bluuebunny/crossref_metadata_2025_split huggingface dataset:
Perturbation of the synaptic release machinery in hippocampal neurons by overexpression of SNAP‐25 with the Semliki Forest virus vector
Authors: Owe‐Larsson Björn , Berglund , Magnus M. , Kristensson Krister , Garoff , Henrik , Larhammar Dan , Brodin Lennart , Löw Peter 
Published: March 2003

AbstractWe have examined whether the Semliki Forest virus (SFV) expression vector can be used to manipulate the exocytotic machinery in cultured hippocampal neurons. Autaptic responses were recorded in individually identified neurons which overexpressed either a non‐synaptic protein, the transferrin receptor, or the synaptic SNARE protein SNAP‐25 (synaptosomal‐associated protein of 25 kDA). In neurons overexpressing the transferrin receptor, autaptic responses occurred in a similar proportion and had similar amplitudes (12–18 h postinfection) as in uninfected control neurons. With increasing time after the infection, an increasing proportion of the transferrin receptor‐overexpressing neurons showed changes in the shape of the cell body, but the autaptic responses appeared normal as long as recordings could be performed (up to 30 h postinfection). In contrast, in SNAP‐25‐overexpressing neurons, the proportion of responding cells was reduced 12–18 h after the infection, and the amplitude of the autaptic current in responding neurons was also reduced. The sensitivity to exogenously applied glutamate was, however, unchanged. Biochemical analysis showed that 50% of the overexpressed SNAP‐25 was palmitoylated. The levels of two other SNAREs, syntaxin and synaptobrevin (also called vesicle‐associated membrane protein), were not affected. Our results indicate that the SFV vector can provide an effective tool to study the function of proteins participating in neurotransmitter release.
Heparan sulphates upregulate regeneration of transected sciatic nerves of adult guinea‐pigs
Authors: Chau C. H. , Shum D. K. Y. , Chan Y. S. , So K. ‐F. 
Published: March 2003

AbstractThe increased content of soluble glycosaminoglycan‐containing forms in sciatic nerves during recovery from crush injury [Shum & Chau (1996) J. Neurosci. Res., 46, 465] suggests that the glycosaminoglycans modulate the environment for post‐traumatic tissue remodelling and axonal regrowth. To test this, defined amounts of soluble heparan sulphates from bovine kidney or guinea‐pig nerve were introduced into the regenerating environment via silicone conduits that bridged 8‐mm gaps of transected sciatic nerves of adult guinea‐pigs. Controls were bridged using the phosphate‐buffered saline (PBS) vehicle or a chondroition sulphate preparation from whale cartilage. After timed periods of recovery, the animals were assessed for electromyographic signals at the target gastrocnemius muscle to determine the conduction velocity across the bridged nerve. Sections of the bridge were also histologically examined for nerve fibres. Transected sciatic nerves bridged with heparan sulphates or chondroitin sulphate showed earlier stimulated myelination of axons (week 5–6) than PBS‐bridged nerves (week 9). Initial electromyographic indication of reconnection with the target was at week 9 post‐transection. In the course of 20 weeks, transected sections of the bridge indicated similar numbers of unmyelinated axons irrespective of bridge material, but distinctly higher numbers of myelinated axons in heparan sulphate‐bridged nerves than either PBS‐ or chondroitin sulphate‐bridged nerves. At the end of the same period, heparan sulphate‐bridged nerves resumed normal conduction velocities, but both PBS‐ and chondroitin sulphate‐bridged nerves remained at 50% of that of the intact contralateral nerves. These results are the first to demonstrate that supplementation of soluble heparan sulphate to the fluid regenerative neural environment can restore functional, axonal reconnection of the severed nerve with the target muscle.
Real‐time imaging of glucocorticoid receptor dynamics in living neurons and glial cells in comparison with non‐neural cells
Authors: Nishi Mayumi , Takenaka Nobuyuki , Morita Noriyuki , Ito Takao , Ozawa Hitoshi , Kawata Mitsuhiro 
Published: March 2003

