bluuebunny/crossref_metadata_2025 huggingface dataset:
Nucleus Reticularis Thalami and Neocortical Paroxysms in the Rat
Authors: Marini Gabriella , Giglio Roberto , Macchi Giorgio , Mancia Mauro
Published: April 2006

The role of the nucleus reticularis thalami in spike‐wave discharges in rats with genetic absence epilepsy has already been demonstrated. This study further investigated the role of the nucleus reticularis thalami in paroxysmal synchronizations in Sprague‐Dawley rats; this strain shows no propensity to epileptic activity. Electroencephalographic patterns were followed in chronically implanted, unrestrained rats. After both electrolytic and chemical unilateral lesions, stereotaxically placed in the anterolateral sectors of this nucleus (verified post mortem), abnormal electroencephalographic rhythms (high‐voltage polyspikes and spike‐wave complexes) were recorded from the frontoparietal cortex, primarily in the contralateral hemisphere. Stereotyped discharges at 3 Hz developed progressively from multiple spikes within the alpha frequency range through the lengthening of the wave component. The excessive synchronized activity recorded from the intact hemisphere was of greater amplitude and occurred slightly earlier than from the lesioned hemisphere. These EEG patterns were associated with behavioural manifestations closely resembling those seen during absence seizures in humans. Bilateral lesions did not induce paroxysmal activity, both hemispheres being characterized by dominant delta/theta activity without signs of EEG‐synchronized sleep. The seizures may thus have been due to disinhibition of the contralateral reticularis nucleus, recently shown to project to the reticularis nucleus of the other side in rats. This working hypothesis is supported by callosal cuts. The results indicate that the reticular neurons exert a control over neocortical paroxysmal activity even in animals which do not present genetic absence epilepsy.
Receptive Field Properties of Starburst Cholinergic Amacrine Cells in the Rabbit Retina
Authors: Taylor W. Rowland , Wässle Heinz
Published: April 2006

Patch‐clamp recordings were made from ON starburst cholinergic amacrine cells with somas located in the ganglion cell layer of an isolated, dark‐adapted rabbit retina preparation. Light responses were analysed and cell identity was confirmed anatomically. The centre light response had a linear current‐voltage relation with a reversal potential close to 0 mV. The receptive field size was similar to the dendritic field size. Cholinergic amacrine cells displayed significant surround inhibition. The receptive field profile consisted of a central excitatory region flanked by an inhibitory surround. The surround attenuated the central response to 36% of the maximum. The surround was probably mediated by a combination of presynaptic and postsynaptic inhibition. Starburst amacrine cells did not display action potentials and the presence of a large, voltage‐dependent outward current limited depolarizing responses to a maximum potential of about ‐40 mV. Light responses were completely suppressed during application of 100 μM D, L‐2‐amino‐4‐phosphonobutyric acid (APB), consistent with activation exclusively through rod bipolar cells (on) and ON‐cone bipolar cells. In darkness the cells displayed a tonic inward current that could be blocked by 100 μM APB and 2 μM CNQX.
Distinct Temporal Patterns of Expression of Sodium Channel‐like Immunoreactivity During the Prenatal Development of the Monkey and Cat Retina
Authors: Miguel‐Hidalgo José J. , Angelides Kimon J. , Chalupa Leo M.
Published: April 2006

Polyclonal and monoclonal antibodies prepared against the α‐subunit of the voltage‐gated sodium channel (αNaCh) were used to examine the distribution of sodium channel‐like immunoreactivity during the prenatal development of the cat and rhesus monkey (Macaca mulatta) retina. At all prenatal ages studied, beginning on embryonic day 29 (E29) in the cat and E52 in the monkey, both antibodies labelled optic axons. With the polyclonal antibodies, the appearance of positive cells largely mirrored the onset of their morphological maturation. Immunoreactivity appeared first in the somata of ganglion cells, and subsequently the inner plexiform layer could be distinguished by its intense immunolabelling. A few weeks later horizontal cells displayed immunolabelling that extended to their dendrites in the developing outer plexiform layer. This was followed by immunoreactive cones, with bipolar cells labelled only postnatally. By contrast, with the monoclonal antibody some cells were found to be immunoreactive while their somata were still in the ventricular layer (E33 in cat and E52 in monkey). Many of these cells appeared to migrate to the outer portion of the prospective inner nuclear layer, where they gradually acquired the morphological appearance of bipolar cells. Transient expression of immunolabelling with monoclonal sodium channel antibody was found in the cones of the cat and cones and rods of the monkey. These results indicate that different types of αNaCh‐like proteins are expressed in the mammalian retina at distinct developmental periods. Their presence at very early stages during development suggests that these proteins could play a specific role in the commitment and/or differentiation of specific retinal cell types.
Interleukin‐1β Stimulates both Central and Peripheral Release of Vasopressin and Oxytocin in the Rat
Authors: Landgraf Rainer , Neumann Inga , Holsboer Florian , Pittman Quentin J.
Published: April 2006

