bluuebunny/crossref_metadata_2025 huggingface dataset:
Decrease of zif‐268 and c‐fos and Increase of c‐jun mRNA in the Cat Areas 17, 18 and 19 Following Complete Visual Deafferentation
Authors: Zhang Fan , Vanduffel Wim , Schiffmann Serge N. , Mailleux Pierre , Arckens Lutgarde , Vandesande Frans , Orban Guy A. , Vanderhaeghen Jean‐Jacques
Published: April 2006

We used in situ hybridization to investigate the effect of complete visual deafferentation on immediate early gene expression in adult cat visual cortex. Deafferentation was obtained by unilateral section of the optic tract and sections of both the corpus callosum and anterior commissure. In this model, one hemisphere served as control for the other within the same animal. A decrease in zinc finger protein (zif)‐268 and c‐fos mRNA was observed in the superficial and deep layers of areas 17 and 18, and all layers of area 19 in the deafferented hemisphere. This decrease, present 3 days after surgery, was maximal after 30 days. An increase of c‐jun mRNA was observed in the deep layers of areas 17, 18 and 19 in the deafferented hemisphere 3, 10 and 30 days after surgery. These results suggest that visual input activates zif‐268 and c‐fos expression and tonically depresses c‐jun expression in the primary visual complex yielding similar levels of c‐jun and c‐fos expression in normal conditions.
Polymer Encapsulated Cell Lines Genetically Engineered to Release Ciliary Neurotrophic Factor Can Slow Down Progressive Motor Neuronopathy in the Mouse
Authors: Sagot Y. , Tan S. A. , Baetge E. , Schmalbruch H. , Kato A. C. , Aebischer P.
Published: April 2006

Ciliary neurotrophic factor (CNTF) has recently generated great interest due to its potential as a therapeutic agent for the treatment of human neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS). Because the systemic half‐life of CNTF is only in the order of a few minutes, continuous delivery of this trophic factor could be attractive or even necessary in the therapy of these diseases. One promising technique involves the polymer encapsulation of cells which have been genetically modified to secrete neurotrophic factors. The polymer capsules can be implanted into animals and effect the slow release of the protein for several months. The encapsulation technique immuno‐isolates the foreign cells from host immune cells and at the same time prevents tumour formation by the transplanted cells. In this study, we have used progressive motoneuronopathy (pmn) mice to determine the extent to which encapsulated cell lines secreting CNTF could alter the course of the disease. pmn/pmn homozygotes present severe loss of myelinated motor fibres and a significant reduction of facial motoneuron cell bodies. The mice develop weakness of the hindlimbs and die during the sixth week after birth. We found that CNTF delayed the disease progression by increasing the survival time by 40% and by improving motor function as assessed by three behavioural tests. Moreover, histological counts of the phrenic nerve myelinated axons and facial nucleus motoneurons indicated a significant reduction of motoneuron loss. These results suggest that polymer‐encapsulated cells releasing neurotrophic factors may provide a potential delivery system for treating neurodegenerative diseases such as ALS.
HIV‐1 gp120 Glycoprotein Induces [Ca2+]i Responses not only in Type‐2 but also Type‐1 Astrocytes and Oligodendrocytes of the Rat Cerebellum
Authors: Codazzi Franca , Menegon Andrea , Zacchetti Daniele , Ciardo Alberto , Grohovaz Fabio , Meldolesi Jacopo
Published: April 2006

Cultures of cerebellar cortex cells were exposed to the HIV‐1 envelope glycoprotein, gp120, and investigated for cytosolic Ca2+ ion concentration ([Ca2+]i) changes by the fura‐2 ratio videoimaging technique while bathed in complete, Na+‐free or Mg2+‐free Krebs‐Ringer media. At the end of the [Ca2+]i experiments the cells were fixed and immunoidentified through the revelation of markers specific for neurons (microtubule associated protein‐2), type‐2 (A2B5) or all (glial fibrillary acidic protein) astrocytes, oligodendrocytes (galactocerebroside) or microglia (F4/80 antibody). In complete medium, rapid biphasic (spike‐plateau) responses induced by gp120 (0.1–1 nM) were observed in a subpopulation of type‐2 astrocytes. In addition, slow but progressive responses were observed in other type‐2 cells and oligodendrocytes, whereas type‐1 astrocytes showed small responses, if any, and granule neurons did not respond at all. Use of Na+‐free medium (a condition that blocked another gp120‐induced response, cytosolic alkalinization) resulted in an increase in [Ca2+]i response that was appreciable not only in type‐2 but also in most type‐1 astrocytes, possibly because of the inhibition of the Na+/Ca2+ exchanger and the ensuing decrease in Ca2+ extrusion. Granule neurons, including those in direct contact with responsive astrocytes, remained unresponsive, even when the experiments were carried out in Mg2+‐free medium supplemented with glycine, a condition that favours activation of the glutamatergic N‐methyl‐D‐aspartate (NMDA) receptor. The results obtained demonstrate that sensitivity to gp120 is a property of not only a few type‐2 astrocytes but of the majority of cerebellar glial cells, which, however, do not respond to the protein with glutamate release, as indicated by the negative results obtained with NMDA‐receptor‐expressing granule neurons. Single glial cell [Ca2+]i increase, the faster and most sensitive effect of gp120 revealed in the brain so far, could be ultimately employed to reveal CD4‐independent transmembrane signalling machanisms of the viral protein that, at the moment, remain almost entirely unknown.
Exposure to gp120 of HIV‐1 Induces an Increased Release of Arachidonic Acid in Rat Primary Neuronal Cell Culture Followed by NMDA Receptor‐mediated Neurotoxicity
Authors: Ushijima Hiroshi , Nishio Osamu , Klöcking Renate , Perovic Sanja , Müller Werner E.G.
Published: April 2006

