bluuebunny/crossref_metadata_2025 huggingface dataset:
Neuronal distribution and functional characterization of the calcium channel α2δ‐2 subunit
Authors: Hobom M. , Dai S. , Marais E. , Lacinova L. , Hofmann F. , Klugbauer N.
Published: March 2003

The auxiliary calcium channel α2δ subunit comprises a family of three genes, α2δ‐1 to 3, which are expressed in a tissue‐specific manner. α2δ‐2 mRNA is found in the heart, skeletal muscle, brain, kidney, liver and pancreas. We report here for the first time the identification and functional characterization of α2δ‐2 splice variants and their mRNA distribution in the mouse brain. The splice variants differ in the α2 and δ protein by eight and three amino acid residues, respectively, and are differentially expressed in cardiac tissue and human medullary thyroid carcinoma (hMTC) cells. In situ hybridization of mouse brain sections revealed the highest expression of α2δ‐2 mRNA in the Purkinje cell layer of the cerebellum, habenulae and septal nuclei, and a lower expression in the cerebral cortex, olfactory bulb, thalamic and hypothalamic nuclei, as well as the inferior and superior colliculus. As the in situ data did not suggest a specific colocalization with any α1 subunit, coexpression studies of α2δ‐2 were carried out either with the high‐voltage‐gated calcium channels, α1C, α1E or α1A, or with the low‐voltage‐gated calcium channel, α1G. Coexpression of α2δ‐2 increased the current density, shifted the voltage dependence of channel activation and inactivation of α1C, α1E and α1A subunits in a hyperpolarizing direction, and accelerated the decay and shifted the steady‐state inactivation of the α1G current.
Opioids intrinsically inhibit the genesis of mouse cerebellar granule neuron precursors in vitro: differential impact of μ and δ receptor activation on proliferation and neurite elongation
Authors: Hauser Kurt F. , Houdi Abdulghani A. , Turbek Carol S. , Elde Robert P. , Iii William Maxson
Published: March 2003

Although opioids are known to affect neurogenesis in vivo, it is uncertain the extent to which opioids directly or indirectly affect the proliferation, differentiation or death of neuronal precursors. To address these questions, the intrinsic role of the opioid system in neurogenesis was systematically explored in cerebellar external granular layer (EGL) neuronal precursors isolated from postnatal mice and maintained in vitro. Isolated neuronal precursors expressed proenkephalin‐derived peptides, as well as specific μ and δ, but negligible κ, opioid receptors. The developmental effects of opioids were highly selective. Morphine‐induced μ receptor activation inhibited DNA synthesis, while a preferential δ2‐receptor agonist ([d‐Ala2]‐deltorphin II) or Met‐enkephalin, but not the δ1 agonist [d‐Pen2, d‐Pen5]‐enkephalin, inhibited differentiation within the same neuronal population. If similar patterns occur in the developing cerebellum, spatiotemporal differences in endogenous μ and δ opioid ligand–receptor interactions may coordinate distinct aspects of granule neuron maturation. The data additionally suggest that perinatal exposure to opiate drugs of abuse directly interfere with cerebellar maturation by disrupting normal opioid signalling and inhibiting the proliferation of granule neuron precursors.
Evidence for nonvesicular nitric oxide release evoked by nerve activation
Authors: Olgart Caroline , Gustafsson Lars E. , Wiklund N. Peter
Published: March 2003

The gaseous nature of nitric oxide (NO) has led to the general assumption that its release from neurons during nerve stimulation is independent of vesicular storage. However, recent findings have shown that NO can exist intracellularly as part of more stable bioactive molecules, suggesting that the role of vesicular exocytosis for NO release cannot be excluded simply based on the chemical nature of NO itself. We have used botulinum toxin B (BTX B) to directly address the role of vesicular exocytosis for NO release. BTX B cleaves the synaptic vesicle protein synaptobrevin/VAMP, and by this inhibits Ca++‐mediated exocytic release of neurotransmitters. As a target organ we used the guinea‐pig enteric nervous system, which innervates the gastrointestinal tract, and in which both classical neurotransmitters as well as NO are released and influence smooth muscle activity. As expected, BTX B (0.1 μm) blocked the nerve stimulation‐induced cholinergic and tachykininergic smooth muscle contractions, and markedly inhibited the nerve stimulation‐evoked release of [3H]‐choline. In contrast, BTX B (0.1 μm) had no effect on nerve stimulation‐evoked relaxations, which were equally inhibited by an NO‐synthase inhibitor as well as by a selective inhibitor of soluble guanylyl cyclase. In addition, nerve stimulation‐evoked NO synthase‐dependent outflow of NO/NO2– was unaffected by BTX B (0.1 μm). These findings suggest that the neuronal release of endogenous NO is independent of intact synaptobrevin/VAMP, and therefore provide further evidence that nerve‐mediated release of further NO is nonvesicular.
The human eye expresses high levels of CB1 cannabinoid receptor mRNA and protein
Authors: Porcella Anna , Maxia Chiara , Gessa Gian Luigi , Pani Luca
Published: March 2003

