal and frontal lobes. Last, but not least, in temporal lobe epilepsy and depression astrocytes play a role that reaches far beyond their involvement in hippocampal sclerosis and ultimately, therapeutic regulation of glial‐neuronal interactions may be a target for future research. All these mechanisms are strongly intertwined and probably bidirectional such that the structural and functional alterations from one disease increase the risk for developing the other. This review provides an integrative update of the most relevant experimental and clinical data on temporal lobe epilepsy and its association with depression.
Immunomodulatory effects of etanercept in a model of brain injury act through attenuation of the acute‐phase response
Authors: Campbell Sandra J. , Jiang Yanyan , Davis Andrew E. M. , Farrands Rebecca , Holbrook Joanna , Leppert David , Anthony Daniel C.
Published: September 2007

TNF‐α has proved to be a successful target in the treatment of many peripheral inflammatory diseases, but the same interventions worsen immune‐mediated CNS disease. However, anti‐TNF‐α strategies may offer promise as therapy for non‐immune CNS injury. In this study, we have microinjected IL‐1β or lipopolysaccharide (LPS) into the rat brain as a simple model of brain injury and have systemically administered the TNF‐α antagonist etanercept to discover whether hepatic TNF‐α, produced as part of the acute‐phase response to CNS injury, modulates the inflammatory response in the brain. We report a significant reduction in neutrophil numbers recruited to the IL‐1β‐ or LPS‐challenged brain as a result of TNF‐α inhibition. We also show an attenuation in the levels of hepatic mRNA including TNF‐α mRNA and of TNF‐α‐induced genes, such as the chemokines CCL‐2, CXCL‐5, and CXCL‐10, although other chemokines elevated by the injury were not significantly changed. The reduction in hepatic chemokine synthesis results in reduced numbers of circulating neutrophils, and also a reduction in the numbers recruited to the liver as a consequence of brain injury. These findings suggest that TNF‐α inhibitors may reduce CNS inflammatory responses by targeting the hepatic acute‐phase response, and thus therapies for brain injury need not cross the blood–brain barrier to be effective.
Glycogen synthase kinase‐3β: homologous regulation of cell surface insulin receptor level via controlling insulin receptor mRNA stability in adrenal chromaffin cells
Authors: Yokoo Hiroki , Nemoto Takayuki , Yanagita Toshihiko , Satoh Shinya , Yoshikawa Norie , Maruta Toyoaki , Wada Akihiko
Published: September 2007

In cultured bovine adrenal chromaffin cells, 48 h‐treatment with 20 mmol/L LiCl, 1 mmol/L valproic acid, 30 μmol/L SB216763, 30 μmol/L SB415286, or 100 nmol/L insulin, a condition that inhibits constitutive active glycogen synthase kinase‐3 (GSK‐3), decreased cell surface 125I‐insulin binding capacity by ∼39%, without altering the Kd value; LiCl, SB216763 or insulin decreased insulin receptor (IR) and IR precursor levels, attenuating insulin‐induced Tyr‐autophosphorylation of IR. LiCl increased inhibitory Ser9‐phosphorylation of GSK‐3β at 6 h, decreasing 125I‐insulin binding at 24 h. SB216763‐induced 125I‐insulin binding reduction (IC50 = 3 μmol/L) was preceded by β‐catenin level increase by SB216763 (EC50 = 11 μmol/L), a hallmark of GSK‐3 inhibition. Insulin‐induced rapid (> 1 min) Ser9‐phosphorylation of GSK‐3β (Nemoto et al. 2006) was followed by ∼48% decrease of IR level. LiCl did not stimulate endocytosis, nor proteolysis of IR. LiCl destabilized IR mRNA (t1/2 = 9.3 vs. 6.5 h), decreasing IR mRNA level by ∼47%, without altering IR gene transcription. Decreases of 125I‐insulin binding and IR level, as well as increased Ser9‐phosphorylation of GSK‐3β were restored to the control levels by washing the test compound‐treated cells. Thus, GSK‐3β regulates IR level via controlling IR mRNA stability.
Key role of the nicotinic receptor in neurotransmitter exocytosis in human chromaffin cells
Authors: Pérez‐Alvarez Alberto , Albillos Almudena
Published: September 2007