Simultaneous microdialysis in the brain and blood was used to monitor the release of vasopressin and oxytocin within the hypothalamic supraoptic (SON) and paraventricular (PVN) nuclei and into the systemic circulation of urethane‐anaesthetized male rats before and after central administration of interleukin‐1β (IL‐1β). Following intracerebroventricular infusion of the cytokine (200 ng/5 μl), the content of vasopressin (up to 278% compared to vehicle‐treated control, P < 0.01 compared to vehicle‐treated control and preinfusion baseline) but not oxytocin (up to 148%, not significant) in 30‐min blood microdialysates was found to be increased. This peripheral release was accompanied by a transient rise in vasopressin (up to 163%, P < 0.05) and oxytocin (up to 182%, P < 0.05) release within the SON, the peak typically occurring during the first and second 30‐min collection intervals after IL‐1 β respectively. In contrast, in the simultaneously microdialysed PVN, both vasopressin and oxytocin failed to respond to intracerebroventricular IL‐1 β. In another series of experiments, IL‐1 β was directly infused (20 ng0.5 μl) into either the SON or PVN during microdialysis of the corresponding nucleus. The cytokine caused a significant and immediate rise in intra‐SON release of both vasopressin (up to 225%, P < 0.01) and oxytocin (up to 178%, P < 0.05). Again, in the PVN, nonapeptide release, although tending to be stimulated in response to intranuclear IL‐1 β, failed to reach statistical significance. The cytokine‐induced central and peripheral release pattern appeared to be independent of the rise in body temperature observed after IL‐1 β administration. In a third series of experiments, bilateral administration of IL‐1 β into the SON (20 ng/0.5 μl) failed to alter peripheral release of both vasopressin and oxytocin into the systemic circulation. The increase in central nonapeptide release in response to IL‐1 β shown in this paper supports the hypothesis that at least vasopressin might act to oppose central effects of the cytokine, including those on thermoregulation and behaviour, in this way contributing to the neuroendocrine‐immune dialogue at brain level.
α‐Bungarotoxin‐sensitive Nicotinic Receptors on Bovine Chromaffin Cells: Molecular Cloning, Functional Expression and Alternative Splicing of the α7 Subunit
Authors: García‐Guzmán Miguel , Sala Francisco , Sala Salvador , Campos‐Caro Antonio , Stühmer Walter , Gutiérrez Luis M. , Criado Manuel
Published: April 2006

Chromaffin cells from the bovine adrenal medulla express α‐bungarotoxin‐sensitive acetylcholine receptors whose subunit composition is unknown. Northern blot analysis showed that the α7 subunit, a main component of these α‐bungarotoxin‐sensitive acetylcholine receptors in avian and rat brain, is expressed in chromaffin cells. The cDNA of this bovine α7 subunit was cloned by polymerase chain reaction amplification of adrenal medulla RNA for detailed characterization of structure and function. The protein‐coding region revealed 92% amino acid sequence identity to rat α7 and 89% to chicken α7 subunits. The α‐bungarotoxin affinity of α7 homomers expressed in Xenopus oocytes was similar to that observed previously with native chromaffin α‐bungarotoxin sensitive acetylcholine receptors. Cross‐linking and sucrose gradient experiments suggested that, like the muscular and neuronal acetylcholine receptors, the α7 receptor has a pentameric structure. Upon activation with nicotinic agonists the α7 receptor exhibited rapidly desensitizing cation currents that were blocked by nicotinic antagonists and showed inward rectification. The amplification of adrenal medulla RNA by reverse transcription polymerase chain reaction methods revealed an alternatively spliced isoform of the bovine α7 subunit, where the exon that codes for the M2 transmembrane segment was skipped during mRNA processing. Oocyte expression of this isoform does not yield functional channels. However, this alternative mRNA exhibits dose‐dependent inhibition of α7 homomer expression when coinjected with the undeleted isoform.
α‐Latrotoxin is a Potent Inducer of Neurotransmitter Release in Torpedo Electric Organ—Functional and Morphological Characterization
Authors: Linial Michal , Llouz Nili , Feinstein Naomi
Published: April 2006

In this report we show that α‐latrotoxin from black widow spider venom is a potent activator of neurotransmitter release in synaptosomes from the Torpedo electric organ. Binding of the purified toxin (5 nM) to the synaptosomal fraction occurs already at 4°C and is dependent on the presence of divalent ions. However, neurotransmitter release commences only after temperature elevation (22°C) and is completed within 2 min. The effect of α‐latrotoxin on release is achieved at 1 nM and is already saturated at 5 nM. The release is stimulated by the presence of Ca2+ ions. Activation of release by α‐latrotoxin is accompanied by morphological changes in electric organ synaptosomes. The synaptosomes swell, resulting in a 55% increase in section area. Moreover, the number of synaptic vesicles per unit area decreases about three‐fold, and rows of docked synaptic vesicles are rarely detected as opposed to control synaptosomes. These morphological changes indicate that the massive release is mainly due to synaptic vesicle fusion. α‐Latrotoxin binding sites are highly concentrated in the innervated face of the electrocytes. lmmunoelectron microscopy on electric organ sections reveals alatrotoxin binding sites over the entire plasma membrane at release sites and facing Schwann cells surrounding Torpedo nerve terminals. Surprisingly, a high concentration of binding sites is also found at structures surrounding branching unmyelinated axons. This staining is in close proximity to Schwann cell envelopes and to the basal lamina around axonal tips. The mode of action of α‐latrotoxin in view of the localization of its binding sites is discussed.
Neuronal Sianallina of Information Imoortant to Visual Recognition‐Memory in Rat Rhinal aid Neighbouring Cortices
Authors: Zhu X. O. , Brown M. W. , Aggleton J. P.
Published: April 2006