We used reverse transcriptase polymerase chain reaction to detect the expression of the central and peripheral cannabinoid receptors (CB1 and CB2, respectively) mRNA, and Western blotting to show the presence of the CB1 protein in subregions of the human eye. CB2 mRNA transcripts were undetectable, while levels of CB1 mRNA were significantly expressed in the human retina (25.8 ± 2.46%), ciliary body (210 ± 11.55%) and iris (62.7 ± 5.94%) when compared with those of the normalizing reference gene β2 microglobulin. The CB1 gene encodes a functional protein which is detected in its glycosylated (63 kDa) and unglycosylated (54 kDa) form in the same areas by a specific purified antibody raised against the amino terminus (residues 1–77) of the CB1 receptor. These results further support the proposed role of the CB1 receptor in controlling intraocular pressure, helping to explain the antiglaucoma properties of marijuana.
Long‐term potentiation of primary afferent neurotransmission at trigeminal synapses of juvenile rats
Authors: Hamba Michiko , Onodera Kayoko , Takahashi Tomoyuki
Published: March 2003

Primary afferent monosynaptic and polysynaptic excitatory postsynaptic currents (EPSCs) were recorded from brainstem trigeminal neurons by stimulation of the mandibular nerve attached to the brainstem preparation of juvenile rats. A high‐frequency conditioning stimulus induced long‐term potentiation (LTP) of high‐threshold EPSCs in the majority of trigeminal caudal neurons in substantia gelatinosa, where both A‐ and C‐fibres terminate. However, the same conditioning stimulus did not potentiate low‐threshold EPSCs in caudal neurons or EPSCs recorded from neurons in the middle part of trigeminal interpolar nucleus, where C‐fibres rarely terminate. LTP in caudal neurons could be induced after blocking N‐methyl‐ d‐aspartate (NMDA) receptors with D(–)‐2‐amino‐5‐phosphonopentanoic acid (D‐AP5, 50 μm), after postsynaptic loading of the Ca2+ chelator BAPTA (10 mm), or even after completely blocking excitatory transmission with kynurenic acid during conditioning. However, LTP was blocked by the metabotropic glutamate receptor antagonist (+)‐α‐methyl‐4‐carboxyphenylglycine (1 mm). We suggest that LTP of the trigeminal primary afferent EPSCs is induced preferentially in the C‐fibre inputs and that the induction mechanism involves metabotropic glutamate receptors, possibly at the presynaptic terminals.
Correlation between gene functions and developmental expression patterns in the mouse cerebellum
Authors: Matoba Ryo , Kato Kikuya , Kurooka Chika , Maruyama Chiyuri , Sakakibara Yoshimasa , Matsubara Kenichi
Published: March 2003

Quantitative changes of 419 gene transcripts during postnatal mouse cerebellar development were accurately determined with a novel polymerase chain reaction (PCR)‐based technique. About 70% of the genes showed differences in expression levels, and the magnitude of difference was relatively small. By hierarchic cluster analysis of developmental expression patterns, the genes were categorized into 19 clusters, which were subsequently assembled into four major groups: group 1, with elevation of gene expression throughout the time course; group 2, with relatively unchanged levels; group 3, with transiently high expression at ∼ 12 days; and group 4, with highest expression at ∼ 4 days. Genes related to brain functions were segregated into several clusters of group 1 and group 3: the same clusters in which cerebellum‐specific genes were also segregated. Genes for protein synthesis belonged to group 4. Genes with housekeeping functions belonged to group 2. Western blotting analysis of representative protein products of each group revealed correlation with the mRNA level for those belonging to group 1 and group 4, but not necessarily in the other groups. The close correlation of algorithmically categorized temporal expression patterns of genes with their functions will be useful for estimating the functions of thousands of novel genes.
Layer‐specific immunocytochemical localization of GABABR1a and GABABR1b receptors in the rat piriform cortex
Authors: Princivalle Alessandra , Spreafico Roberto , Bowery Norman , De Curtis Marco
Published: March 2003

A peculiar, layer‐segregated immunoreactive distribution of GABABR1a and GABABR1b receptor antibodies is present in the piriform cortex of adult rats. The GABABR1a antibody selectively marked the neuropile in layer Ia, where afferent olfactory fibres and intrinsic GABAergic (γ‐aminobutyric acid) axons terminate on the distal apical dendrites of pyramidal neurons. The GABABR1b antibody was detected in the soma and the large basal dendrites of layer II and III neurons. The pattern of distribution observed supports the hypothesis that (presynaptic) GABABR1a receptors in the superficial molecular layer modulate neurotransmitter release in a feedforward synaptic circuit, whereas GABABR1b (postsynaptic) receptors mediate feedback inhibitory potentials on principal cells.
Orientation discrimination of objects and gratings compared: an fMRI study
Authors: Faillenot Isabelle , Sunaert Stefan , Van Hecke Paul , Orban Guy A.
Published: March 2003