The whole‐cell secretory response evoked by acetylcholine (ACh) in human chromaffin cells was examined using a new protocol based on quickly switching from the voltage‐clamp to the current‐clamp (CC) configuration of the patch‐clamp technique. Our experiments revealed that Ca2+ entry through the nicotinic receptor at hyperpolarized membrane potentials contributed as much to the exocytosis (100.4 ± 27.3 fF) evoked by 200 ms pulses of ACh, as Ca2+ flux through voltage‐dependent Ca2+ channels at depolarized membrane potentials. The nicotinic current triggered a depolarization event with a peak at +49.3 mV and a ‘plateau’ phase that ended at −23.9 mV, which was blocked by 10 μmol/L mecamylamine. When a long ACh stimulus (15 s) was applied, the nicotinic current at the end of the pulse reached a value of 15.45 ± 3.6 pA, but the membrane potential depolarization still remained at the ‘plateau’ stage until withdrawal of the agonist. Perfusion with 200 μmol/L Cd2+ during the 15 s ACh pulse completely abolished the plasma membrane depolarization at the end of the pulse, indicating that Ca2+ entry through Ca2+ channels contributed to the membrane potential depolarization provoked by prolonged ACh pulses. These findings also reflect that voltage‐dependent Ca2+ channels were recruited by the small current flowing through the desensitized nicotinic receptor to maintain the depolarization. Finally, muscarinic receptor activation triggered a delayed exocytotic process after prolonged ACh stimulation, dependent on Ca2+ mobilization from the endoplasmic reticulum. In summary, we show here that nicotinic and muscarinic receptors contribute to the exocytosis of neurotransmitters in human chromaffin cells, and that the nicotinic receptor plays a key role in several stages of the stimulus–secretion coupling process in these cells.
Functional protein kinase arrays reveal inhibition of p‐21‐activated kinase 4 by α‐synuclein oligomers
Authors: Danzer Karin M. , Schnack Cathrin , Sutcliffe Andrew , Hengerer Bastian , Gillardon Frank
Published: September 2007

There is increasing evidence that aggregation of α‐synuclein contributes to the functional and structural deterioration in the CNS of Parkinson’s disease patients and transgenic animal models. α‐Synuclein binds to various cellular proteins and aggregated α‐synuclein species may affect their physiological function. In the present study, we used protein arrays spotted with 178 active human kinases for a large‐scale analysis of the effects of recombinant α‐synuclein on kinase activities. Incubation with globular α‐synuclein oligomers significantly inhibited autophosphorylation of p21‐activated kinase (PAK4) compared to treatment with monomeric α‐synuclein or β‐synuclein. A concentration‐dependent inhibition was also observed in a solution‐based kinase assay. To show in vivo relevance, we analyzed brainstem protein extracts from α‐synuclein (A30P) transgenic mice where accumulation of α‐synuclein oligomers has been demonstrated. By immunoblotting using a phospho‐specific antibody, we detected a significant decline in phosphorylation of LIM kinase 1, a physiological substrate for PAK4. Suppression of PAK activity by amyloid‐β oligomers has been reported in Alzheimer’s disease. Thus, PAKs may represent a target for various neurotoxic protein oligomers, and signaling deficits may contribute to the behavioral defects in chronic neurodegenerative diseases.
Hierarchical recruitment by AMPA but not staurosporine of pro‐apoptotic mitochondrial signaling in cultured cortical neurons: evidence for caspase‐dependent/independent cross‐talk
Authors: Beart Philip M. , Lim Maria L. R. , Chen Baohong , Diwakarla Shanti , Mercer Linda D. , Cheung Nam Sang , Nagley Phillip
Published: September 2007

Excitotoxicity mediated via the (S)‐α‐amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionate (AMPA) subtype of receptor for l‐glutamate contributes to various neuropathologies involving acute brain injury and chronic degenerative disorders. In this study, AMPA‐induced neuronal injury and staurosporine (STS)‐mediated apoptosis were compared in primary neuronal cultures of murine cerebral cortex by analyzing indices up‐ and downstream of mitochondrial activation. AMPA‐mediated apoptosis involved induction of Bax, loss of mitochondrial transmembrane potential (ΔΨm), early release of cytochrome c (cyt c), and more delayed release of second mitochondrial activator of caspases (SMAC), Omi, and apoptosis‐inducing factor (AIF) with early calpain and minor late activation of caspase 3. STS‐induced apoptosis was characterized by a number of differences, a more rapid time course, non‐involvement of ΔΨm, and relatively early recruitment of SMAC and caspase 3. The AMPA‐induced rise in intracellular calcium appeared insufficient to evoke ΔΨm as release of cyt c preceded mitochondrial depolarization, which was followed by the cytosolic translocation of SMAC, Omi, and AIF. Bax translocation preceded cyt c release for both stimuli inferring its involvement in apoptotic induction. Inclusion of the broad spectrum caspase inhibitor zVAD‐fmk reduced the AMPA‐induced release of cyt c, SMAC, and AIF, while only affecting the redistribution of Omi and AIF in the STS‐treated neurons. Only AIF release was affected by a calpain inhibitor (calpastatin) which exerted relatively minor effects on the progression of cellular injury. AMPA‐mediated release of apoptogenic proteins was more hierarchical relative to STS with its calpain activation and caspase‐dependent AIF redistribution arguing for a model with cross‐talk between caspase‐dependent/independent apoptosis.
Excitotoxic loss of post‐synaptic sites is distinct temporally and mechanistically from neuronal death
Authors: Waataja Jonathan J. , Kim Hee Jung , Roloff Alan M. , Thayer Stanley A.
Published: September 2007