AbstractTo investigate the intracellular trafficking of glucocorticoid receptor (GR) in response to various conditions in a single living cell, a green fluorescent protein (GFP) and rat GR chimera construct (GFP‐GR) was prepared. We transiently transfected GFP‐GR into primary cultured rat hippocampal neurons, cortical glial cells, and non‐neural cells, e.g. COS‐1 cells and CV‐1 cells, and compared the dynamic changes in subcellular localization of GFP‐GR in these cells. When GFP‐GR was expressed in the cells, GFP‐GR efficiently transactivated the mouse mammary tumour virus promoter in response to dexamethasone (DEX). The cytoplasm‐to‐nuclear translocation of GFP‐GR induced with 10–7 m DEX, a specific agonist of GR, at 37 °C was completed within 30 min in all cell types used, and the rate of nuclear translocation was dependent on the ligand dose. The translocation of GFP‐GR into the nucleus from the cytoplasm was induced in a ligand‐specific manner, similar to that of the native GR. The disruption of microtubules by colchicine or nocodazole showed no significant effect on the DEX‐induced GFP‐GR translocation from the cytoplasmic region to the nuclear region. The cells were not deteriorated during time‐lapse imaging analysis for 1 h at 37 °C. The present findings suggest that the subcellular localization of GFP‐GR is dynamically changed in response to extracellular and intracellular conditions, and that there are no conspicuous variations in the manner of trafficking of GR among different types of cells in vitro.
Transcriptional repression by the zinc finger protein REST is mediated by titratable nuclear factors
Authors: Leichter Michael , Thiel Gerald 
Published: March 2003

AbstractThe zinc finger protein REST (RE‐1 silencing transcription factor) is a transcriptional repressor that inhibits neuronal gene transcription in non‐neuronal tissues. REST may represent a master regulator of neuronal gene expression. REST contains two repressor domains located at the N‐ and C‐termini of the molecule. To investigate the molecular mechanism of transcriptional repression by REST, in vivo competition experiments were performed. Both repression domains were expressed in the nucleus as fusion proteins with S. japonicum glutathione S‐transferase (GST). The ability of these fusion proteins to block transcriptional repression mediated by the repressor domains of REST was tested. The results show that transcriptional repression by the N‐terminal repression domain of REST could be overcome by expression of a GST fusion protein encoding the N‐terminal, but not C‐terminal repression domain, and vice versa, suggesting that both repression domains have to interact with distinct nuclear factors to exhibit biological activity. The GST‐REST fusion proteins had no effect upon transcriptional repression mediated by the KRAB (Krüppel‐associated box) domain, a strong mammalian repressor domain, or the repressor domain derived from the thyroid hormone receptors α. We conclude that REST has to interact with at least two distinct nuclear factors to inhibit transcription. These factors are distinct from the mammalian corepressor proteins KAP‐1/KRIP‐1 and N‐CoR that mediate repression by the KRAB domain or the thryoid hormone receptor α. Thus, mammalian transcriptional repressors utilize different mechanisms to inhibit transcription by using different kinds of protein–protein interactions.
K+‐dependence of Na+–Ca2+ exchange in type I vestibular sensory cells of guinea‐pig
Authors: Boyer C. , Sans A. , Vautrin J. , Chabbert C. , Lehouelleur J. 
Published: March 2003

AbstractThe properties of the vestibular Na+–Ca2+ exchanger in mammalian type I vestibular sensory cells were studied using fura‐2 fluorescence and immunocytochemical techniques. In the absence of external Na+, the activation of Na+–Ca2+ exchange in reverse mode required the presence of external K+ (K+o) and depended on K+o concentration. Alkali cations Rb+ and NH4+ but not Li+ or Cs+ substituted for K+o to activate the exchange. For pressure applications of 10 mm K+, the contribution of voltage‐sensitive calcium channels to the increase in [Ca2+]i was < 15%. The dependence of the exchange on [K+]o was also recorded when the membrane potential was clamped using carbonyl cyanide p‐trifluoromethoxy‐phenylhydrazone (FCCP) and monensin ionophores. In these conditions, where there was no intracellular Na+, the increase in [Ca2+]i was completely blocked. These physiological results suggest that in reverse mode, Ca2+ entry is driven by both an outward transport of Na+ and an inward transport of K+. The dependence of the vestibular Na+–Ca2+ exchanger on K+ is more reminiscent of the properties of the retinal type Na+–Ca2+ exchanger than those of the more widely distributed cardiac type exchanger. Moreover, the immunocytochemical localization of both types of exchange proteins in the vestibular sensory epithelium confirmed the presence in the vestibular sensory cells of a Na+–Ca2+ exchanger which is recognized by an antibody raised against retinal type and not by an antibody raised against the cardiac type.
Control of neuronal excitability by an ion‐sensing receptor
Authors: Washburn David L. S. , Smith Pauline M. , Ferguson Alastair V. 
Published: March 2003