After incubation of highly enriched neurons from rat cerebral cortex with the HIV‐1 coat protein gp120 for 18 h, cells showed fragmentation of DNA at internucleosomal linkers followed by NMDA receptor‐mediated neurotoxicity. We report that in response to exposure to gp120 cells react with an increased release of arachidonic acid (AA) via activation of phospholipase A2. This process was not inhibited by NMDA receptor antagonists. To investigate the role of AA on the sensitivity of the NMDA receptor towards its agonist, low concentrations of NMDA were co‐administered with AA. This condition enhanced the NMDA‐mediated cytotoxicity. Administration of mepacrine reduced cytotoxicity caused by gp120. We conclude that gp120 causes an activation of phospholipase A2, resulting in the increased release of AA, which may in turn sensitize the NMDA receptor.
Differential Expression Patterns of Five Acetylcholine Receptor Subunit Genes in Rat Muscle During Development
Authors: Kues W. A. , Sakmann B. , Witzemann V.
Published: April 2006

The spatial and temporal expression patterns of five genes which encode the α‐, β‐, γ‐, δ‐ and ε‐subunits of the nicotinic acetylcholine receptor in skeletal muscle were followed during development in the rat by in situ hybridization analysis. Three major developmental phases, characterized by specific expression patterns, could be distinguished. (i) During myogenic differentiation α‐, β‐, γ‐ and δ‐subunit genes are activated and transcripts are expressed in muscle precursor cells at embryonic day 12 (E12) and during subsequent cell fusion. (ii) Following innervation of myotubes at ˜E15‐E17 the mRNA of the α‐, β‐, γ‐ and δ‐subunit genes accumulate in synaptic and decrease in extrasynaptic fibre regions during early synaptogenesis. The mRNA of the δ‐subunit gene becomes detectable first in subsynaptic nuclei 2–3 days after innervation has occurred. (iii) During postnatal development α‐, β‐ and δ‐ subunit transcript levels are reduced predominantly in extrasynaptic fibre segments and show significant differences in distribution depending on the muscle subtype whereas the γ‐subunit mRNA disappears completely within the first postnatal week in all muscles. In contrast, the γ‐subunit gene is transcribed only in subsynaptic myonuclei throughout development and in the adult muscle.
Physiological and Morphological Heterogeneity of Dentate Gyrus—Hilus Interneurons in the Gerbil Hippocampus In Vivo
Authors: Buckmaster Paul S. , Schwartzkroin Philip A.
Published: April 2006

A variety of morphological types of dentate gyrus/hilus interneurons have been described, but little is known about their corresponding physiological characteristics. To address this issue, intracellular responses to current injection and perforant path stimulation were obtained from putative dentate interneurons in anaesthetized adult gerbils. Our sample of interneurons showed heterogeneity in their intrinsic physiological characteristics and spike thresholds to perforant path stimulation, suggesting the existence of distinct physiologically‐defined classes. ‘Fast‐spiking’interneurons had a low threshold to perforant path stimulation, whereas ‘slow‐spiking’interneurons responded with predominantly inhibitory potentials. In several cases, cells were intracellularly labelled with biocytin for visualization. Interneurons with different physiological traits had distinct morphological features. These results confirm that, as in hippocampus proper, morphologically identifiable interneurons in the dentate hilus show electrophysiological features that are likely to reflect functionally specific roles in informational processing.
Flip and Flop Variants of AMPA Receptors in the Rat Lumbar Spinal Cord
Authors: Tölle T. R. , Berthele A. , Zieglgänsberger W. , Seeburg P. H. , Wisden W.
Published: April 2006

The expression of eight messenger RNA splice forms encoding the Flip and Flop variants of AMPA receptor subunits GluR‐A to ‐D in the rat lumbar spinal cord was examined by in situ hybridization using specific oligonucleotides. In the dorsal horn (laminae I, II and III) the predominant mRNA was GluR‐B Flip. Much lower levels of GluR‐A Flip were found in lamina I and in superficial parts of lamina II outer. In the ventral horn, motor neurons expressed mainly GluR‐B Flip, GluR‐C Flip and Flop, and GluR‐D Flip. Serial sectioning through large motor neurons indicated that a given cell contained, for example, both GluR‐C Flip and Flop splice types.
The Neurotrophin Receptors TrkA and TrkB are Inhibitory for Neurite Outgrowth
Authors: Tannahill Lesley , Klein Rüdiger , Schachner Melitta
Published: April 2006

To investigate the possibility that the neurotrophin tyrosine kinase receptors are also recognition molecules by virtue of their immunoglobulin‐like domains, the ability of TrkA and TrkB to influence neurite outgrowth was tested in vitro. Cell monolayers of fibroblasts transfected to express either the TrkA or TrkB receptor reduced neurite outgrowth of phaeochromocytoma PC12 cells by 50–60% when compared to mock transfected fibroblasts or fibroblasts transfected with the epidermal growth factor receptor. Neurite outgrowth from cerebellar neurons was inhibited by 30–40% on these substrates. When a recombinantly expressed fragment of TrkA comprising the two immunoglobulin‐like domains was coated as a substrate in combination with poly‐L‐lysine and laminin, neurite outgrowth was inhibited in a dose‐dependent manner. This inhibition of neurite outgrowth was not mediated via an interaction with laminin as there is no specific binding of the TrkA fragment to laminin. The adhesion of cell bodies to this substrate was not affected by the immunoglobulin‐like domains. These observations suggest that the mammalian neurotrophin receptors not only influence neurite outgrowth by neurotrophin triggered activation of the receptor, but also by cell surface recognition processes conveyed by the immunoglobulin‐like domains.
Target Preference of Embryonic Retina Cells and Retinal Cell Lines is Cell‐autonomous, Position‐specific, Early Determined and Heritable
Authors: Pollerberg G. E. , Eickholt B. J.
Published: April 2006