Dendritic degeneration and loss of synaptic proteins are early events correlated with functional decline in neurodegenerative disease. The temporal and mechanistic relationship between synapse loss and cell death, however, remains unclear. We used confocal microscopy and image processing to count post‐synaptic sites on rat hippocampal neurons by expressing post‐synaptic density protein 95 fused to green fluorescent protein. Fluorescent puncta co‐localized with neurotransmitter release sites, NMDA‐induced Ca2+ increases and NMDA receptor immunoreactivity. During excitotoxic neurodegeneration, synaptic sites were lost and synaptic transmission impaired. These changes were mediated by NMDA receptors and required Ca2+‐dependent activation of the proteasome pathway. Tracking synapses from the same cell following brief neurotoxic insult revealed transient loss followed by recovery. The time‐course, concentration‐dependence and mechanism for loss of post‐synaptic sites were distinct from those leading to cell death. Cells expressing p14ARF, which inhibits ubiquitination of post‐synaptic density protein 95 and prevents loss of synaptic sites, displayed an increased sensitivity to glutamate‐induced cell death. Thus, excitotoxic synapse loss may be a disease‐modifying process rather than an obligatory step leading to cell death. These results demonstrate the importance of assessing synaptic function independent of neuronal survival during neurodegeneration and indicate that this approach will be useful for identifying toxins that degrade synaptic connections and for screening for agents that protect synaptic function.
Seizure‐induced formation of isofurans: novel products of lipid peroxidation whose formation is positively modulated by oxygen tension
Authors: Patel Manisha , Liang Li‐Ping , Hou Huagang , Williams Benjamin B. , Kmiec Maciej , Swartz Harold M. , Fessel Joshua P. , Roberts L. Jackson
Published: September 2007

We have previously shown that seizures induce the formation of F2‐isoprostanes (F2‐IsoPs), one of the most reliable indices of oxidative stress in vivo. Isofurans (IsoFs) are novel products of lipid peroxidation whose formation is favored by high oxygen tensions. In contrast, high oxygen tensions suppress the formation of F2‐IsoPs. The present study determined seizure‐induced formation of IsoFs and its relationship with cellular oxygen levels (pO2). Status epilepticus (SE) resulted in F2‐IsoP and IsoF formation, with overlapping but distinct time courses in hippocampal subregions. IsoF, but not F2‐IsoP formation coincided with mitochondrial oxidative stress. SE resulted in a transient decrease in hippocampal pO2 measured by in vivo electron paramagnetic resonance oximetry suggesting an early phase of seizure‐induced hypoxia. Seizure‐induced F2‐IsoP formation coincided with the peak hypoxia phase, whereas IsoF formation coincided with the ‘reoxygenation’ phase. These results demonstrate seizure‐induced increase in IsoF formation and its correlation with changes in hippocampal pO2 and mitochondrial dysfunction.
GSK3β mediates the induced expression of synaptic acetylcholinesterase during apoptosis
Authors: Jing Peng , Jin Qihuang , Wu Jun , Zhang Xue‐Jun
Published: September 2007