We used functional magnetic resonance imaging to compare the human brain regions involved in orientation discrimination of two‐dimensional (2D) objects and gratings. The orientation discrimination tasks, identification and successive discrimination, were contrasted to a dimming detection control condition with identical retinal input. Regions involved in orientation discrimination were very similar for the two types of tasks and for the two types of stimuli and both belonged to the dorsal and ventral visual pathways. They included posterior occipital, lingual, posterior fusiform, inferior temporal, dorsal intraparietal and medial parietal regions. The main difference between the two types of stimuli was a larger activation of precuneus when 2D objects were used compared to gratings. The main difference between discrimination tasks was an enhanced activity, at the group level, in superior frontal sulcus in identification compared to successive discrimination, and at least at the single subject level, a larger activity in right fusiform cortex in successive discriminations compared to identification. Thus, in contradiction to generally accepted views, orientation discrimination of gratings and objects involve largely similar networks including both ventral and dorsal visual regions.
Differential expression of fibroblast growth factor‐2 and fibroblast growth factor receptor 1 in a scarring and nonscarring model of CNS injury in the rat
Authors: Smith Conrad , Berry Martin , Clarke Wendy E. , Logan Ann
Published: March 2003

Injury to the adult brain results in abortive axon regeneration and the deposition of a dense fibrous glial scar. Therapeutic strategies to promote postinjury axon regeneration are likely to require antiscarring strategies. In neonatal brain wounds, scar material is not laid down and axons grow across the lesion site, either by de novo growth or regeneration. To achieve the therapeutic goal of recapitulating the nonscarring neonatal response in the injured adult, an understanding of how ontogenic differences in scarring reflect developmental diversities in the trophic response to injury is required. Fibrobast growth factor‐2 (FGF‐2) expression is developmentally regulated and has been implicated as a regulator of the wounding response of the adult rat central nervous system. We have investigated the expression of FGF‐2 and fibroblast growth factor receptor 1 (FGFR1) after penetrating lesions to the cerebral cortex of 5 days post partum (dpp) (nonscarring) and 16 dpp and adult (scarring) rats. In situ hybridization, immunohistochemistry and Western blotting showed robust and sustained increases in FGF‐2 and FGFR1 mRNA and protein in reactive astrocytes around the lesion in scarring rats, a response that was attenuated substantially in the nonscarring neonate. These results demonstrate that changes in astrocyte FGF‐2 and FGFR1 expression are coincident with the establishment of a mature pattern of glial scarring after injury in the maturing central nervous system, but it is premature to infer a causal relationship without further experiments.
Input‐dependent synaptic targeting of α2‐subunit‐containing GABAA receptors in synapses of hippocampal pyramidal cells of the rat
Authors: Nyíri Gábor , Freund Tamás F. , Somogyi Péter
Published: March 2003

Pyramidal cells, expressing at least 14 subunits of the heteropentameric GABAA receptor, receive GABAergic input on their soma and proximal dendrites from basket cells, activating GABAA receptors and containing either parvalbumin or cholecystokinin and vasoactive intestinal polypeptide. The properties of GABAA receptors are determined by the subunit composition, and synaptic receptor content governs the effect of the presynaptic neuron. Using a quantitative electron microscopic immunogold technique, we tested whether the synapses formed by the two types of basket cell show a difference in the subunit composition of GABAA receptors. Terminals of one of the basket cells were identified by antibodies to parvalbumin. Synapses made by parvalbumin‐negative terminals showed five times more immunoreactivity for the α2 subunit than synapses made by parvalbumin‐positive basket cells, whose synapses were frequently immunonegative. This difference is likely to be due to specific GABAA receptor α subunit composition, because neither synaptic size nor immunoreactivity for the β2/3 subunits, indicating total receptor content, was different in these two synapse populations. Synapses established by axo‐axonic cells on axon initial segments showed an intermediate number of immunoparticles for the α2 subunit compared to those made by basket cells but, due to their smaller size, the density of the α2 subunit immunoreactivity was higher in synapses on the axon. Because the two basket cell types innervate the same domain of the pyramidal cell, the results indicate that pyramidal cells have mechanisms to target GABAA receptors, under presynaptic influence, preferentially to distinct synapses. The two basket cell types act via partially distinct GABAA receptor populations.
GDNF reverses priming for dyskinesia in MPTP‐treated, l‐DOPA‐primed common marmosets
Authors: Iravani Mahmoud M. , Costa Sergio , Jackson Michael J. , Tel Banu C. , Cannizzaro Carla , Pearce Ronald K. B. , Jenner Peter
Published: March 2003