Retinal ganglion cells (RGCs) form the topographic connection between retina and optic tectum in the developing avian embryo. In vitro, neurons with the morphological traits and marker expression of RGCs were found both in single‐cell cultures from embryonic day (E) 6 chick retina and in retinal cell lines derived from E3.5 quail retina. Rapid and substantial differentiation of RGC‐like cells could be induced in the lines by addition of fibroblast growth factor aFGF or bFGF. RGC‐like cells were examined with respect to their target discrimination properties as single cells in the stripe carpet assay. In this assay system, alternating stripes of membrane vesicles prepared from the anterior and posterior tectum are offered to growing axonal processes as a substrate. Temporal RGC‐like cells, both primary cells prepared from the temporal retina and immortalized cells of those retinal lines derived from the temporal retina, avoid stripes of membrane vesicles from posterior tectum; they prefer to grow on membrane vesicles from the anterior tectum, which is their in vivo target. Nasal RGC‐like cells did not exhibit a target preference, in accordance with previous findings. Together the experiments show that target preference of RGCs is a cell‐autonomous and heritable mechanism that is determined early and is position‐dependent.
Behaviour of Small Inhibitory Interneurons in Early Postnatal Mouse Cerebellar Microexplant Cultures: A Video Time‐lapse Analysis
Authors: Magyar‐Lehmann Stefanie , Suter Caterina Savi , Stahel Werner , Schachner Melitta
Published: April 2006

The aim of this work was to investigate how the environment of the neuropil determines the positioning and differentiation of neurons that are postsynaptic to them. We investigated how stellate and basket cells, the small inhibitory interneurons of the cerebellar cortex, find their perpendicular orientation to the direction of fasciculated granule cell axons. Cultures of early postnatal mouse cerebellar microexplants showing this cellular behaviour in vitro were analysed by video time‐lapse cinematography and evaluated by morphometry. The small interneurons were first detectable when they migrated, intermingled with granule cells, away from the explant along the radial fascicles of granule cell neurites. During migration some cells suddenly changed their orientation by extending neurites in perpendicular orientation to the radial fascicles. These cells were all GABA‐immunoreactive and expressed the cytoskeletal markers tau in the thin axon‐like process and MAP2 in the thicker dendrite‐like arborizations at the opposite pole of the cell body. After having translocated in perpendicular orientation, these neurons were again able to turn back to move along the radial neurite bundles to another position. Furthermore, while in perpendicular orientation, the processes of these cells repelled each other upon contact of their growth cones, leading to equal spacing between the cell bodies with time in culture.
Fasciculation of Granule Cell Neurites is Responsible for the Perpendicular Orientation of Small Inhibitory Interneurons in Mouse Cerebellar Microexplant Cultures In Vitro
Authors: Magyar‐Lehmann Stefanie , Frei Thomas , Schachner Melitta
Published: April 2006

To study the cellular and molecular mechanisms involved in the perpendicular orientation of stellate and/or basket cells to the direction of fasciculating granule cell neurites, we have used cultures of microexplants from early postnatal mouse cerebellum that show this cellular behaviour in vitro. When these cultures were maintained in the presence of antibodies to the neural cell adhesion molecules L1 and N‐CAM or immunoglobulin‐like domains I, II and IV of N‐CAM, the resulting decrease or increase in the fasciculation of granule cell neurites changed the perpendicular orientation and morphology of the small inhibitory interneurons. Additives which did not perturb fasciculation did not affect the perpendicular orientation and morphology of stellate and/or basket cells. Furthermore, when perturbation of fasciculation was prevented, neither L1 nor N‐CAM antibodies modified the positioning or morphology of interneurons. These observations indicate that ordered fasciculation of granule cell neurites is an important parameter in the perpendicular orientation and elaboration of the typical morphology of the small cerebellar inhibitory interneurons.
Expression of the mRNA of Seven Metabotropic Glutamate Receptors (mGluR1 to 7) in the Rat Retina. An In Situ Hybridization Study on Tissue Sections and Isolated Cells
Authors: Hartveit E. , Brandstätter J. H. , Enz R. , Wässle H.
Published: April 2006

We have studied the expression of mRNAs for seven metabotropic glutamate receptors (mGluR1–7) in the retina of the adult rat by in situ hybridization with tissue sections and isolated cells using [α35S]dATP‐labelled oligonucleotide probes. Hybridization revealed the expression of six of the metabotropic receptor mRNAs, mGluR1, 2 and 4–7, in the retina, while mGluR3 was not detected. Each of the expressed receptor mRNAs showed a distinct pattern of expression. In the outer nuclear layer, corresponding to photoreceptor somata, no labelling was detected. In the outer part of the inner nuclear layer, putative horizontal cells were labelled for mGluR5. More proximal in this layer, corresponding to the position of bipolar cell somata, there was strong labelling for mGluR6. A small number of bipolar cells were also labelled for mGluR5 and mGluR7. In situ hybridization with isolated cells showed that mGluR6 was expressed by rod bipolar cells. Subsets of amacrine cells, with cell bodies along the border between the inner nuclear layer and the inner plexiform layer, were positive for mGluR1, 2, 4 and 7, suggesting considerable heterogeneity of these receptors among amacrine cells. None of the seven metabotropic receptor mRNAs was expressed in isolated Müller glial cells. In the ganglion cell layer, virtually every ganglion cell and displaced amacrine cell was labelled for mGluR1 and mGluR4. Some cells in this layer (˜20% of the total), most likely both ganglion cells and displaced amacrine cells, were also labelled for mGluR2 and mGluR7. These findings suggest that metabotropic glutamate receptors are considerably more widespread among neurons in the retina than indicated by previous physiological and pharmacological investigations.
Immunocytochemical Localization of trkA Receptors in Chemically Identified Subgroups of Adult Rat Sensory Neurons
Authors: Averill S. , McMahon S. B. , Clary D. O. , Reichardt L. F. , Priestley J. V.
Published: April 2006