AbstractWhile most central nervous system (CNS) neurons receive the majority of their input through direct synaptic connections, there is evidence suggesting that they are in fact susceptible to modulation by changes in extracellular ionic composition during both physiological and pathophysiological conditions. In many regions of the CNS, there exists an identified extracellular receptor with the ability to sense levels of cations, most notably calcium. Here we report that activation of this calcium receptor (CaR) in neurons of the subfornical organ (SFO), a forebrain circumventricular structure, results in profound effects on neuronal excitability through metabotropic actions on a non‐selective cation channel. Activation of the CaR by NPS R‐467, an allosteric agonist of the CaR, evoked depolarizing plateau potentials ranging in duration from 5 to 30 s. Similarly, 5 mm CaCl2 caused depolarization and increased action potential frequency. NPS R‐467 was found to activate a non‐selective cation channel with a reversal potential of −48 ± 4 mV, and a slope conductance of 2.54 ± 11 nS. This current could also be elicted by spermine, a known agonist of the CaR. CaR‐mediated activation of this channel was dependent upon both G proteins and intracellular Ca2+ signalling, as disruption of these pathways through inclusion of guanosine 5′‐O‐(2‐thiodiphosphate) (GDP‐β‐S) and 1,2‐bis(2‐aminophenoxy)ethane‐N,N,N ′,N ′‐tetraacetic acid (BAPTA), respectively, in the recording pipette prevented activation of the current. Microinjection of CaR agonists into the SFO of anaesthetized rats resulted in a significant, site‐specific elevation of blood pressure (mean area under curve, 141 ± 50 mmHg.s). Together, these results indicate that the CaR can play an important role in transducing the effects of changes in the extracellular ionic composition, and that these effects have implications for the neural control of fluid balance.
Visualization of local afferent inputs to magnocellular oxytocin neurons in vitro
Authors: Jourdain P. , Dupouy B. , Bonhomme R. , Poulain D. A. , Israel J. M. , Theodosis D. T. 
Published: March 2003

AbstractWe recently showed that oxytocin (OT) neurons in organotypic slice cultures obtained from postnatal rat hypothalamus display complex patterns of electrical activity, similar to those of adult magnocellular OT neurons in vivo. Here we used such cultures to investigate the identity and, in particular, the origin of afferent inputs responsible for this activity. Multiple immunostaining with light and confocal microscopy showed that the somata and dendrites of oxytocinergic neurons were contacted by numerous synapses, visualized by their reaction to the synaptic markers, synaptophysin or synapsin. Many were GABAergic, displaying immunoreactivities for glutamic acid decarboxylase or γ‐aminobutyric acid (GABA); others were enriched in glutamate immunoreactivity. Such afferents presumably arose from GABA‐ or glutamate‐immunoreactive neurons, respectively, with distinct and characteristic morphologies and topographies. A few dopaminergic boutons (tyrosine hydroxylase‐ or dopamine‐immunopositive) impinged on OT neurons; they arose from dopamine‐positive neurons located along the third ventricle. No noradrenergic profiles were detected. Despite the presence of choline acetyl‐transferase (ChAT)‐immunoreactive neurons, there were no cholinergic contacts. Lastly, we found oxytocinergic synapses, identified by immunoreaction for OT‐related neurophysin and synapsin, contacting OT somata and dendrites.Our observations thus demonstrate that inhibitory and excitatory inputs to OT neurons derive from local intrahypothalamic GABA and glutamate neurons, in close proximity to the neurons. They also reveal that OT neurons are innervated by hypothalamic dopaminergic neurons. Finally, they confirm the existence of homotypic OT synaptic contacts which derive from local OT neurons.
Social intelligence in the normal and autistic brain: an fMRI study
Authors: Baron‐Cohen Simon , Ring Howard A. , Wheelwright Sally , Bullmore Edward T. , Brammer Mick J. , Simmons Andrew , Williams Steve C. R. 
Published: March 2003

AbstractThere is increasing support for the existence of ‘social intelligence’[Humphrey (1984) Consciousness Regained], independent of general intelligence. Brothers et al. (1990) J. Cog. Neurosci., 4, 107–118] proposed a network of neural regions that comprise the ‘social brain’: the orbito‐frontal cortex (OFC), superior temporal gyrus (STG) and amygdala. We tested Brothers' theory by examining both normal subjects as well as patients with high‐functioning autism or Asperger syndrome (AS), who are well known to have deficits in social intelligence, and perhaps deficits in amygdala function [Bauman & Kemper (1988) J. Neuropath. Exp. Neurol., 47, 369]. We used a test of judging from the expressions of another person's eyes what that other person might be thinking or feeling. Using functional magnetic resonance imaging (fMRI) we confirmed Brothers' prediction that the STG and amygdala show increased activation when using social intelligence. Some areas of the prefrontal cortex also showed activation. In contrast, patients with autism or AS activated the fronto‐temporal regions but not the amygdala when making mentalistic inferences from the eyes. These results provide support for the social brain theory of normal function, and the amygdala theory of autism.
Localization of synapse‐associated proteins during postnatal development of the rat retina
Authors: Koulen Peter 
Published: March 2003