Besides its role in terminating acetylcholine‐mediated neurotransmission, acetylcholinesterase (AChE) is found to be expressed and participate in the process of apoptosis in various cell types. However, the mechanisms underlying AChE up‐regulation in neuronal cells remain elusive. Herein we demonstrated that glycogen synthase kinase‐3β (GSK3β) mediates induced AChE‐S expression during apoptosis. In this study, A23187 and thapsigargin (TG) were employed to induce apoptosis in neuroendocrine PC12 cells. The results showed that exposure of PC12 cells to A23187 and TG up‐regulated AChE activity significantly. The same treatment also led to activation of GSK3β. Two different inhibitors of GSK3β (lithium and GSK3β‐specific inhibitor VIII) could block A23187‐ or TG‐induced up‐regulation of AChE activity, AChE‐S mRNA level and protein expression. However, lithium could not inhibit the induction of AChE‐R mRNA and protein under similar conditions. Taken together, our results show that GSK3β is specifically involved in the induction of AChE‐S expression in PC12 cells during apoptosis.
Iron trafficking inside the brain
Authors: Moos Torben , Nielsen Thomas Rosengren , Skjørringe Tina , Morgan Evan H.
Published: September 2007

Iron, an essential element for all cells of the body, including those of the brain, is transported bound to transferrin in the blood and the general extracellular fluid of the body. The demonstration of transferrin receptors on brain capillary endothelial cells (BCECs) more than 20 years ago provided the evidence for the now accepted view that the first step in blood to brain transport of iron is receptor‐mediated endocytosis of transferrin. Subsequent steps are less clear. However, recent investigations which form the basis of this review have shed some light on them and also indicate possible fruitful avenues for future research. They provide new evidence on how iron is released from transferrin on the abluminal surface of BCECs, including the role of astrocytes in this process, how iron is transported in brain extracellular fluid, and how iron is taken up by neurons and glial cells. We propose that the divalent metal transporter 1 is not involved in iron transport through the BCECs. Instead, iron is probably released from transferrin on the abluminal surface of these cells by the action of citrate and ATP that are released by astrocytes, which form a very close relationship with BCECs. Complexes of iron with citrate and ATP can then circulate in brain extracellular fluid and may be taken up in these low‐molecular weight forms by all types of brain cells or be bound by transferrin and taken up by cells which express transferrin receptors. Some iron most likely also circulates bound to transferrin, as neurons contain both transferrin receptors and divalent metal transporter 1 and can take up transferrin‐bound iron. The most likely source for transferrin in the brain interstitium derives from diffusion from the ventricles. Neurons express the iron exporting carrier, ferroportin, which probably allows them to excrete unneeded iron. Astrocytes lack transferrin receptors. Their source of iron is probably that released from transferrin on the abluminal surface of BCECs. They probably to export iron by a mechanism involving a membrane‐bound form of the ferroxidase, ceruloplasmin. Oligodendrocytes also lack transferrin receptors. They probably take up non‐transferrin bound iron that gets incorporated in newly synthesized transferrin, which may play an important role for intracellular iron transport.
Localization of the membrane‐anchored MMP‐regulator RECK at the neuromuscular junctions
Authors: Kawashima Satoshi , Imamura Yukio , Chandana Ediriweera P. S. , Noda Toru , Takahashi Rei , Adachi Eijiro , Takahashi Chiaki , Noda Makoto
Published: September 2007

Nerve apposition on nicotinic acetylcholine receptor clusters and invagination of the post‐synaptic membrane (i.e. secondary fold formation) occur by embryonic day 18.5 at the neuromuscular junctions (NMJs) in mouse skeletal muscles. Finding the molecules expressed at the NMJ at this stage of development may help elucidating how the strong linkage between a nerve terminal and a muscle fiber is established. Immunohistochemical analyses indicated that the membrane‐anchored matrix metalloproteinase regulator RECK was enriched at the NMJ in adult skeletal muscles. Confocal and electron microscopy revealed the localization of RECK immunoreactivity in secondary folds and subsynaptic intracellular compartments in muscles. Time course studies indicated that RECK immunoreactivity becomes associated with the NMJ in the diaphragm at around embryonic day 18.5 and thereafter. These findings, together with known properties of RECK, support the hypothesis that RECK participates in NMJ formation and/or maintenance, possibly by protecting extracellular components, such as synaptic basal laminae, from proteolytic degradation.
Kir4.1 channels regulate swelling of astroglial processes in experimental spinal cord edema
Authors: Dibaj Payam , Kaiser Melanie , Hirrlinger Johannes , Kirchhoff Frank , Neusch Clemens
Published: September 2007