Parkinson's disease (PD) is associated with a progressive loss of dopamine neurons in the substantia nigra and degeneration of dopaminergic terminals in the striatum. Although l‐DOPA treatment provides the most effective symptomatic relief for PD it does not prevent the progression of the disease, and its long‐term use is associated with the onset of dyskinesia. In rodent and primate studies, glial cell line‐derived neurotrophic factor (GDNF) may prevent 6‐OHDA‐ or MPTP‐induced nigral degeneration and so may be beneficial in the treatment of PD. In this study, we investigate the effects of GDNF on the expression of dyskinesia in l‐DOPA‐primed MPTP‐treated common marmosets, exhibiting dyskinesia. GDNF or saline was administered by two intraventricular injections, 4 weeks apart, to MPTP‐treated, l‐DOPA‐treated common marmosets primed to exhibit dyskinesia. Prior to GDNF or saline administration, all animals displayed marked dyskinesia when treated with l‐DOPA. GDNF administration produced a significant improvement in motor disability and, following the second injection of GDNF, a significant improvement in the locomotor activity was observed. Following the administration of l‐DOPA there was a greater reversal of disability and a reduction in the intensity of l‐DOPA‐induced dyskinesia in GDNF‐treated animals compared to saline‐treated controls. However, there was no significant difference in l‐DOPA's ability to increase locomotor activity between GDNF‐treated and saline‐treated animals. GDNF treatment caused a significant increase in the number of tyrosine hydroxylase‐positive neurons in the substantia nigra, but no change in [3H]mazindol binding to dopamine terminals was found in the striatum of GDNF‐treated animals compared to saline‐treated controls. In GDNF‐treated animals a small but significant reduction in enkephalin mRNA was observed in the caudate nucleus but not in the putamen or the nucleus accumbens. Substance P mRNA expression was equally reduced in the caudate nucleus and the putamen of the GDNF‐treated animals but not in the nucleus accumbens. Intraventricular administration of GDNF improved MPTP‐induced disability and reversed dopamine cell loss in the substantia nigra. GDNF also diminished l‐DOPA‐induced dyskinesia, which may relate to its ability to partly restore nigral dopaminergic transmission or to modify the activity of striatal output pathways.
Cloning and functional expression of GABAB receptors from Drosophila
Authors: Mezler Mario , Müller Thomas , Raming Klaus
Published: March 2003

The neurotransmitter GABA (γ‐aminobutyric acid) functions as the major inhibitory neurotransmitter in the central nervous system of vertebrates and invertebrates. In vertebrates GABA signals both through ionotropic receptors (GABAA, GABAC), which induce fast synaptic inhibitory responses, and through metabotropic receptors (GABAB), which play a fundamental role in the reduction of presynaptic transmitter release and postsynaptic inhibitory potentials. Whilst GABAA and GABAC receptors have been cloned from vertebrates as well as invertebrates, GABAB receptors have only been identified in vertebrate species to date, although indirect evidence suggests their existence in arthropods, too. Here we report the cloning of three putative invertebrate GABAB receptor subtypes (D‐GABABR1, R2 and R3) isolated from Drosophila melanogaster. Whilst D‐GABABR1 and R2 show high sequence identity to mammalian GABABR1 and R2, respectively, the receptor D‐GABABR3 seems to be an insect‐specific subtype with no known mammalian counterpart so far. All three D‐GABABR subtypes are expressed in the embryonic central nervous system. In situ hybridization of Drosophila melanogaster embryos shows that two of the D‐GABABRs (D‐GABABR1 and R2) are expressed in similar regions, suggesting a coexpression of the two receptors, whilst the third D‐GABABR (D‐GABABR3) displays a unique expression pattern. In agreement with these results we have only been able to functionally characterize D‐GABABR1 and R2 when the two subtypes are coexpressed either in Xenopus laevis oocytes or mammalian cell lines, whilst D‐GABABR3 was inactive in any combination. The pharmacology of the coexpressed D‐GABABR1/2 receptor was different from the mammalian GABABRs: e.g. baclofen, an agonist of mammalian GABABRs, showed no effect.
Activation of murine microglial cell lines by lipopolysaccharide and interferon‐γ causes NO‐mediated decreases in mitochondrial and cellular function
Authors: Moss David W. , Bates Timothy E.
Published: March 2003