This study was conducted to discover whether the rat cortex contains neurons that signal information concerning the previous occurrence of stimuli, as has been found in the primate. Recordings of the activity of 396 single neurons were made while unanaesthetized rats were shown objects. The effects on neuronal responsiveness of stimulus repetition and of the relative familiarity of the stimuli were sought. The areas sampled were the rhinal (entorhinal and perirhinal) cortex, area TE of the temporal cortex, the lateral occipital cortex and the hippocampal formation. The response to the first presentations of objects was significantly different from that to their second presentations for 63 (34%) of the 185 responsive neurons; for 39 of the neurons the response was smaller when the stimulus was repeated, whereas for 24 it was larger. The incidence of decremental responses was higher in the non‐hippocampal cortex than in the hippocampal formation, while the incidence of incremental responses was higher in the hippocampal formation than other cortical areas. The response to unfamiliar objects was significantly different from that to highly familiar objects for 15 (22%) of 67 responsive neurons so tested; for 12 of the neurons the response was smaller when the stimulus was repeated, and for three it was larger; most of these neurons were found in area TE. The responses of ten familiarity neurons varied significantly with the relative familiarity of the stimuli but not with stimulus repetition; the responses of seven recency neurons varied significantly upon stimulus repetition but not with the relative familiarity of the stimuli. Thus information concerning stimulus repetition and familiarity is separably encoded at the single neuron level in the rat cortex. The results demonstrate that in the rat cortex as in the monkey cortex there are neurons that signal information concerning the prior occurrence of stimuli; such information is of importance to recognition memory, working memory and priming memory.
Functional Involvement of Sciatic Nerve‐derived Versicanand Decorin‐like Molecules and other Chondroitin Sulphate Proteoglycans in ECM‐mediated Cell Adhesion and Neurite Outgrowth
Authors: Braunewell Karl‐Heinz , Pesheva Penka , McCarthy James B. , Furcht Leo T. , Schmitz Brigitte , Schachner Melitta
Published: April 2006

We have previously described two proteoglycans from human sciatic nerve which are immunochemically related to the chondroitin sulphate proteoglycans versican and decorin. The chondroitin sulphate of the versican‐like molecule and the core protein of the decorin‐like molecule have been found previously to be up‐regulated after lesioning the adult mouse sciatic nerve. To investigate if the versican‐ and decorin‐like molecules are involved in cell‐extracellular matrix interactions, we studied the effect of both molecules on cell adhesion. The versican‐ and decorin‐like molecules, substrate‐coated in a mixture with fibronectin, but not with laminin or collagen types I or IV, inhibited the adhesion of several cell lines, neonatal dorsal root ganglion neurons and Schwann cells. The inhibitory activity was concentration‐dependent and mediated by the chondroitin sulphate. Furthermore, when different proteoglycans were incubated with fibronectin, only the versican‐ and decorin‐like molecules and the chondroitin sulphate proteoglycan aggrecan, but not the heparan sulphate proteoglycan perlecan, were able to inhibit fibronectin‐mediated cell adhesion. The versican‐ and decorin‐like molecules, substrate‐coated alone or in a mixture with fibronectin or laminin, were at most slightly inhibitory to neurite outgrowth from PC12 phaeochromocytoma cells and neonatal dorsal root ganglion neurons. In a solid‐phase ligand‐binding assay theversican‐ and decorin‐like molecules interacted with fibronectin, but not with laminin or collagen types I and IV. Binding of the versican‐like molecule to fibronectin and inhibition of cell adhesion by this molecule was mediated via the heparin and cell‐binding domains of fibronectin. These observations suggest that binding of the two proteoglycans to fibronectin is involved in the modulation of adhesion of cells to fibronectin.
Electrophysiological Characterization of a Novel Conotoxin That Blocks Molluscan Sodium Channels
Authors: Hasson Arik , Fainzilber Michael , Zlotkin Eliahu , Spira Micha
Published: April 2006

A novel peptide toxin, PnIVB, isolated from the venom of Conus pennaceus blocks voltage‐gated sodium current in Aplysia neurons. Complete blockade is obtained at a PnIVB concentration of 80±2.2 nM and 50% blockade at 16±0.86 nM. The potency of PnIVB in blocking Aplysia sodium current is four orders of magnitude larger than that of tetrodotoxin. The toxin has no paralytic activity when injected into fish. The rapid blockade of sodium current by PnIVB is not associated with a change in the activation or inactivation kinetics of the current, or with the reversal potential. Sodium current blockade is reversible after a 30 min wash with 50 times the bath volume. The novel conotoxin PnlVB can be used as a powerful tool for mollusc neurobiological research and as a molecular probe to explore the structure‐function relations of voltage‐gated sodium channel subtypes.
Morphological Fate of Rhombomeres in Quail/Chick Chimeras: A Segmental Analysis of Hindbrain Nuclei
Authors: Marín Faustino , Puelles Luis
Published: April 2006

Quail rhombomeres two to six (r2‐r6) were individually grafted homotopically into the hindbrain of chick embryos at 2 days of incubation. Nine to 10 days after the operation the chimeric embryos were fixed and processed for parallel cytoarchitectural and immunocytochemical study (with an anti‐quail antibody) in order to map the anatomical fate of the grafted tissue. Emphasis was placed on conventionally identified and distinct neuronal populations composing the sensory and motor longitudinal columns. Grafted rhombomeres consistently developed as complete transverse slices of the chimeric hindbrain. Interrhombomeric cell migration was either sparse or restricted to specific nuclei. The cranial nerve motor nuclei showed rhombomeric origins consistent with the patterns described in early embryos. Unexpectedly, alar r2 was found to form the auricular part of the cerebellum. As regards the cochlear nuclei, we found that nucleus angularis derives from r3 to r6, nucleus laminaris from r5 to r6, nucleus magnocellularis from r6 to r7 and nucleus olivaris superior from r5. The nuclei of the lateral lemniscus originated between r1 and r3. We also delimited the respective rhombomeric subdivisions of the sensory vestibular and trigeminal columns, both of which extend from r1 caudalwards throughout the hindbrain. There were consistently some interrhombomeric neuronal migrations inside the vestibular column, some motor nuclei and the reticular formation, involving only one rhombomere length. The pontine nuclei, which extended from r1 to r7, showed neuronal migrations that crossed several rhombomeres. On the whole, these results represent the first anatomical analysis of the mature avian hindbrain in terms of rhombomere‐derived domains.
Immunohistochemical Visualization of Brain‐derived Neurotrophic Factor in the Rat Brain
Authors: Dugich‐Djordjevic Millicent M. , Peterson Christine , Isono Fujio , Ohsawa Fukuichi , Widmer Hans R. , Denton Timothy L. , Bennett Gregory L. , Hefti Franz
Published: April 2006