AbstractThe expression of synapse‐associated proteins (SAPs) was monitored throughout postnatal development of the rat retina using specific antibodies and immunocytochemistry. The distribution of chapsin‐110/postsynaptic density protein (PSD)‐93, SAP90/PSD‐95, SAP97 and SAP102 immunoreactivity was characterized. All SAPs were found to be expressed in the inner plexiform layer (IPL) from birth on or soon after birth. With the exception of SAP97, the IPL labelling changed from a diffuse pattern staining the whole developing IPL to the typical adult punctate synaptic staining in the second postnatal week. Staining in the outer retina was first observed at postnatal day 5 (P5) for all proteins at the onset of outer plexiform layer (OPL) development. All SAPs showed a differential cellular and temporal distribution being either exclusively pre‐ or postsynaptically localized. Except for SAP90/PSD‐95, immunoreactivity was also detected in the nerve fibre layer throughout postnatal development. Possible functions of the early expression of SAPs well before differentiation and maturation of glutamatergic ribbon synapses are discussed.
Neuroprotective actions of peripherally administered insulin‐like growth factor I in the injured olivo‐cerebellar pathway
Authors: Fernandez A. M. , de la Vega A. Gonzalez , Planas B. , Torres‐Aleman I. 
Published: March 2003

AbstractExogenous administration of insulin‐like growth factor I (IGF‐I) restores motor function in rats with neurotoxin‐induced cerebellar deafferentation. We first determined that endogenous IGFs are directly involved in the recovery process because infusion of an IGF‐I receptor antagonist into the lateral ventricle blocks gradual recovery of limb coordination that spontaneously occurs after partial deafferentation of the olivo‐cerebellar circuitry. We then analysed mechanisms whereby exogenous IGF‐I restores motor function in rats with complete damage of the olivo‐cerebellar pathway. Treatment with IGF‐I normalized several markers of cell function in the cerebellum, including calbindin, glutamate receptor 1 (GluR1), γ‐aminobutyric acid (GABA) and glutamate, which are all depressed after 3‐acetylpyridine (3AP)‐induced deafferentation. IGF‐I also promoted functional reinnervation of the cerebellar cortex by inferior olive (IO) axons. In the IO, increased expression of bax in neurons and bcl‐X in astrocytes after 3AP was significantly reduced by IGF‐I treatment. On the contrary, IGF‐I prevented the decrease in poly‐sialic‐acid neural cell adhesion molecule (PSA‐NCAM) and GAP‐43 expression induced by 3AP in IO cells. IGF‐I also significantly increased the number of neurons expressing bcl‐2 in brainstem areas surrounding the IO. Altogether, these results indicate that subcutaneous IGF‐I therapy promotes functional recovery of the olivo‐cerebellar pathway by acting at two sites within this circuitry: (i) by modulating death‐ and plasticity‐related proteins in IO neurons; and (ii) by impinging on homeostatic mechanisms leading to normalization of cell function in the cerebellum. These results provide insight into the neuroprotective actions of IGF‐I and may be of practical consequence in the design of new therapeutic approaches for neurodegenerative diseases.
Stimulation of μ‐ and δ‐opioid receptors enhances phosphoinositide metabolism in mouse spinal cord: evidence for subtypes of δ‐receptors
Authors: Sánchez‐Blázquez Pilar , Rodríguez‐Díaz Marta , Frejo María‐Teresa , Garzón Javier 
Published: March 2003