Immunocytochemistry has been used to examine the location of trkA, the high‐affinity receptor for nerve growth factor, in adult rat dorsal root ganglia, trigeminal ganglia and spinal cord. TrkA immunoreactivity was observed in small and medium sized ganglion cells and in the dorsal horn of the spinal cord. In lumbar L4 and L5 ganglia trkA‐immunoreactive cells constitute 40% of dorsal root ganglion cells and range in size from 15 to 45 μm in diameter. Double labelling using markers for various dorsal root ganglion subpopulations revealed that virtually all (92%) trkA‐immunoreactive cells express calcitonin gene‐related peptide (CGRP) immunoreactivity. In contrast only 4 and 13% of trkA‐immunoreactive cells are labelled by the monoclonal antibody LA4 or the lectin Griffonia simplicifolia IB4, markers for small non‐peptide‐containing cells. Eighteen percent of trkA‐immunoreactive cells belong to the ‘large light’subpopulation, identified by their strong immunostaining by the neurofilament antibody RT97. TrkA immunoreactivity in the dorsal horn is heaviest in laminae I and II outer, has a similar distribution to CGRP, and is depleted by dorsal rhizotomy. Our results show that trkA‐expressing cells in dorsal root ganglia correspond almost exactly with the CGRP, peptide‐producing population. The receptor is present not only on cell bodies but also on central terminals. Non‐peptide‐containing small cells, which constitute 30% of dorsal root ganglion cells, are not trkA‐immunoreactive and therefore most probably are functionally independent of nerve growth factor.
Expression of GABA Receptor ρ1 and ρ2 Subunits in the Retina and Brain of the Rat
Authors: Enz Ralf , Brandstätter Johann Helmut , Hartveit Espen , Wässle Heinz , Bormann Joachim
Published: April 2006

We have investigated the distribution of GABA receptor ρ1 and ρ2 subunits in the rat central nervous system. Cloning of rat ρ1 and ρ2 cDNA fragments revealed similarities to the corresponding human sequences of 99% (ρ1) and 88% (ρ2) at the protein level. Whereas the human ρ2 subunit has no consensus sequence for phosphorylation by protein kinase C, the cytoplasmic loop of the rat sequence contains two such sites. Use of the polymerase chain reaction with reverse‐transcribed total RNA (RT‐PCR) from different brain tissues revealed that transcript for the ρ1 subunit was present in the retina only. The ρ2 mRNA was detected in all brain regions, with the highest level of expression in the retina.In situhybridization of retinal sections revealed that ρ1 and ρ2 transcripts are present in the inner nuclear layer. RT‐PCR andin situhybridization of isolated retinal cells showed that both ρ subunits are present in rod bipolar cells. Since these cells express bicuculline‐insensitive GABA receptors, our results further support the idea that ρ subunits are part of the GABACreceptor.
BDNF Protein Measured by a Novel Enzyme Immunoassay in Normal Brain and after Seizure: Partial Disagreement with mRNA Levels
Authors: Nawa Hiroyuki , Carnahan Josette , Gall Christine
Published: April 2006

Messenger RNA for brain‐derived neurotrophic factor (BDNF) is distributed in many brain regions and regulated by excitatory neuronal activity. Despite numerous studies of BDNF mRNA, the distribution and regulation of BDNF protein are poorly understood because of the difficulty of its quantitative measurement. We have established a two‐site enzyme immunoassay that detects trace amounts of BDNF protein (>1 pg/assay) but not other neurotrophins or growth factors. The highest levels of BDNF in adult rat brain were found in the hippocampus, followed by the hypothalamus, neocortex, cerebellum, thalamus and striatum. This pattern is similar, but not identical, to the distribution of BDNF mRNA. A similar disparity between BDNF protein and mRNA levels was observed in their changes after hilus lesion‐induced limbic seizures. In limbic structures, BDNF concentrations remained elevated 4 days after seizure onset, whereas BDNF mRNA has been reported previously to return to basal levels within 46 h. The temporal and spatial differences between the dynamics of protein and mRNA levels suggest the importance of post‐translational and/or subcellular processes for BDNF production. The persistence of the increases in BDNF content was also reflected in its biological activity, e.g. peptidergic differentiation activity. After limbic seizures, neuropeptide Y content was most markedly and persistently elevated in the entorhinal/amygdaloid region, where the most sustained up‐regulation of BDNF protein was observed. These results suggest that the sustained increase of BDNF protein in these limbic structures is involved in prolonged post‐seizure phenomena, including peptidergic alterations.
Properties of K+ Conductances in Cat Retinal Ganglion Cells During the Period of Activity‐mediated Refinements in Retinofugal Pathways
Authors: Skaliora I. , Robinson D. W. , Scobey R. P. , Chalupa L. M.
Published: April 2006