A purified polyclonal antibody preparation was made against recombinant human brain‐derived neurotrophic factor (BDNF) in guinea pig and characterized for use in immunoassays and immunohistochemistry. The anti‐BDNF antibodies specifically recognized BDNF in Western blots and immunoprecipitation. There was no cross‐reactivity with the other known mammalian members of the neurotrophin family, nerve growth factor, neurotrophin‐3 and neurotrophin‐4/5. In immunohistochemical analysis, the anti‐BDNF recognized exogenous BDNF injected into the brain of rats, whereas no signal was obtained with the other neurotrophins. Preabsorption with native BDNF abolished the immunoreactivity in brain sections. These studies identify the anti‐BDNF as a tool for immunocytochemistry and the development of an immunoassay. Immunohistochemical analysis revealed widespread neuronal localization of BDNF in many brain areas. BDNF was localized in all subpopulations of hippocampal neurons. The distribution in the hippocampus suggests localization in the cytoplasm of cell bodies and dendrites.
Early Changes in Prodynorphin mRNA and ir‐Dynorphin A Levels after Kindled Seizures in the Rat
Authors: Romualdi P. , Donatini A. , Bregola G. , Bianchi C. , Beani L. , Ferri S. , Simonato M.
Published: April 2006

Prodynorphin mRNA and immunoreactive dynorphin A (ir‐dynorphin A) levels were measured in different brain areas at various time points after amygdala kindled seizures. In the hippocampus, striatum and hypothalamus, prodynorphin mRNA levels were not significantly changed in kindled rats (killed 1 week after the last stimulus‐evoked seizure), but they were significantly increased 1 h after seizures. The relative increase was the highest in the hippocampus (∼3‐fold). In the brainstem, midbrain and cerebral cortex no changes in prodynorphin mRNA were detected in kindled rats, 1 h or 1 week after a kindled seizure. ir‐Dynorphin A levels were significantly reduced in the hippocampus and in the striatum of kindled rats, as well as 5 and 60 min after kindled seizures, but they were increased back to control levels after 120 min. In the hypothalamus, ir‐dynorphin A levels were significantly increased 120 min after a kindled seizure. ir‐Dynorphin A levels were also significantly reduced in the brainstem and in the frontal, parietal and temporal cortex 120 min, but not 5 or 60 min, after a kindled seizure. Taken together, these data support the hypothesis that the dynorphinergic system is activated after amygdala kindled seizures, with different kinetics in different brain areas.
Glial Cells in the Mouse Hippocampus Express AMPA Receptors with an Intermediate Ca2+ Permeability
Authors: Seifert Gerald , Steinhäuser Christian
Published: April 2006

Recently, we could demonstrate that ‘complex’ glial cells in mouse hippocampal slices express glutamate receptor channels of the α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA)/kainate subtypes. In the present study, we further characterized this glial receptor. Since voltage‐clamp control is imperfect and diffusion barriers hinder the quantitative analysis of the receptor currents in situ, the patch‐clamp technique was applied to glial cells acutely isolated from the mouse hippocampal CA1 stratum radiatum subregion. A concentration‐clamp technique was used which enabled very fast exchange of the extracellular solutions. Thus, it was possible to characterize the transient receptor currents with high time resolution. Application of L‐glutamate, AMPA and L‐homocysteate induced rapidly activating and fast desensitizing receptor currents in the suspended glial cells. In contrast, kainate induced non‐desensitizing currents. The corresponding dose‐response curve revealed a half‐maximum of current activation at 350 μM. The current/voltage relationship of the kainate‐evoked response was linear, with a reversal potential at ∼9 mV. Analysis of the reversal potential in solutions containing high concentrations of CaCl2 confirmed earlier in situ data by demonstrating significant Ca2+ permeability of the glial glutamate receptor channels in the hippocampus. The kainate‐induced receptor currents were markedly increased by cyclothiazide, a substance which selectively potentiates glutamate receptors of the AMPA subtype. We conclude that glial cells of the juvenile hippocampus mainly express heteromeric high‐affinity AMPA receptors. Most probably, the receptor channels are assembled from the low Ca2+‐permeable glutamate receptor‐2 subunit together with Ca2+‐permeable AMPA‐preferring subunits.
Activation of Class II or III Metabotropic Glutamate Receptors Protects Cultured Cortical Neurons Against Excitotoxic Degeneration
Authors: Bruno V. , Battaglia G. , Copani A. , Giffard R. G. , Raciti G. , Raffaele R. , Shinozaki H. , Nicoletti F.
Published: April 2006