AbstractThe accumulation of inositol phosphates (IPs) induced by agonist‐activated opioid receptors was analysed in mouse spinal cord slices pre‐labelled with myo‐[3H]inositol. Agonists showing selectivity to μ‐opioid receptors, morphine and [d‐Ala2,MePhe4, Gly(ol)5]enkephalin (DAMGO), promoted concentration‐dependent increases in the formation of IPs. The activation of δ‐opioid receptors by the selective agonists [d‐Pen2,5]enkephalin (DPDPE) and [d‐Ala2]deltorphin II produced similar increases in phosphoinositide (PI) metabolism. Pre‐treatment of the slices with pertussis toxin (PTX) blocked the effect of opioid agonists on IP production. The involvement of Gi/Go‐protein (guanine nucleotide‐binding protein) classes in this opioid effect is therefore suggested. The activity of the opioid agonists was reduced by the opioid antagonists naltrexone and naloxone. The antagonist at δ1‐receptors, 7‐benzylidenenaltrexone (BNTX), exhibited greater potency than the antagonists at δ2‐receptors, naltriben methanesulphonate (NTB) or naltrindrole 5′‐isothiocyanate (NT II), in reducing the activating effect of DPDPE on phosphoinositide metabolism. Conversely, NTB and NT II were more potent antagonists of the activity of [d‐Ala2]deltorphin II than BNTX. This work demonstrates the coupling of spinal μ‐ and δ‐opioid receptors to phospholipase C and the generation of IPs. It also provides biochemical evidence for pharmacological subtypes of δ‐opioid receptors in the activation of this signalling pathway.
Involvement of substance P in the anti‐inflammatory effects of the peripherally selective κ‐opioid asimadoline and the NK1 antagonist GR205171
Authors: Binder Waltraud , Scott Caroline , Walker Judith S. 
Published: March 2003

AbstractWe have previously shown that kappa‐opioids have antiarthritic properties. In this study, using two differently acting drugs (the peripherally selective κ‐agonist, asimadoline, and the NK1‐antagonist, GR205171), we have examined possible roles of the neuropeptide substance P (SP) in the pathogenesis and maintenance of experimental arthritis in rats. The anti‐inflammatory actions and the time dependence of these drugs were compared, and concentrations of SP determined in joint tissue. In untreated animals, SP levels in ankle joint tissue increased late in the disease (by day 21) but substantially lagged behind development of clinical disease. Prolonged (days 1–21 or days 12–18) but not early, short‐term (days 1–3) treatment with the NK1‐antagonist GR205171 (1 mg/kg/day i.p.) significantly attenuated joint damage; SP levels showed multiphasic dose dependence over the 21‐day treatment. The data suggest that GR205171 antagonizes the action of SP by presynaptic as well as postsynaptic mechanisms. Treatment with asimadoline (5 mg/kg/day i.p.) produced marked (and sustained) attenuation of the disease with all three time regimes. The effect of asimadoline on SP levels was time dependent: reduction of SP content after 3 days but an increase after 12 or 21 days treatment, paradoxically with clinical improvement in each case. Drug‐induced changes in SP content could follow from changed release or synthesis from either neural or immune cells. The results suggest that both drugs have potential therapeutic value at different stages of inflammatory joint disease.
Activation of the extracellular signal‐regulated kinase 2 by metabotropic glutamate receptors
Authors: Ferraguti Francesco , Baldani‐Guerra Barbara , Corsi Mauro , Nakanishi Shigetada , Corti Corrado 
Published: March 2003

AbstractActivation of metabotropic glutamate receptors (mGluRs) leads to modulation of a variety of second messenger pathways probably including the mitogen‐activated protein kinase (MAPK) extracellular signal‐regulated protein kinases (ERK). MAPK play a key role in the control of cellular responses to changes in the external environment by regulating transcriptional activity and the phosphorylation state of several cytoplasmic targets. In this study, Chinese hamster ovary (CHO) cells permanently transfected with rat mGluR1a, mGluR2 and mGluR4 were employed as a model to examine the activation of MAPK by glutamate through mGluRs. All three mGluR subtypes rapidly stimulated ERK activation. In particular, mGluR1a and mGluR2 preferentially mediated phosphorylation and activation of ERK2 in a pertussis toxin (PTX)‐sensitive and concentration‐dependent manner. The activation was blocked completely by pretreatment with the antagonist (rs)‐α‐methyl‐4‐carboxyphenylglycine (MCPG) or with the MEK inhibitor PD098059. Furthermore, mGluR1a‐mediated ERK activation was suppressed by the depletion of endogenous protein kinase C (PKC) activity and by the PKC inhibitors staurosporine and calphostin C, but not chelerythrine. When cAMP was elevated in mGluR2‐expressing cells, by forskolin or dibutyryl‐cAMP, slight elevation of ERK activity was observed. However, glutamate‐stimulated ERK activation remained unaffected. In these cells, the phosphatidylinositol 3 kinase (PI3K) inhibitor wortmannin produced a significant, albeit only partial, inhibition of mGluR2‐mediated ERK activation. These findings raise the possibility of a MAPK cascade involvement in glutamate‐dependent neuronal plasticity mediated through stimulation of mGluRs.