In glial cells, inwardly rectifying K+ channels (Kir) control extracellular [K+]o homeostasis by uptake of K+ from the extracellular space and release of K+ into the microvasculature. Kir channels were also recently implicated in K+‐associated water influx and cell swelling. We studied the time‐dependent expression and functional implication of the glial Kir4.1 channel for astroglial swelling in a spinal cord edema model. In this CNS region, Kir4.1 is expressed on astrocytes from the second postnatal week on and co‐localizes with aquaporin 4 (AQP4). Swelling of individual astrocytes in response to osmotic stress and to pharmacological Kir blockade were analyzed by time‐lapse‐two‐photon laser‐scanning microscopy in situ. Application of 30% hypotonic solution induced astroglial soma swelling whereas no swelling was observed on astroglial processes or endfeet. Co‐application of hypotonic solution and Ba2+, a Kir channel blocker, induced prominent swelling of astroglial processes. In Kir4.1−/− mice, however, somatic as well as process swelling was observed upon application of 30% hypotonic solutions. No additional effect was provoked upon co‐application with Ba2+. Our experiments show that Kir channels prevent glial process swelling under osmotic stress. The underlying Kir channel subunit that controls glial process swelling is Kir4.1, whereas changes of the glial soma are not substantially related to Kir4.1.
The α1 subunit of nicotinic acetylcholine receptors in the inner ear: transcriptional regulation by ATOH1 and co‐expression with the γ subunit in hair cells
Authors: Scheffer Deborah , Sage Cyrille , Plazas Paola V. , Huang Mingqian , Wedemeyer Carolina , Zhang Duan‐Sun , Chen Zheng‐Yi , Elgoyhen A. Belen , Corey David P. , Pingault Veronique
Published: September 2007

Acetylcholine is a key neurotransmitter of the inner ear efferent system. In this study, we identify two novel nAChR subunits in the inner ear: α1 and γ, encoded by Chrna1 and Chrng, respectively. In situ hybridization shows that the messages of these two subunits are present in vestibular and cochlear hair cells during early development. Chrna1 and Chrng expression begin at embryonic stage E13.5 in the vestibular system and E17.5 in the organ of Corti. Chrna1 message continues through P7, whereas Chrng is undetectable at post‐natal stage P6. The α1 and γ subunits are known as muscle‐type nAChR subunits and are surprisingly expressed in hair cells which are sensory‐neural cells. We also show that ATOH1/MATH1, a transcription factor essential for hair cell development, directly activates CHRNA1 transcription. Electrophoretic mobility‐shift assays and supershift assays showed that ATOH1/E47 heterodimers selectively bind on two E boxes located in the proximal promoter of CHRNA1. Thus, Chrna1 could be the first transcriptional target of ATOH1 in the inner ear. Co‐expression in Xenopus oocytes of the α1 subunit does not change the electrophysiological properties of the α9α10 receptor. We suggest that hair cells transiently express α1γ‐containing nAChRs in addition to α9α10, and that these may have a role during development of the inner ear innervation.
The role of the cannabinoid type 1 receptor and down‐stream cAMP/DARPP‐32 signal in the nucleus accumbens of methamphetamine‐sensitized rats
Authors: Chiang Yao‐Chang , Chen Jin‐Chung
Published: September 2007

Blockade of the cannabinoid type 1 (CB1) receptor could suppress methamphetamine self‐administration; however, the cellular mechanism remains unclear. In this study, we intended to investigate the significance of brain CB1receptors on the development of behavioral sensitization to methamphetamine. Male Sprague‐Dawley rats treated with chronic methamphetamine (4 mg/kg, i.p.) for either 7 or 14 days developed behavioral sensitization to methamphetamine (1 mg/kg) at withdrawal day 7. A progressive decrease in numbers of CB1receptor (bothBmaxand mRNA) but increase in binding affinity (Kd) was noticed during withdrawal days 3 to 7. Microinjection of CB1antagonist [5‐(4‐bromophenyl)‐1‐(2,4‐dichlorophenyl)‐4‐ethyl‐N‐(1‐piperidinyl)‐1H‐pyrazole‐3‐carboxamide] into the nucleus accumbens (NAc) at withdrawal day 7, significantly suppressed the behavioral sensitization to methamphetamine. In NAc brain slices preparation, acute incubation with CB1agonist (1R,3R,4R)‐3‐[2‐hydroxy‐4‐(1,1‐dimethylheptyl)phenyl]‐4‐(3‐hydroxypropyl)cyclohexan‐1‐ol (CP 55940) dose‐dependently enhanced cAMP accumulation in sensitized rats; no change was noticed in control groups. Consequently, treatment of CP 55940 induced a dose‐dependent (10 nmol/L–10 μmol/L) phosphorylation on down‐stream dopamine and cAMP‐regulated phosphoprotein of Mr 32 000 (DARPP‐32)/Thr34 in sensitized rats, while only 10 μmol/L CP 55940 was able to enhance the phosphoDARPP‐32/T34 in control groups. Alternatively, both basal activity of calcineurin (PP‐2B) and CP 55940‐induced changes in the amount of PP‐2B in the NAc were both decreased in sensitized rats, but not in controls. Overall, we demonstrated that brain CB1receptor and its down‐stream cAMP/DARPP‐32/T34/PP‐2B signaling are profoundly altered in methamphetamine‐sensitized animals.
Replacement of histidine in position 105 in the α5subunit by cysteine stimulates zolpidem sensitivity of α5β2γ2GABAAreceptors
Authors: Baur Roland , Sigel Erwin
Published: September 2007