Activation of murine microglial and macrophage cell lines with lipopolysaccharide (LPS) and interferon‐γ (IFN‐γ) resulted in the induction of the inducible form of nitric oxide synthase (NOS) and the release of micromolar amounts of NO into the surrounding medium. The synthesis of NO was associated with increased cellular membrane damage as assessed by trypan blue dye exclusion and the leakage of lactate dehydrogenase into the cell culture medium. However, the synthesis and release of cytokines was largely unaffected. NO‐mediated cell damage was also accompanied by a marked decrease in the intracellular levels of reduced glutathione and ATP. In addition, significant inhibition of mitochondrial respiratory chain enzyme activities was seen following cellular activation. However, citrate synthase activity (a mitochondrial matrix enzyme) was not detectable in the extracellular supernatants, suggesting preservation of the integrity of the mitochondrial inner membrane following activation. These effects were largely prevented by the addition of the NOS inhibitor, N‐guanidino monomethyl l‐arginine during the activation period. Our observations demonstrate that induction of NOS activity in microglia results in damage to the plasma membrane leading to a loss of glutathione, complex‐specific inhibition of the mitochondrial electron transport chain and depletion of cellular ATP. Our data suggest that pharmacological modulation of NOS activity in activated microglia in vivo may prevent cellular damage to bystander cells such as neurons, astrocytes and oligodendrocytes, as well as to microglia themselves.
Functional projection distances of spinal interneurons mediating reciprocal inhibition during swimming in Xenopus tadpoles
Authors: Soffe S. R. , Zhao F.‐Y. , Roberts Alan
Published: March 2003

The basis for longitudinal coordination among spinal neurons during locomotion is still poorly understood. We have now examined the functional projection distances for the longitudinal axons of reciprocal inhibitory ‘commissural interneurons’ in the spinal cord of young Xenopus tadpoles. In quiescent animals, glycinergic inhibitory postsynaptic potentials (IPSPs) were evoked in ventral spinal neurons by stimulating small rostral and caudal groups of commissural interneuron somata at different distances on the opposite side of the hindbrain and spinal cord. Unitary IPSPs, produced by single synaptic contacts, could be distinguished from background noise. Local cord stimulation at different distances revealed maximum functional projection distances up to ∼ 0.5 mm for both descending and ascending axons, but with the probability of recording connections falling steeply over this distance. These maximum longitudinal projection distances are smaller than predicted by axonal anatomy (∼ 1.2 mm). We then measured functional projection distances during swimming by examining the synaptic output of a surgically isolated group of rostral commissural interneurons, mapping the occurrence of the mid‐cycle, reciprocal IPSPs they produced in more caudal neurons. IPSPs occurred with high probability up to 0.9 mm away, nearly twice the projection distance found in quiescent tadpoles. These results show that synaptic contacts from commissural interneurons could influence longitudinal coupling during swimming at distances of up to 0.9 mm (∼ 4–5 myotome segments or ∼ 25% of the spinal cord). They provide direct evidence for functional projection distances of a characterized class of interneurons belonging to a spinal locomotor pattern generator.
The selective cyclooxygenase‐2 inhibitor rofecoxib reduces kainate‐induced cell death in the rat hippocampus
Authors: Kunz Tina , Oliw Ernst H.
Published: March 2003

Treatment of male Sprague–Dawley rats with kainic acid (10 mg/kg, i.p.) triggered limbic seizures in 60% of the animals starting within 30 min and lasting for about 6 h. Cyclooxygenase‐2 (COX‐2) mRNA was strongly induced in the pyramidal cells of the hippocampus, in the amygdala and the piriform cortex after 8 h, as shown by in situ hybridization, and returned to control levels after 72 h. At this time marked cell loss occurred in the CA1–CA3 areas of the hippocampus. We hypothesize that rofecoxib, a selective COX‐2 inhibitor, might abbreviate the late neurotoxicity, possibly associated with COX‐2 induction. Animals which developed seizures were treated for 3 days with rofecoxib (10 mg/kg, i.p., n = 12) starting 6 or 8 h after kainic acid injection. Histological staining of viable cells confirmed that rofecoxib treatment selectively diminished cell loss in the hippocampus. The TdT‐mediated dUTP nick end labelling (TUNEL) technique was used to estimate delayed cell death. Abundant TUNEL‐positive cells were detected in seizure rats 72 h after kainic acid injection in pyramidal cells of the hippocampus (CA1–CA3), in cells of the thalamus, the amygdala and the piriform cortex. Treatment with rofecoxib selectively and significantly (P < 0.05) attenuated the number of TUNEL‐positive cells in the hippocampus, whereas the cells of the thalamus, amygdala and piriform cortex were not protected. Therefore we conclude that COX‐2 might contribute to cell death of pyramidal cells of the hippocampus as a consequence of limbic seizures.
Subunits of the nitric oxide receptor, soluble guanylyl cyclase, expressed in rat brain
Authors: Gibb Barry J. , Garthwaite John
Published: March 2003