Trans‐1‐aminocyclopentane‐1,3‐dicarboxylic acid, a mixed agonist of all metabotropic glutamate receptor (mGluR) subtypes, is known to produce either neurotoxic or neuroprotective effects. We have therefore hypothesized that individual mGluR subtypes differentially affect neurodegenerative processes. Selective agonists of subtypes which belong to mGluR class II or III, such as (2s, 1′R,2′R,3′R)‐2‐(2,3‐dicarboxycyclopropyl)‐glycine (DCG‐IV) (specific for subtypes mGluR2 or 3) or L‐2‐amino‐4‐phosphonobutanoate and L‐serine‐O‐phosphate (specific for subtypes mGluR4, 6 or 7), were highly potent and efficacious in protecting cultured cortical neurons against toxicity induced by either a transient exposure to N‐methyl‐D‐aspartate (NMDA) or a prolonged exposure to kainate. In contrast, agonists that preferentially activate class I mGluR subtypes (mGluR1 or 5), such as quisqualate or trans‐azetidine‐2,3‐dicarboxylic acid, were inactive. DCG‐IV was still neuroprotective when applied to cultures after the toxic pulse with NMDA. This delayed rescue effect was associated with a reduction in the release of endogenous glutamate, a process that contributes to the maturation of neuronal damage. We conclude that agonists of class II or III mGluRs are of potential interest in the experimental therapy of acute or chronic neurodegenerative disorders.
Optical Imaging of the Layout of Functional Domains in Area 17 and Across the Area 17/18 Border in Cat Visual Cortex
Authors: Bonhoeffer Tobias , Kim Dae‐Shik , Malonek Dov , Shoham Doron , Grinvald Amiram
Published: April 2006

Optical imaging based on intrinsic signals was used to investigate the functional architecture of cat area 17 and the border between areas 17 and 18. The visual stimuli were gratings of different spatial frequencies moving at different angles, in different directions and with different speeds. In area 17 the iso‐orientation domains were usually organized in patches rather than as elongated bands. Patches with different orientation preferences were arranged radially forming ‘pinwheels’ around ‘orientation centres’. The pinwheel density was ∼1.7‐fold higher than in area 18. To explore clustering according to direction of motion, stimuli having the same orientation but moving in opposite directions were used. These two stimuli yielded very similar activity maps giving no indication of robust directionality clustering. Using near infrared light we were able to simultaneously image ocular‐dominance and iso‐orientation domains. A quantitative assessment of the relative strengths of the two subsystems showed that in upper cortical layers clustering according to orientation preference was three‐fold stronger than clustering according to ocular dominance. The functional organization of spatial frequency was also examined. When we compared the activated regions by stimuli having different spatial frequency and moving at different velocities we observed that neurons were clustered also in these respects. We also investigated the functional architecture at the area 17/18 border and found that orientation maps at both sides of the border were not independent of each other. The map of area 17 smoothly blended into that of area 18. Similarly, the preferred spatial frequency of the neurons changed gradually over a distance of ∼0.8 mm at the region of the area 17/18 border.
Alternative RNA Splicing Generates Diversity of Neuropeptide Expression in the Brain of the Snail Lymnaea: In Situ Analysis of Mutually Exclusive Transcripts of the FMRF amide Gene
Authors: Santama Niovi , Benjamin Paul R. , Burke Julian F.
Published: April 2006

In the CNS of the snail Lymnaea stagnalis, Phe‐Met‐Arg‐Phe‐amide (FMRFamide)‐like and additional novel neuropeptides are encoded by a common, multi‐exon gene. This complex locus, comprising at least five exons, is subject to post‐transcriptional regulation at the level of alternative RNA splicing. Our aim was first to analyse the pattern by which exons of this neuropeptide locus combine during splicing of the primary RNA transcript, and second to investigate the functional significance of splicing by mapping the expression and neuronal localization in the CNS of the alternative mRNA transcripts, in the context of defined neuronal networks and single identified neurons. The approach was a combination of comparative in situ hybridization and immunocytochemistry, using a battery of exon‐specific oligonucleotides and anti‐peptide antisera. The analysis illustrated that exons III, IV and V were always coexpressed and colocalized whereas the expression of exon II was always differential and mutually exclusive. Both sets of exons were, however, coexpressed with exon I: the total number of exon I‐expressing neurons was equal to the combined number of neurons expressing exon III/IVA/ and neurons expressing exon II. In addition, it was revealed that the extreme 5’of exon II, encoding a potential hydrophobic leader signal, was not expressed in the CNS of Lymnaea but was apparently spliced out during RNA processing. Both mRNA transcripts of the FMRFamide locus, type I (exons I/II) and type 2 (exons I/III/IV/V), were translated in the CNS and the resulting protein precursors were also expressed in a mutually exclusive fashion, as were their respective transcripts. The expression of alternative transcripts within identified networks or neuronal clusters was heterogeneous, as exemplified by the cardiorespiratory network. On the basis of this work and a previous cDNA analysis, we put forward a revised model of differential splicing and expression of the FMRFamide gene in the CNS of Lymnaea.
Fibroblast Growth Factors and Insulin Growth Factors Combine to Promote Survival of Rat Schwann Cell Precursors Without Induction of DNA Synthesis
Authors: Gavrilovic Jelena , Brennan Angela , Mirsky Rhona , Jessen Kristjan R.
Published: April 2006

In embryonic rat nerves, we recently identified an early cell in the Schwann cell lineage, the Schwann cell precursor. We found that when these cells were removed from contact with axons they underwent rapid apoptotic death, and that in a proportion of the cells this death could be prevented by basic fibroblast growth factor (bFGF, FGF‐2). We now report that 100% of Schwann cell precursors isolated from peripheral nerves of 14‐day‐old‐rat embryos can be rescued by a combination of insulin‐like growth factor (IGF) 1 or 2 in combination with either acidic FGF (aFGF, FGF‐1), bFGF or Kaposi's sarcoma FGF (K‐FGF; FGF‐4). The precursors display an absolute requirement for both an IGF and an FGF to achieve maximal survival. Elevation of intracellular levels of cAMP by forskolin does not result in a significant shift in the IGF/FGF dose‐response curves. In contrast, the percentage of precursors rescued by FGF in the presence of insulin is dramatically increased by elevation of cAMP. These growth factor combinations did not stimulate DNA synthesis significantly in Schwann cell precursors. These findings show that cooperation between growth factors is required to suppress cell death in Schwann cell precursors, and suggest that survival and DNA synthesis are regulated by distinct growth factor combinations in these cells. The observations are consistent with the idea that survival regulation by FGFs and IGFs plays an important role in the development of glial cells in early embryonic nerves.
Distinct Populations of Identified Glial Cells in the Developing Rat Spinal Cord Slice: Ion Channel Properties and Cell Morphology
Authors: Chvátal Alexandr , Pastor Andrea , Mauch Marianne , Syková Eva , Kettenmann Helmut
Published: April 2006