Zolpidem is a positive allosteric modulator of GABAAreceptors with sensitivity to subunit composition. While it acts with high affinity and efficacy at GABAAreceptors containing the α1subunit, it has a lower affinity to GABAAreceptors containing α2, α3, or α5subunits and has a very weak efficacy at receptors containing the α5subunit. Here, we show that replacing histidine in position 105 in the α5subunit by cysteine strongly stimulates the effect of zolpidem in receptors containing the α5subunit. The side chain volume of the amino acid residue in this position does not correlate with the modulation by zolpidem. Interestingly, serine is not able to promote the potentiation by zolpidem. The homologous residues to α5H105 in α1, α2, and α3are well‐known determinants of the action of classical benzodiazepines. Other studies have shown that replacement of these histidines α1H101, α2H101, and α3H126 by arginine, as naturally present in α4and α6, leads to benzodiazepine insensitivity of these receptors. Thus, the nature of the amino acid residue in this position is not only crucial for the action of classical benzodiazepines but in α5containing receptors also for the action of zolpidem.
An exposed carboxyl group on sialic acid is essential for gangliosides to inhibit calcium uptake via the sarco/endoplasmic reticulum Ca2+‐ATPase: relevance to gangliosidoses
Authors: Ginzburg Luba , Li Su‐Chen , Li Yu‐Teh , Futerman Anthony H.
Published: September 2007

We previously observed that gangliosides GM2, GM1, and GM3 inhibit Ca2+‐uptake via the sarco/endoplasmic reticulum Ca2+‐ATPase (SERCA) in neurons and in brain microsomes. We now systematically examine the effect of various gangliosides and their analogs on Ca2+‐uptake via SERCA and demonstrate that an exposed carboxyl group on the ganglioside sialic acid residue is required for inhibition. Thus, asialo‐GM2 and asialo‐GM1 have no inhibitory effect, and modifications of the carboxyl group of GM1 and GM2 into a hydroxymethyl residue (CH2OH), a methyl ester (COOCH3) or a taurine‐conjugated amide (CONHCH2CH2SO3H) drastically diminish their inhibitory activities. We also demonstrate that the saccharides must be attached to a ceramide backbone in order to inhibit SERCA as the ceramide‐free ganglioside saccharides only inhibit SERCA to a minimal extent. Finally, we attempted to use the ceramide‐free ganglioside saccharides to antagonize the effects of the gangliosides on SERCA; although some reversal was observed, the inhibitory effects of the gangliosides were not completely abolished.
Up‐regulation of L‐type high voltage‐gated calcium channel subunits by sustained exposure to 1,4‐ and 1,5‐benzodiazepines in cerebrocortical neurons
Authors: Katsura Masashi , Shibasaki Masahiro , Kurokawa Kazuhiro , Tsujimura Atsushi , Ohkuma Seitaro
Published: September 2007