Despite the widespread use of nitric oxide as a signalling molecule in the central nervous system, the molecular makeup of its receptor, soluble guanylyl cyclase (sGC), therein is poorly understood. Accordingly, RT‐PCR and in situ hybridization were used to identify sGC subunits expressed in rat brain. In addition to the expected mRNA for α1 and β1 subunits, message for the β2 subunit was detected in the cerebellum at all developmental stages investigated (1–150 days postnatum). The use of degenerate primers allowed the identification of mRNA coding for the rat α2 subunit, which was also expressed at every age studied. All but β2 were detected by in situ hybridization in the brains of both 8‐day‐old and adult rats. The distribution patterns indicated that in some areas, e.g. caudate–putamen and nucleus accumbens, sGC probably exists mainly as the α1β1 heterodimer. In others, e.g. hippocampus and olfactory bulb, α2β1 is likely to be dominant. In the cerebellum, α1 and β1 message was strong in the Purkinje cell layer but was not confined to Purkinje cells: smaller cells, presumed to be the Bergmann glia, were also labelled. In contrast, α2 mRNA was concentrated in cerebellar granule cells. Western blotting indicated an excess of α1 over β1 protein in the cerebellum, the reverse of what was found in the lung. It is concluded that, in molecular terms, sGC is likely to be more complex and exhibit more regional variation in the brain than previously thought. The functional consequences of this heterogeneity require investigation.
Olfactory learning is associated with increased spine density along apical dendrites of pyramidal neurons in the rat piriform cortex
Authors: Knafo Shira , Grossman Yoram , Barkai Edi , Benshalom Gadi
Published: March 2003

We studied the effect of olfactory learning on the dendritic spine density of pyramidal neurons in the rat piriform (olfactory) cortex. Rats were trained to distinguish between two pairs of odours in an olfactory discrimination task. Three days after training completion, rats were killed and layer II pyramidal neurons identified by Golgi impregnation were examined with a light microscope. Counts of visible spines were performed along the secondary and tertiary branches of both the apical dendrites and the basal dendrites, which are the sites of intracortical synaptic inputs. An estimate of the true spine density was obtained using Feldman and Peters' method (1979, The Journal of Comparative Neurology, 188, 527–542). The estimated true spine density along apical dendrites was higher in neurons from trained rats than those in pseudotrained and naive rats by 15%. As length of spiny dendrites did not change significantly after learning, the learning‐related increase in spine density in neurons from trained rats may indicate on an increased number of excitatory synapses interconnecting pyramidal neurons in the piriform cortex, following olfactory learning.
Peripheral auditory asymmetry in infantile autism
Authors: Khalfa S. , Bruneau N. , Rogé B. , Georgieff N. , Veuillet E. , Adrien J.‐L. , Barthélémy C. , Collet L.
Published: March 2003

Difficulty in filtering relevant auditory information in background noise is one of the features of autism. Auditory filtering processes can be investigated at the peripheral level as they are hypothesized to involve active cochlear mechanisms which are regulated by the efferent activity of the medial olivocochlear (MOC) system. The aim of the present work was therefore to assess these peripheral auditory processes in 22 children and adolescents with autism compared with age‐ and gender‐matched normal controls. Active cochlear mechanisms were evaluated with transiently evoked otoacoustic emissions (TEOAEs) and MOC system efficiency was assessed via TEOAEs which are decreased when stimulating the contralateral ear with noise. The MOC system evaluation was performed on 18 of the 22 children. In both studies, results were analysed according to age (from 4 to 10 years and from 11 to 20 years). The main result concerns the asymmetry of the efferent system which differs in individuals with autism. Several neural processes might be hypothesized as involved in the results obtained as the MOC system which originates in the brainstem received regulating controls from upper brain structures including auditory cortex. Lateralization abnormalities at the auditory periphery may reflect indirectly a problem at a higher level of auditory processing. A second important result shows a decrease in TEOAE amplitude with age, in patients, that may correspond to a decrease in hearing sensitivity.
Influence of injury and cytokines on synthesis of monocyte chemoattractant protein‐1 mRNA in peripheral nervous tissue
Authors: Subang M. C. , Richardson P. M.
Published: March 2003