Four types of glial cells could be distinguished in the grey matter of rat spinal cord slices at postnatal days 1‐19 (P1‐P19), based on their pattern of membrane currents as revealed by the whole cell patch clamp technique, and by their morphological and immunocytochemical features. The recorded cells were labelled with Lucifer Yellow, which allowed the subsequent identification of cells using cell‐type‐specific markers. Astrocytes were identified by positive staining for glial fibrillary acidic protein (GFAP). These were morphologically characterized by multiple, very fine and short processes and electrophysiologically by symmetrical, non‐decaying K+ selective currents. Oligodendrocytes were identified by a typical oligodendrocyte‐like morphology, lack of GFAP staining and positive labelling with a combination of O1 and O4 antibodies (markers of the oligodendrocyte lineage), and their membrane was dominated by symmetrical, passive, decaying K+ currents. The third population of glial cells was also characterized by positive staining for O1/O4 or only for O4 antigens, lack of GFAP staining and, in some cells, oligodendrocyte‐like morphology. However, these cells could be distinguished by the presence of inwardly rectifying (KDR), delayed outwardly rectifying (KDR) and A‐type K+ currents (KA), representing the most likely glial precursor cells of the oligodendrocyte lineage. The fourth population of glial cells had small somata and a widespread network of long processes with no apparent orientation preference. In one case, processes were positively labelled with GFAP, while 30% were characterized by faint, diffuse staining. These cells expressed a complex pattern of voltage‐gated channels, namely Na+, KDR, KA and KIR channels. In contrast to neurons, the amplitude of Na+ currents was at least one order of magnitude smaller than the K+ currents, and none of these cells showed the ability to generate action potentials in the current clamp mode. Since none of these cells could be labelled by oligodendrocyte markers we assume that they were either astrocytes or glial precursor cells of the astrocyte lineage. The four cell types were found in all regions of the grey matter. When randomly accessing the glial cells, the probability of recording from the oligodendrocyte precursor cells and the glial cells with Na+ currents decreased during development. At P1‐P3, 50% of the cells revealed the Na+ current, while at P13‐P15 only 18% did. Concomitantly, the number of glial cells with astrocyte‐ and oligodendrocyte‐like membrane currents increased from 19 and 12% to 41 and 35.5% respectively. We conclude that the glial cells in the spinal cord slices possess distinct morphological, immunohistochemical and physiological properties, and that the glial populations undergo changes during postnatal development.
Glutamate Induces c‐fos Proto‐oncogene Expression and Inhibits Proliferation in Oligodendrocyte Progenitors: Receptor Characterization
Authors: Liu Hsueh‐Ning , Almazan Guillermina
Published: April 2006

The effect of glutamate on c‐fos expression in oligodendrocyte progenitors was investigated by Northern blot analysis. Glutamate caused rapid and transient induction. Both 6‐cyano‐7‐nitro‐quinoxaline‐2,3‐dione (CNQX) and 6,7‐dinitroquinoxaline‐2,3‐dione (DNQX), two competitive non‐NMDA ionotropic receptor antagonists, reduced glutamate‐induced c‐fos expression, whereas the NMDA antagonist MK‐801 was ineffective. In addition, the glutamate receptor agonists (±)‐α‐amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionic acid hydrobromide (AMPA) and kainate strongly induced c‐fos. However, the metabotropic receptor agonist trans‐(±)‐1‐amino‐(1S,3R)‐cyclopentanedicarboxylic acid (trans‐(±)‐ACPD) did not increase c‐fos mRNA level and the antagonist L‐(+)‐2‐amino‐3‐phosphonopropionic acid did not block glutamate‐induced c‐fos mRNA. These findings indicate that c‐fos induction in oligodendrocyte progenitors is mediated through the AMPA/kainate receptors, while NMDA and metabotropic receptor subtypes are not involved. Chelation of extracellular calcium by EDTA prevented glutamate‐induced c‐fos expression. Similarly, the protein kinase C inhibitor 1‐(5‐isoquinoline‐sulphonyl)‐2‐methylpiperazine dihydrochloride (H7) and down‐regulation of protein kinase C by prolonged exposure to phorbol‐12‐myristate 13‐acetate blocked c‐fos induction. These results suggest that induction of c‐fos through AMPA/kainate receptors is dependent on extracellular calcium influx and involves downstream activation of phorbol ester‐sensitive protein kinase C. The effect of glutamate on oligodendrocyte progenitor proliferation was assessed by [3H]thymidine incorporation. Glutamate and the agonists kainate and AMPA, but not trans‐(±)‐ACPD, caused a dose‐dependent decrease in [3H]thymidine incorporation. All these pharmacological agents were not toxic to oligodendrocyte progenitors. CNQX reversed the inhibitory effects produced by glutamate and the various agonists. These results suggest that glutamate may modulate the growth and differentiation of oligodendrocytes in the central nervous system.
Noradrenaline Increases K‐conductance and Reduces Glutamatergic Transmission in the Mouse Entorhinal Cortex by Activation of α2‐Adrenoreceptors
Authors: Pralong Etienne , Magistretti Pierre J.
Published: April 2006