During ontogeny retinal ganglion cells manifest pronounced changes in excitable membrane properties. To further our understanding of the ionic conductances underlying such functional changes, the whole‐cell voltage‐clamp variation of the patch‐clamp technique was used to record potassium currents in 220 ganglion cells dissociated from cat retinas ranging in age from embryonic day 31 to postnatal day 10. Potassium currents were isolated by blocking voltage‐gated Na+ and Ca2+ currents with tetrodoxin (TTX) and CoCl2 respectively and were characterized by their pharmacology, kinetics and voltage‐dependence of activation and inactivation. In all cases, a combination of three currents accounted for the total outward calcium‐independent K+ current: (i) a steady linear conductance; (ii) a voltage‐gated transient current, lA, and (iii) a voltage‐gated sustained current, lk. Both voltage‐gated currents were affected by the application of 4‐aminopyridine and tetraethylammonia (TEA): lA showed a greater sensitivity to 4‐aminopyridine, while lk was more sensitive to TEA. Both voltagegated currents were present throughout the developmental period examined; however, the percentage of retinal ganglion cells (RGCs) expressing lA showed a marked decline from 82% at E31 to 45% at postnatal ages. During this developmental period there was an increase in the density of the two voltage‐gated and the linear conductance. Additionally, with maturation, significantly slower inactivation kinetics were observed for lK. These findings, and our previous results dealing with maturational changes in the TTX‐sensitive voltage‐gated Na current, are related to the generation of excitability in developing retinal ganglion cells. Furthermore, the presence of cells with and without transient K+ conductance throughout development suggests that the different spiking patterns observed in RGC classes may be partially due to differences in their membrane properties.
Calcium Deposit Formation and Glial Reaction in Rat Brain after Ibotenic Acid‐induced Basal Forebrain Lesion
Authors: Saura J. , Boatell M. L. , Bendahan G. , Mahy N.
Published: April 2006

The mechanisms underlying amino acid neurotoxicity may involve a rise in the intracellular concentration of calcium. Some neurons appear to die as a consequence of increased intracellular calcium levels induced by excitatory amino acids. One month after injection of ibotenic acid in the rat basal forebrain, the induced formation of calcium deposits and concomitant glial reaction were studied. Alizarine Red‐positive calcium deposits were observed after ibotenic acid injection in the ventral part of the globus pallidus, but not in the medial septum. These deposits were present in the globus pallidus, ventral pallidum, substantia innominata, zona incerta, lateral hypothalamic area, entopeduncular nucleus, medial amygdaloid nucleus and several thalamic nuclei. Three types of round shaped deposit were identified morphologically. Differential astroglial and microglial reactions, studied autoradiographically with the monoamine oxidase‐B marker [3H]Ro 19‐6327 and the peripheral benzodiazepine receptor marker [3H]Ro5‐4864 respectively, were observed after both lesions. Our data suggest that excitotoxic lesions in the globus pallidus and medial septum lead with time to different neurodegenerative consequences and glial reactions. This differential sensitivity is discussed on the basis of the presence of different glutamate receptor subtypes and calcium‐binding proteins.
The Rate of Wallerian Degeneration in Cultured Neurons from Wild Type and C57BL/WldS Mice Depends on Time in Culture and may be Extended in the Presence of Elevated K+ Levels
Authors: Buckmaster E. A. , Perry V. H. , Brown M. C.
Published: April 2006

Wallerian degeneration of severed axons is delayed in C57BL/Wlds mice. We have examined this further in cultured sympathetic, sensory and CNS neurons using superior cervical ganglion (SCG), dorsal root ganglion (DRG) and cerebellar granule neurons respectively from neonatal mice. We found that the time taken for the neurites to degenerate depends upon the length of time in culture before cutting, reaching a maximum by ‐7 days when C57BL/WldS neurites survive for >6 days after axotomy. The onset of degeneration could also be extended in SCG and DRG neurites from wild type C57BL/6J mice. After 7 days in culture these neurites normally degenerate within ‐12–16 h of axotomy, but in the presence of raised K+ (50 mM) degeneration often did not begin until a further 2 days had lapsed. Under similar conditions degeneration of neurites from C57BL/WldS mice was also found to be further delayed, extending survival from ‐5–6 days to >7 days. The L‐type Ca2+ channel blockers nifedipine (5 μM) and verapamil (10 μM) both blocked the effect of raised [K+], although not completely. Thapsigargin, which raises cytoplasmic [Ca2+], and the cAMP analogue 8‐(4‐chlorophenylthio)cAMP were also able to delay degeneration, but only when added 24 h prior to axotomy. These results show that it is possible to influence the course of Wallerian degeneration and that increases in levels of cytoplasmic Ca2+ can protect neurites from its onset.
Persistence of Neuromuscular Junctions after Axotomy in Mice with Slow Wallerian Degeneration (C57BL/Wlds)
Authors: Ribchester R. R. , Tsao J. W. , Barry J. A. , Asgari‐Jirhandeh N. , Perry V. H. , Brown M. C.
Published: April 2006

The present study was undertaken to examine the fate of neuromuscular junctions in C57BL/Wlds mice (formerly known as OLA mice) after nerve injury. When a peripheral nerve is injured, the distal axons normally degenerate within 1‐3 days. For motor axons, an early event is deterioration of motor nerve terminals at neuromuscular junctions. Previously, the vulnerability of motor terminals has been attributed either to a ‘signal’originating at the site of nerve injury and transported rapidly to the terminals or to their continual requirement for essential maintenance factors synthesized in the motor neuron cell body and supplied to the terminals by fast axonal transport. Mice of the Wlds strain have normal axoplasmic transport but show an abnormally slow rate of axon and myelin degeneration. Structure and function are retained in the axons of distal nerve stumps for several days or even weeks after nerve injury in these mice. The results of the present study show that Wlds neuromuscular junctions are also preserved and continue to release neurotransmitter and recycle synaptic vesicle membrane for at least 3 days and in some cases up to 2 weeks after nerve injury. Varying the site of the nerve lesion delayed degeneration by ‐1‐2 days per centimetre of distal nerve remaining. These findings suggest that the mechanisms of nerve terminal degeneration after injury are more complex than can be accounted for simply by the failure of motor neuron cell bodies to supply their terminals with essential maintenance factors. Rather, the data support the view that nerve section normally activates cellular components or processes already present, but latent, in motor nerve endings, and that in Wlds mice either the trigger or the cellular response is abnormal.
Neuroanatomy of Cardiac Activity‐regulating Circuitry: A Transneuronal Retrograde Viral Labelling Study in the Rat
Authors: Ter Horst Gert J. , Hautvast Raymond W. M. , De Jongste Mike J. L. , Korf Jakob
Published: April 2006