The signals and the source of the signals for monocyte/macrophage entry into the injured peripheral nervous tissue are not yet defined. This study was undertaken to determine the distribution of the chemokine monocyte chemoattractant protein‐1 mRNA in injured rat and mouse nerves and to investigate the mechanisms that regulate its synthesis in rat Schwann cells. Results from RNase protection assays showed that, following sciatic nerve transection in rats, mRNA for monocyte chemoattractant protein‐1 was induced at the site of lesion within 3 h of surgery and in more distal segments from 24 h for at least 8 days. In cultured Schwann cells, tumour necrosis factor‐α but not interleukin‐1β, interleukin‐6, transforming growth factor‐β1, platelet‐derived growth factor‐BB or nerve growth factor induced monocyte chemoattractant protein‐1 mRNA in a time‐ and dose‐dependent fashion. The induction of monocyte chemoattractant protein‐1 mRNA in Schwann cells treated with tumour necrosis factor‐α was reduced by inhibitors of nuclear factor‐κB and the p38 mitogen‐activated protein kinase. In mice that lack the two receptors for tumour necrosis factor, the message for JE, a murine homologue of monocyte chemoattractant protein‐1, was still induced within 6 h of injury at the lesion site. However, in more distal segments 4 days after transection the concentration of JE mRNA was lower than that of control mice. Tumor necrosis factor‐α is the only cytokine that was shown to induce monocyte chemoattractant protein‐1 mRNA in cultured Schwann cells and is one of the factors that regulate the synthesis of monocyte chemoattractant protein‐1 in injured nerves.
Developmental and cellular expression pattern of epithelial sodium channel α, β and γ subunits in the inner ear of the rat
Authors: Gründer Stefan , Müller Andrea , Ruppersberg J. Peter
Published: March 2003

Endolymphatic ion composition in the adult inner ear is characterized by high K+ and low Na+ concentration. This unique ion composition is essential for proper functioning of sensory processing. Although a lot has been learned in recent years about molecules involved in K+ transport in inner ear, the molecules involved in Na+ transport are only beginning to emerge. The epithelial Na+ channel (ENaC) is a highly selective Na+ channel that is expressed in many Na+‐reabsorbing tissues. The aim of our study was to investigate whether ENaC is expressed in inner ear of rats and could account for Na+ reabsorption from endolymph. We detected mRNA for the three channel‐forming subunits (α, β and γENaC) in cochlea, vestibular system and endolymphatic sac. mRNA abundance increased during the first 12 days of life in cochlea and vestibular system, coinciding with decreasing Na+ concentration in endolymph. Expression was strongest in epithelial cells lining scala media, most notably Claudius' cells. As these cells are characterized by a very negative resting potential they would be ideally suited for reabsorption of Na+. mRNA abundance in endolymphatic sac decreased during the first 6 days of life, suggesting that ENaC might be implicated in reabsorption of endolymph in the endolymphatic sac of neonatal animals. Together, our results suggest that the epithelial Na+ channel is a good candidate for a molecule involved in Na+ homeostasis in inner ear.
The highly sialylated isoform of the neural cell adhesion molecule is required for estradiol‐induced morphological synaptic plasticity in the adult arcuate nucleus
Authors: Hoyk ZS. , Parducz A. , Theodosis D. T.
Published: March 2003

The large quantities of polysialic acid (PSA) characterizing highly sialylated isoform of the neural cell adhesion molecule (PSA‐NCAM), greatly reduce cell adhesion and render this particular cell surface adhesion molecule a likely candidate to intervene in dynamic neuronal phenomena, such as synaptic plasticity. The hypothalamic arcuate nucleus expresses high levels of PSA‐NCAM and maintains a high capacity for neuroplastic changes in the adult. Thus, in the arcuate nucleus of female rats, varying circulating levels of estrogen give rise to a reversible reduction in the number of axo‐somatic GABA synapses, together with a changing ensheathing of neuronal somata by astrocytes. To examine the role of PSA in such changes, we perturbed its expression, either by blockade with antibodies raised against this carbohydrate moiety (delivered intracerebroventricularly), or by its enzymatic cleavage after microinjection of endoneuraminidase N over the arcuate nucleus. Either procedure was performed in ovariectomized adult rats that received concurrent treatment with 17β‐estradiol. Morphological synaptic plasticity was analysed using the unbiased disector method to assess synaptic densities in ultrathin sections of the arcuate nucleus immunogold‐labelled for GABA. As expected, 17β‐estradiol induced a significant reduction in the number of GABAergic axo‐somatic synapses, a reduction which did not occur after infusion of anti‐PSA antibodies or in vivo enzymatic removal of PSA from NCAM. Taken together, our results provide strong evidence that the presence of large quantities of the PSA moiety on NCAM is a necessary prerequisite for estrogen‐induced phasic remodelling of synapses in the adult female arcuate nucleus.
Progression of spontaneous seizures after status epilepticus is associated with mossy fibre sprouting and extensive bilateral loss of hilar parvalbumin and somatostatin‐immunoreactive neurons
Authors: Gorter J. A. , Van Vliet E. A. , Aronica E. , Da Silva F. H. Lopes
Published: March 2003