The aim of this study is to examine how sustained exposure to two 1,4‐benzodiazepines (BZDs) with different action period, diazepam and brotizolam, and a 1,5‐BZD, clobazam, affects L‐type high voltage‐gated calcium channel (HVCC) functions and its mechanisms using primary cultures of mouse cerebral cortical neurons. The sustained exposure to these three BZDs increased [45Ca2+] influx, which was due to the enhanced [45Ca2+] entry through L‐type HVCCs but not through of Cav2.1 and Cav2.2. Increase in [3H]diltiazem binding after the exposure to these three BZDs was due to the increase in the binding sites of [3H]diltiazem. Western blot analysis showed increase of Cav1.2 and Cav1.3 in association with the increased expression of α2/δ1 subunit. Similar changes in [3H]diltiazem binding and L‐type HVCC subunit expression were found in the cerebral cortex from mouse with BZD physical dependence. These results indicate that BZDs examined here have the potential to increase L‐type HVCC functions mediated via the enhanced expression of not only Cav1.2 and Cav1.3 but also α2/δ1 subunit after their sustained exposure, which may participate in the development of physical dependence by these BZDs.
Cerebrospinal fluid concentrations of peptides derived from chromogranin B and secretogranin II are decreased in multiple sclerosis
Authors: Mattsson Niklas , Rüetschi Ulla , Podust Vladimir N. , Stridsberg Mats , Li Susann , Andersen Oluf , Haghighi Sara , Blennow Kaj , Zetterberg Henrik
Published: September 2007

Novel biomarkers for multiple sclerosis (MS) could improve diagnosis and provide clues to pathogenesis. In this study surface‐enhanced laser desorption/ionization time‐of‐flight mass spectrometry was used to analyze protein expression in CSF from 46 MS patients, 46 healthy siblings to the patients, and 50 unrelated healthy controls. Twenty‐four proteins in the mass range 2–10 kDa were expressed at significantly different levels (p < 0.01) in a robust manner when comparing the three groups. Identities of three proteins were determined using biochemical purification followed by tandem mass spectrometric analysis. Immunoprecipitation experiments confirmed the identities for two peptides derived from chromogranin B (m/z 6252) and from secretogranin II (m/z 3679). These peptides were all decreased in MS when compared with siblings or controls. Radioimmunoassays specific for each peptide confirmed these differences. The lowered concentrations did not correlate to the axonal damage marker neurofilament light protein and may thus reflect functional changes rather than neurodegeneration. Further studies will investigate the involvement of these peptides in MS pathogenesis.
Nociceptive and behavioural sensitisation by protein kinase Cɛ signalling in the CNS
Authors: Van Kolen Kristof , Pullan Shirley , Neefs Jean‐Marc , Dautzenberg Frank M.
Published: September 2007

Despite the apparent homology in the protein kinase C (PKC) family, it has become clear that slight structural differences are sufficient to have unique signalling properties for each individual isoform. For PKCɛ in depth investigation of these aspects revealed unique actions in the CNS and lead to development of specific modulators with clinical perspective. In this review, we describe to which extent PKCɛ is distinct from other isoforms on the level of tissue expression and protein structure. As this kinase is highly expressed in the brain, we outline three main aspects of PKCɛ signalling in the CNS. First, its ability to alter the permeability of N‐type Ca2+channels in dorsal root ganglia has been shown to enhance nociception. Secondly, PKCɛ increases anxiety by diminishing GABAAR‐induced inhibitory post‐synaptic currents in the prefrontal cortex. Another important aspect of the latter inhibition is the reduced sensitivity of GABAAreceptors to ethanol, a mechanism potentially contributing to abuse. A third signalling cascade improves cognitive functions by facilitating cholinergic signalling in the hippocampus. Collectively, these findings point to a physical and behavioural sensitising role for this kinase.
Treadmill exercise counteracts the suppressive effects of peripheral lipopolysaccharide on hippocampal neurogenesis and learning and memory
Authors: Wu Chih‐Wei , Chen Yi‐Chieh , Yu Lung , Chen Hsiun‐ing , Jen Chauying J. , Huang A‐Min , Tsai Hsing‐Jung , Chang Ya‐Ting , Kuo Yu‐Min
Published: September 2007

New neurons are continuously generated in hippocampal subgranular zone throughout life, and the amount of neurogenesis is suggested to be correlated with the hippocampus‐dependent function. Several extrinsic stimuli are known to modulate the neurogenesis process. Among them, physical exercise has advantageous effects on neurogenesis and brain function, while inflammation shows the opposite. Herein we showed that a moderate running exercise successfully restored the peripheral lipopolysaccharide (LPS)‐impaired neurogenesis in the dentate area. LPS treatment obstructed neuronal differentiation, but not proliferation. Exercise training facilitated both the proliferation of the neural stem cells and their differentiation into neurons. Interestingly, exercise replenished the LPS‐reduced levels of brain‐derived neurotrophic factor and its receptor, TrkB, and rescued the LPS‐disturbed performance in water maze; while the LPS‐elicited up‐regulation of tumor necrosis factor‐alpha and interleukin‐1β remained unaltered. In conclusion, our findings suggest that running exercise effectively ameliorates the LPS‐disturbed hippocampal neurogenesis and learning and memory performance. Such advantageous effects of running exercise are not due to the alteration of inflammatory response, but possibly by the restoring the LPS‐lessened brain‐derived neurotrophic factor signaling pathway.
Soluble aggregates of the amyloid‐β protein activate endothelial monolayers for adhesion and subsequent transmigration of monocyte cells
Authors: Gonzalez‐Velasquez Francisco J. , Moss Melissa A.
Published: September 2007