The anatomy of cardiac activity‐regulating circuitry was studied with retrograde transneuronal viral labelling after pseudorabies virus injections into different parts of the rat heart. Transection of the spinal cord at Th1 was used to reveal selectively the parasympathetic neuronal networks. Virus‐labelled sympathetic preganglionic cells were found in the Th1‐Th7 thoracic intermediolateral cell groups, with some additional infections at Th8‐Thll after inoculations of the ventricular myocardium. After ventricular injections the thoracic spinal labelling pattern was bilateral and after right atrial infection it was contralateral. Approximately 20% of the parasympathetic preganglionic cells were located in the dorsal motor vagus nucleus; the rest occupied positions in the peri‐ambiguus area ventrolateral to the nucleus ambiguus. Here and in the ventrolateral reticular formation myocardiotopy was found. Supraspinal transneuronal infections were bilateral, showed no apparent side dominance and were found in the nucleus of the solitary tract, the area postrema, the raphe nuclei, the A5 group, the parabrachial region, the periaqueductal grey, the hypothalamus, the amygdala and the cortex, in particular the anterior cingulate, the frontal, prelimbic, infralimbic and insular cortices. Spinal transections at Th1 reduced the number of labelled cells, gave a right side labelling dominance and affected the infection patterns in the ventrolateral reticular area, the raphe nuclei, the periaqueductal grey matter, the perifornical and retrochiasmatic area and the rostra1 parts of the insular cortex. The latter structures are linked selectively to the sympathetic innervation of the heart. The anatomical and functional aspects of these findings are discussed in relation to the autonomic control of heart activity.
Dopamine Agonists Increase Nitric Oxide Production in the Paraventricular Nucleus of the Hypothalamus: Correlation with Penile Erection and Yawning
Authors: Melis Maria Rosaria , Succu Salvatora , Argiolas Antonio
Published: April 2006

A low dose of apomorphine (80 μg/kg s.c.), a mixed D1/D2 agonist that induces penile erection and yawning, increased the concentration of NO2−from 1.12 ± 0.45 μM to 3.8 ± 0.75 μM and NO3−from 5.53 ± 0.82 to 11.25 ± 2.30 μM in the dialysate collected from the paraventricular nucleus of the hypothalamus of male rats byin vivomicrodialysis. The NO2−concentration was also increased by LY 171555 (50 μg/kg s.c.), a D2 agonist that induces penile erection and yawning, but not by SKF 38393 (5 mg/kg s.c.), a D1 agonist with no effect on these responses. Conversely, apomorphine's effect on NO2−was prevented by haloperidol (0.5 mg/kg i.p.), a mixed D1/D2 antagonist and L‐sulpiride (25 mg/kg i.p.), a D2 antagonist, but not by the D1 agonist SCH 23390 (50 μg/kg s.c.), although all three compounds prevented penile erection and yawning. The apomorphine effect on NO2−, penile erection and yawning was also prevented by the nitric oxide synthase inhibitorNG‐nitro‐L‐arginine methyl ester (200 μg l.c.v.). The nitric oxide scavenger haemoglobin (200 μg l.c.v.) also prevented the NO2−increase, but was ineffective against penile erection and yawning. In contrast, the oxytocin antagonist d(CH2)5Tyr(Me)‐Orn8‐vasotocin (1 μg i.c.v.) and the guanylate cyclase inhibitor methylene blue (300 μg l.c.v.) had no effect on the NO2−increase, but did prevent the behavioural responses. We infer from this that dopamine agonists induce penile erection and yawning by acting on D2 receptors that increase nitric oxide synthase activity in the cell bodies of paraventricular oxytocinergic neurons projecting to extra‐hypothalamic brain areas.
Kindled Seizure‐induced c‐fos and Prodynorphin mRNA Expressions are Unrelated in the Rat Brain
Authors: Simonato M. , Bregola G. , Donatini A. , Bianchi C. , Beani L. , Ferri S. , Romualdi P.
Published: April 2006

Levels of mRNA for c‐fos and prodynorphin were studied by in situ hybridization in adjacent coronal sections taken from kindled rats 30–60 min after the last seizure. Within this time frame, expression of both genes was induced in multiple brain areas. Anatomical colocalization of the induced gene expressions was found in the hippocampus. Induction of c‐fos in the dentate gyrus was bilateral and symmetrical in a subgroup of rats, ipsilateral in another subgroup and absent in a third subgroup. However, no relative increase was observed in the ipsilateral compared with the contralateral prodynorphin expression in the dentate gyrus when c‐fos expression was induced ipsilaterally only. These observations suggest that, at variance with other experimental situations, Fos is not involved in the mechanisms of kindled seizure‐induced activation of prodynorphin transcription in the rat forebrain.
Cyclic Nucleotide‐gated Channels in Identified Human Olfactory Receptor Neurons
Authors: Thüraüf N. , Gjuric M. , Kobal G. , Hatt H.
Published: April 2006