The entorhinal cortex is a gateway to the hippocampus; it receives inputs from several cortical associative areas as well as subcortical areas. Since there is evidence showing that noradrenaline reduces the epileptic activity generated in the entorhinal cortex, we have examined the action of noradrenaline in the superficial layer of the entorhinal cortex, which is the main source of afferents to the hippocampus. In a previous study we showed that noradrenaline hyperpolarized layer II entorhinal cortex neurons and reduced global synaptic transmission via α2‐adrenoreceptors. Here we present a detailed analysis of the effect of noradrenaline on membrane resistance and on the pharmacologically isolated postsynaptic potentials in layer II entorhinal cortex neurons of mice. Noradrenaline (50 μM) hyperpolarized most layer II entorhinal cortex neurons. This hyperpolarization corresponded to an outward current with a reversal potential following the Nernst equilibrium potential for potassium. The hyperpolarizing effect of noradrenaline was blocked by 10 μM yohimbine. These observations suggest that noradrenaline activates a potassium conductance via an α2‐adrenoreceptor. Noradrenaline (10–50 μM) reversibly reduced the amplitude of the pharmacologically isolated excitatory potentials mediated by both NMDA and α‐amino‐3‐hydroxy‐5‐methyl‐isoxazole‐propionic acid (AMPA) receptors, the former being more strongly affected. Again this effect was blocked by 10 μM yohimbine. In contrast, GABAA‐mediated synaptic transmission was virtually unaffected by noradrenaline. Thus, noradrenaline appears to strongly inhibit the glutamate‐mediated synaptic transmission in the entorhinal cortex without affecting inhibitory post‐synaptic potentials. These observations suggest that α2‐adrenergic receptor agonists may exert a beneficial effect in the control of hyperexcitability in temporal lobe epilepsy.
High Specificity of Neurotrophins in the Embryonic Chicken Trigeminal System
Authors: Pinon Luzia G. P. , Robinson Michelle , Davies Alun M.
Published: April 2006

Studies of cell lines and some cultured neurons have demonstrated potential cross‐talk between neurotrophins and their receptors; high concentrations of neurotrophins can exhibit either agonist or antagonistic actions on heterologous neurotrophin receptors. We have studied neurotrophin discrimination among the sensory neurons of the embryonic chicken trigeminal system. We show that nerve growth factor (NGF) at a concentration that is six orders of magnitude greater than that required to promote the survival of NGF‐dependent dorsomedial trigeminal ganglion (DMTG) neurons has no effect on the survival of brain‐derived neurotrophic factor (BDNF)‐dependent trigeminal mesencephalic nucleus (TMN) neurons and does not affect the dose‐response relationship of these neurons to BDNF. A similar high level of neurotrophin‐3 neither promotes the survival of BDNF‐dependent ventrolateral trigeminal ganglion neurons nor affects the dose response of these neurons to BDNF. High levels of BDNF have a negligible effect on the survival of mid‐embryonic DMTG neurons. These results show that some neurons are able to discriminate completely between neurotrophins at very high concentrations, indicating that neurotrophin responses can be far more highly specific than previously appreciated.
Membrane Potential Oscillations of Neonatal Rat Spinal Motoneurons Evoked by Electrical Stimulation of Dorsal Root Fibres
Authors: Baranauskas Gytis , Nistri Andrea
Published: April 2006

Intracellular recording from lumbar motoneurons of the neonatal rat isolated spinal cord bathed in standard saline solution was used to study membrane potential oscillations which accompanied the decay phase of excitatory postsynaptic potentials (EPSP) induced by single electrical pulses to an adjacent dorsal root. About 60% of motoneurons displayed rhythmic oscillations of 10 ± 2 mV maximal amplitude and 7 ± 0.5 Hz frequency. Ability to generate oscillations could not be correlated to the cell membrane properties or to the age of the preparation (5–13 days). The oscillation frequency was independent of membrane potential (‐100 to ‐45 mV) or of the intensity of dorsal root stimuli. The oscillation amplitude was linearly related to the cell potential within the same voltage level. Fast Fourier transform analysis showed that the power spectrum of oscillations peaked at ˜8 Hz. Electrically evoked activities and spontaneous events displayed similar cut‐off frequencies. When the cell membrane potential was steadily depolarized, a hyperpolarizing pulse applied during the decay phase of the EPSP promptly revealed the presence of oscillatory behaviour. Pharmacological block of neurokinin‐1 or N‐methyl‐D‐aspartate receptors depressed the decay phase of the EPSP and the associated oscillatory responses. It is suggested that rhythmic oscillations were probably due to summated synaptic potentials generated at the premotoneuron level and are perhaps of functional relevance to motoneuron behaviour.
Inhibitors of Type IV Phosphodiesterases Reduce the Toxicity of MPTP in Substantia Nigra Neurons In Vivo
Authors: Hulley Philippa , Hartikka Jukka , Abdel'Al Samir , Engels Peter , Buerki Hans‐Rudolf , Wiederhold Karl‐Heinz , Müller Thomas , Kelly Peter , Lowe David , Lübbert Hermann
Published: April 2006