Increasing evidence suggests that the deposition of amyloid plaques, composed primarily of the amyloid‐β protein (Aβ), within the cerebrovasculature is a frequent occurrence in Alzheimer’s disease and may play a significant role in disease progression. Accordingly, the pathogenic mechanisms by which Aβ can alter vascular function may have therapeutic implications. Despite observations that Aβ elicits a number of physiological responses in endothelial cells, ranging from alteration of protein expression to cell death, the Aβ species accountable for these responses remains unexplored. In the current study, we show that isolated soluble Aβ aggregation intermediates activate human brain microvascular endothelial cells for both adhesion and subsequent transmigration of monocyte cells in the absence of endothelial cell death and monolayer disruption. In contrast, unaggregated Aβ monomer and mature Aβ fibril fail to induce any change in endothelial adhesion or transmigration. Correlations between average Aβ aggregate size and observed increases in adhesion illustrate that smaller soluble aggregates are more potent activators of endothelium. These results support previous studies demonstrating heightened neuronal activity of soluble Aβ aggregates, including Aβ‐derived diffusible ligands, oligomers, and protofibrils, and further show that soluble aggregates also selectively exhibit activity in a vascular cell model.
PERMANENT GENETIC RESOURCES: Development of microsatellite markers and analysis of their inheritance in the Australian reptile tick, Bothriocroton hydrosauri
Authors: GUZINSKI JARO , SAINT KATHLEEN M. , GARDNER MICHAEL G. , DONNELLAN STEPHEN C. , BULL C. MICHAEL
Published: September 2007

Despite long‐term study, the mechanism explaining the parapatric distribution of two Australian reptile tick species is not understood. We describe the development of primers amplifying 10 microsatellite Bothriocroton hydrosauri loci, for the study of population structure and dispersal patterns of this tick. The numbers of alleles per locus ranged from two to seven in ticks from the study site, and the observed heterozygosity between 0.28 and 0.69. Pedigree analysis indicates that one locus is inherited in a non‐Mendelian manner in three families, which was not explained by null allele presence.
PERMANENT GENETIC RESOURCES: Consensus primers of cyp73 genes discriminate willow species and hybrids (Salix, Salicaceae)
Authors: TRUNG LE QUANG , VAN PUYVELDE KAROLIEN , TRIEST LUDWIG
Published: September 2007

Consensus primers, based on exon sequences of the cyp73 gene family coding for cinnamate 4‐hydroxylase (C4H) of the lignin biosynthesis pathway, were designed for the tetraploid willow species Salix alba and Salix fragilis. Diagnostic alleles at species level were observed among introns of three cyp73 genes and allowed unambiguous detection of the first generation and introgressed hybrids in populations. Progeny analysis of a female S. alba with a male introgressed hybrid confirmed the codominant inheritance of each intron. Sequences of the diagnostic alleles of both species were similar to those found in the hybrids.
DNA BARCODING: Barcoding corals: limited by interspecific divergence, not intraspecific variation
Authors: SHEARER T. L. , COFFROTH M. A.
Published: September 2007

The expanding use of DNA barcoding as a tool to identify species and assess biodiversity has recently attracted much attention. An attractive aspect of a barcoding method to identify scleractinian species is that it can be utilized on any life stage (larva, juvenile or adult) and is not influenced by phenotypic plasticity unlike morphological methods of species identification. It has been unclear whether the standard DNA barcoding system, based on cytochromecoxidase subunit 1 (COI), is suitable for species identification of scleractinian corals. Levels of intra‐ and interspecific genetic variation of the scleractinian COI gene were investigated to determine whether threshold values could be implemented to discriminate conspecifics from other taxa. Overlap between intraspecific variation and interspecific divergence due to low genetic divergence among species (0% in many cases), rather than high levels of intraspecific variation, resulted in the inability to establish appropriate threshold values specific for scleractinians; thus, it was impossible to discern most scleractinian species using this gene.