Patch‐clamp recordings revealed the presence of a non‐desensitizing cyclic nucleotide‐gated channel on human olfactory receptor neurons and a fast‐desensitizing non‐specific cation channel activated by nucleotides on human supporting cells. Cyclic nucleotide‐gated channels on olfactory receptor neurons showed selective channel activation by cAMP (K1/2= 5 μM) and cGMP (K1/2= 2 μM), a unitary conductance of ∼20 pS, a reversal potential of single‐channel currents close to 0 mV, a linear current‐voltage relationship over the range of −80 to 80 mV and a strong extracellular but a weaker intracellular blocking effect of Ca2+. The channel activity outlasted the cyclic nucleotide pulses for hundreds of milliseconds when higher agonist concentrations (>50 μM cAMP) were applied. The duration of the response was longer than in cyclic nucleotide‐gated channels from other species studied so far. The plateau duration and the decay remained constant for pulses with a length of 50−150 ms, whereas pulses shorter than 50 ms successively reduced the time required by shortening the plateau phase. A larger difference for the K1/2 values of cAMP (K1/2= 22 μM) and cGMP (K1/2= 2.5 μM) were found for a small group (n = 3) of cyclic nucleotide‐gated channels, pointing to the selective expression of the a‐subunit in a small subgroup of olfactory receptor neurons.
Interaction between Low Voltage‐activated Currents in Reticular Thalamic Neurons in a Rat Model of Absence Epilepsy
Authors: Meis Susanne , Biella Gerardo , Pape Hans‐Christian
Published: April 2006

A transient potassium (K+) outward current (IA) contributes to the distinctive patterns of low‐threshold spike firing observed in various classes of thalamic neurons through a functional interaction with a calcium (Ca2+)‐mediated inward current (IT). The present study was undertaken to investigate the properties of transient K+ currents and their interaction with IT in neurons of the reticular thalamic nucleus, and to compare these properties in reticular thalamic nucleus neurons from a rat model of absence epilepsy, designated the Genetic Absence Epilepsy Rat from Strasbourg (GAERS), with those from a Non‐epileptic Control strain (NEC). This comparative approach appeared to be particularly important in view of the recent finding of a selective increase in IT in reticular thalamic nucleus neurons from GAERS. Neurons were acutely isolated from the reticular thalamic nucleus through enzymatic procedures, and identified by morphological and immunocytochemical criteria. Ionic currents were analysed using whole‐cell patch‐clamp techniques. Transient K+ currents in reticular thalamic nucleus neurons with properties indicative of IA activated at ∼−55 mV (with half‐activation at −27 and −33 mV in NEC and GAERS respectively), declined rapidly with a voltage‐dependent time constant (τ= 4 ms at +45 mV), were 50% steady‐state‐inactivated at −81 and −86 mV in the two strains of rats respectively, and rapidly recovered from inactivation with a monoexponential time course (τ= 31 and 37 ms respectively). No significant differences in IA properties or densities were found between reticular thalamic nucleus neurons from GAERS and NEC rats. Analysis of the interaction between IA and IT indicated a shift in the balance between the two opposing membrane conductances towards the generation of a low‐voltage‐activated inward current in reticular thalamic nucleus neurons from GAERS compared with NEC, and a lack of IA to functionally compensate for this shift, which in turn may contribute to pathological forms of low‐threshold spike firing characterizing spike‐and‐wave discharges.
Interaction Between α and β‐Adrenergic Receptor Agonists Modulating the Slow Ca2+‐activated K+ Current ‐ IAHP in Hippocampd Neurons
Authors: Pedarzani Paola , Storm Johan F.
Published: April 2006

Noradrenaline inhibits the Ca2+‐activated K+ current IAHP, which underlies the slow afterhyperpolarization and spike frequency adaptation in hippocampal and neocortical neurons. The resulting increase in excitability probably contributes to the state control of the forebrain during arousal and attention. The modulation of IAHP by noradrenaline has previously been shown to be mediated by β1 receptors, cyclic AMP and protein kinase A, but not by α receptors. We have now tested the possibility that a receptors also contribute to IAHP modulation through interaction with β receptors, by the use of whole‐cell recordings in CA1 pyramidal cells of rat hippocampal slices. The α‐receptor agonist 6‐fluoro‐noradrenaline strongly potentiated the effect of isoproterenol on IAHP. The synergistic effect of 6‐fluoro‐noradrenaline and soproterenol was blocked by the β‐receptor antagonist timolol, but the receptor type mediating the effect of 6‐fluoro‐noradrenaline could not be unequivocally identified by using α‐receptor antagonists. The effect of high concentrations of noradrenaline on IAHP was only partly blocked by the β‐receptor antagonist timolol, and was further reduced by blocking α receptors, again suggesting a contribution from α receptors. In contrast, the effect of low concentrations of noradrenaline seemed to be potentiated by the α‐receptor antagonist phentolamine in 57% of the cells, suggesting concentration dependent antagonistic interaction between α and β receptors. Further tests indicated that the cross‐talk between 6‐fluoro‐noradrenaline and isoproterenol occurs upstream from cyclic AMP production, and that protein kinase A serves as a final common path for the modulation of IAHP by noradrenaline, and by the combination of 6‐fluoro‐noradrenaline and isoproterenol.
Expression of the Transcription Factor Zif268 in the Temporal Cortex of Monkeys during Visual Paired Associate Learning
Authors: Okuno Hiroyuki , Miyashita Yasushi
Published: April 2006