The neuropathology of Parkinson's disease is characterized by the degeneration of dopaminergic neurons in the substantia nigra. We have recently shown that the activation of protein kinase A improves the survival of dopaminergic neurons in culture and, furthermore, protects them from the dopaminergic neurotoxin, 1‐methyl‐4‐phenylpyridinium ion (MPP+) in vitro. We have now analysed the potential of phosphodiesterase inhibitors to increase cAMP levels in dopaminergic neurons, to improve their survival in culture and to protect them from the toxicity of 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP) in vivo. Increasing intracellular cAMP with phosphodiesterase type IV‐specific inhibitors enhanced the survival of dopaminergic neurons in culture. Inhibitors of other phosphodiesterase types were not active. In vivo, phosphodiesterase type IV inhibitors reduced the MPTP‐induced dopamine depletion in the striatum of C57BL/6 mice. Furthermore, the loss of tyrosine hydroxylase‐immunopositive neurons in the substantia nigra of these animals was diminished. After Nissl staining, a similar reduction of the MPTP‐induced loss of neurons was observed in the substantia nigra. The protective effect of protein kinase A activation did not appear to be due to the blocking of MPP+ uptake into dopaminergic neurons. This was not decreased after treatment with forskolin or 8‐(4‐chlorophenylthio)‐cAMP. Thus, protein kinase A regulates the survival and differentiation of dopaminergic substantia nigra neurons in vivo, implicating a therapeutic potential for substances which regulate cAMP turnover in these neurons.
IL‐6 Expression in Neurons of Transgenic Mice Causes Reactive Astrocytosis and Increase in Ramified Microglial Cells but no Neuronal Damage
Authors: Fattori Elena , Lazzaro Domenico , Musiani Piero , Modesti Andrea , Alonzi Tonino , Ciliberto Gennaro
Published: April 2006

Growing evidence suggests that aberrant production of inflammatory cytokines within the central nervous system (CNS) contributes to the development of pathological conditions. To test the cause—effect relationship between the overproduction of interleukin‐6 (IL‐6) in the CNS and the onset of neuropathological changes, we have generated transgenic mice in which human IL‐6 expression has been targeted to the neurons by using the rat neuron‐specific enolase promoter. These mice develop reactive astrocytosis and an increase in ramified microglial cells but do not show histological or behavioural signs of neuron damage at the light microscope level. We thus conclude that a constant release of human IL‐6 by neuronal subpopulations in mice is sufficient to activate cells potentially capable of modulating the local immune response, but at the same time is compatible with normal neuron functions.
Dopamine Autoreceptor Sensitivity is Unchanged in Rat Nucleus Accumbens After Chronic Haloperidol Treatment: An In Vivo and In Vitro Voltammetric Study
Authors: Chesi Alexander J. R. , Feasey‐Truger Karin J. , Alzheimer Christian , Bruggencate Gerrit ten
Published: April 2006

Fast cyclic voltammetry was used to assess the effects of chronic oral haloperidol treatment (0.7 mg/kg/day for 21 days) on the sensitivity of dopamine autoreceptors in the rat nucleus accumbens both in vivo and in vitro. Evoked dopamine overflow was significantly reduced after chronic haloperidol treatment, but the sensitivity of dopamine overflow to sulpiride, an antagonist at release‐inhibiting dopamine autoreceptors, and quinpirole, an agonist at these receptors, was unchanged. The estimated EC50 values for quinpirole and sulpiride (52 and 60 nM respectively) obtained in vitro and the receptor distribution profiles published in the literature suggest that the autoreceptors involved in this modulation are mainly of the D3 subtype. The finding that the reduced dopamine overflow in the nucleus accumbens observed after chronic treatment with a classical neuroleptic is not due to dopamine autoreceptor supersensitivity may therefore be the first functional evidence for unchanged autoreceptor activity in the nucleus accumbens, supporting biochemical findings of a lack of D3 autoreceptor up‐regulation after chronic haloperidol treatment. It lends further support to the assumption that the long‐term changes occurring during chronic neuroleptic treatment may not lie at the level of presynaptic dopamine receptor regulation.
Prominent Expression of bFGF in Dorsal Root Ganglia after Axotomy
Authors: Ji R.‐R. , Zhang Q. , Zhang X. , Piehl F. , Reilly T. , Pettersson R. F. , Hökfelt T.
Published: April 2006

Using quantitative in situ hybridization and immunohistochemistry the expression of acidic and basic fibroblast growth factors (aFGF, bFGF) in dorsal root ganglia (DRGs) was examined. Around 5% of the small neurons expressed bFGF mRNA in normal DRGs. Nerve injury induced a very dramatic and rapid up‐regulation in bFGF mRNA levels, and around 80% of all DRG neurons expressed bFGF mRNA 3 days after axotomy. A distinct increase in bFGF‐like immunoreactivity (LI) was also detected as early as 15 h after axotomy. The elevation of bFGF mRNA and protein levels declined after 1 week. bFGF mRNA was also up‐regulated in non‐neuronal cells following axotomy. Normally bFGF‐LI was mainly localized in the nuclei of DRG neurons and in some non‐neuronal cells. After nerve section, bFGF‐LI was in addition found in the cytoplasm, and many more bFGF‐positive non‐neuronal cells were observed. By means of confocal microscopy analysis of axotomized DRGs, some bFGF‐LI could be detected in vesicle‐like structures in the cytoplasm as well as in the nucleoli, in addition to the nuclear location. Application of leukaemia inhibitory factor to the transected sciatic nerve significantly increased the number of bFGF‐positive neurons, whereas the bFGF‐LI in non‐neuronal cells was strongly suppressed. About 70% of the normal DRG neurons expressed aFGF mRNA and aFGF‐LI. Axotomy produced a moderate increase in aFGF mRNA levels, but no detectable effect on protein levels. Taken together, the results show that bFGF may be involved in the neuronal response to injury and suggest a role in neuronal survival and regeneration in axotomized DRG neurons.