In humans and monkeys, memory consists of various components which were initially revealed through neuropsychological studies of amnesic patients. One memory component, the long‐term memory about facts and events (declarative memory), has been shown to require the integrity of the medial temporal lobe and the neocortex. To investigate a map of gene expression during declarative memory formation in the primate, we monitored expression of immediate early genes in the temporal lobe of macaque monkeys. We trained the monkeys to learn visual associative memory tasks, and immunohistochemically analysed the expression of protein products of immediate early genes. We found that Zif268, a transcription factor regulated by neuronal activity, accumulated in patches in the anterior temporal lobe during visual stimulus‐stimulus association learning, whereas other transcription factors, c‐Fos and JunD, were not. By contrast, the patchy expression of Zif268 was not observed during another type of learning, visual discrimination learning. These results suggest that Zif268 activates a gene cascade related to the formation of long‐term memory in the primate.
Comparative Antagonism of Kainate‐activated Kainate and AMPA Receptors in Hippocampal Neurons
Authors: Paternain Ana V. , Vicente Angeles , Nielsen Elsebet Ø. , Lerma Juan
Published: April 2006

Native kainate receptors expressed by cultured hippocampal cells were studied in the whole‐cell configuration of the patch‐clamp technique by using a fast perfusion system. About 80% of the neurons expressed kainate receptors independently of the time in culture (0–4 days), which coincided with the number of cells immunoreactive for a monoclonal antibody against the GluR5/6/7 subunits. Three types of cells were considered: neurons in which the rapid application of kainate induced a rapidly desensitizing current, cells in which kainate induced a more slowly rising, non‐desensitizing, response and those in which a mixture of both responses was apparent. Steady responses induced by 300 μM kainate were inhibited by 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (CNQX) in a dose‐dependent manner (IC50= 0.92 μM). CNQX was less potent in blocking transient kainite‐induced responses (IC50= 6.1 μM). Responses to kainate, whether steady or transient, were also inhibited by NS102, showing poor selectivity for the transient response (IC50= 4.1 and 2.2 μM respectively). The new α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole (AMPA) receptor antagonist NS394 was very potent in inhibiting steady kainate‐induced currents (IC50= 0.45 μM), but was even more effective in preventing peak responses (IC50= 0.13 μM). In contrast, cyclothiazide did not affect transient kainate‐induced responses but did potentiate current induced by activation of AMPA receptors by AMPA or kainate. These results demonstrate the lack of complete selectivity amongst some available competitive antagonists for AMPA and kainate receptors, and indicate that kainate receptors expressed by hippocampal cells lack the cyclothiazide modulatory site present at AMPA receptors. In addition, the present data support the idea that low‐affinity kainate binding sites in the brain correspond to receptor channels selectively activated by kainate.
Effects of Glutamate Application on the Rhythm of Low Magnesium‐induced Epileptiform Activity in Hippocampal Slices of Guinea‐pigs
Authors: Lücke A. , Köhling R. , Speckmann E.‐J.
Published: April 2006

The extracellular concentration of glutamate has previously been reported to increase to more than 10‐fold the basal level during seizure activity. In the present study, we tested whether localized increases in extracellular glutamate concentration influence the rhythm of epileptiform discharges in the low‐magnesium epilepsy model. In hippocampal slices of guinea‐pigs, epileptiform activity was induced by omission of magnesium from the bath fluid. Glutamate and its subreceptor agonists N‐methyl‐D‐aspartate (NMDA) and α‐amino‐3‐hydroxy‐5‐methyl‐4‐ isoxazolepropionic acid (AMPA) were ejected into different strata of the CA3 and CA1 regions using microiontophoretic and micropressure application. Glutamate, NMDA and AMPA applied to the CA3 region, but not to the CA1 region, induced a short‐lasting increase in epileptiform discharge frequency, often followed by a transient reduction. The effect was most pronounced with application into the stratum lacunosum‐moleculare of the CA3 region and could only be evoked in slices exceeding 400 μm in thickness. The effects on the rhythm of epileptiform discharges induced by NMDA and AMPA were blocked by their specific receptor antagonists. They were not influenced by application of GABAA and GABAB receptor antagonists. Changes in somatic membrane potential of CA3 pyramidal neurons did not correlate with changes in the rhythm of epileptiform discharges elicited in this region. The transient suppression of epileptiform discharges that followed the increase in discharge frequency was abolished by an adenosine A1 receptor antagonist. We propose that localized increases in extracellular glutamate concentration modify the rhythm of epileptiform discharges due to changes in neuronal network activity.
Group II and Group III Metabotropic Glutamate Receptors in the Rat Retina: Distributions and Developmental Expression Patterns
Authors: Koulen Peter , Malitschek Barbara , Kuhn Rainer , Wässle Heinz , Brandstätter Johann Helmut
Published: April 2006

We have studied the distributions of group II metabotropic glutamate receptors, mGluR2 and mGluR3, and a group III metabotropic glutamate receptor, mGluR4, in the adult rat retina and during postnatal development using receptor specific anti‐peptide antisera. Of the three receptors examined, mGluR3 was not expressed in the retina. MGluR2 showed a distinct stratification pattern in the inner plexiform layer (IPL). Double‐labelling immunocytochemistry revealed that mGluR2 was localized in the processes of cholinergic amacrine cells. MGluR4 was found throughout the entire IPL. At the subcellular level, both mGluR2 and mGluR4 were found to be localized exclusively in processes postsynaptic to bipolar cell synapses in the IPL. During postnatal development, labelling for mGluR2 was detected at around postnatal day five. MGluR4 was already present at postnatal day one, prior to the establishment of synaptic connections in the IPL. The differential expression patterns of individual metabotropic glutamate receptors in the adult and developing rat retina suggest distinct roles for these receptors in retinal synaptic circuitry.