The development of spontaneous limbic seizures was investigated in a rat model in which electrical tetanic stimulation of the angular bundle was applied for up to 90 min. This stimulation produced behavioural and electrographic seizures that led to a status epilepticus (SE) in most rats (71%). Long‐term EEG monitoring showed that the majority of the rats (67%) that underwent SE, displayed a progressive increase of seizure activity once the first seizure was recorded after a latent period of about 1 week. The other SE rats (33%) did not show this progression of seizure activity. We investigated whether these different patterns of evolution of spontaneous seizures could be related to differences in cellular or structural changes in the hippocampus. This was the case regarding the following changes. (i) Cell loss in the hilar region: in progressive SE rats this was extensive and bilateral whereas in nonprogressive SE rats it was mainly unilateral. (ii) Parvalbumin and somatostatin‐immunoreactive neurons: in the hilar region these were almost completely eliminated in progressive SE rats but were still largely present unilaterally in nonprogressive SE rats. (iii) Mossy fibre sprouting: in progressive SE rats, extensive mossy fibre sprouting was prominent in the inner molecular layer. In nonprogressive SE rats, mossy fibre sprouting was also present but less prominent than in progressive SE rats. Although mossy fibre sprouting has been proposed to be a prerequisite for chronic seizure activity in experimental temporal lobe epilepsy, the extent of hilar cell death also appears to be an important factor that differentiates between whether or not seizure progression will occur.
Real‐time, two‐dimensional visualization of ischaemia‐induced glutamate release from hippocampal slices
Authors: Uchino Shigeo , Nakamura Takeshi , Nakamura Kyoko , Nakajima‐Iijima Sadayo , Mishina Masayoshi , Kohsaka Shinichi , Kudo Yoshihisa
Published: March 2003

The involvement of excitatory amino acid (EAA) toxicity in ischaemia‐induced neuronal cell death has long been suggested. However, in the hippocampus, the brain site most vulnerable to ischaemia, the detailed spatial and temporal patterns of EAA release are not yet known. To address this issue, we have developed a novel strategy for the continuous, real‐time, two‐dimensional monitoring of EAA release from brain slices. As EAA detector, we used a cell line transformed with the N‐methyl‐d‐aspartate (NMDA) receptor, which is exclusively activated by EAAs, leading to an increase in the intracellular Ca2+ level. Combined with a calcium imaging technique, the use of this cell line allowed the temporal and regional analysis of EAA release from a brain slice placed directly on top of the clonal cells in a culture dish. Using this strategy, we demonstrated ischaemia‐induced EAA release in rat hippocampal slices. Increased EAA release was seen initially in the CA1 region, about 3 min after the beginning of ischaemia, then in the CA3 region and dentate gyrus, and, finally, throughout the hippocampal slice. Regional differences in extracellular EAA levels were also seen, with more EAA being released from the CA1 region than from the middle dentate gyrus. The present results are especially interesting as neurons in the CA1 region are more vulnerable to ischaemia than those in the CA3 region and dentate gyrus.
Differential role of the nitric oxide pathway on Δ9‐THC‐induced central nervous system effects in the mouse
Authors: Azad Shahnaz Christina , Marsicano Giovanni , Eberlein Irina , Putzke Jörg , Zieglgänsberger Walter , Spanagel Rainer , Lutz Beat
Published: March 2003

This study investigated whether the nitric oxide pathway was involved in the central effects of Δ9‐tetrahydrocannabinol (Δ9‐THC), the major psychoactive constituent of cannabis sativa. Body temperature, nociception and locomotion were measured in neuronal nitric oxide synthase (nNOS) knock‐out (KO) mice and wild‐type (WT) controls after intraperitoneal application of Δ9‐THC. These Δ9‐THC‐induced effects are known to be mediated through the brain‐type cannabinoid receptor 1 (CB1). Therefore, in situ hybridization (ISH) experiments were performed in the adult murine brain to determine possible changes in CB1 mRNA levels in nNOS‐KO, compared with WT mice, and to reveal brain areas where CB1 and nNOS were coexpressed in the same neurons. We found that an intraperitoneal injection of 10 mg/kg Δ9‐THC led to the same increase in the hot plate latencies in both genotypes, suggesting that Δ9‐THC‐mediated antinociception does not involve nNOS. In contrast, a significant Δ9‐THC‐induced decrease of body temperature and locomotor activity was only observed in WT, but not in nNOS‐KO mice. ISH revealed significantly lower levels of CB1 mRNA in the ventromedial hypothalamus (VMH) and the caudate putamen (Cpu) of the nNOS‐KO animals, compared with WT mice. Both areas are known to be among the regions involved in cannabinoid‐induced thermoregulation and decrease of locomotion. A numerical evaluation of nNOS/CB1 coexpression showed that approximately half of the nNOS‐positive cells in the dorsolateral Cpu also express low levels of CB1. ISH of adjacent serial sections with CB1 and nNOS, revealed expression of both transcripts in VMH, suggesting that numerous nNOS‐positive cells of VMH coexpress CB1. Our findings indicate that the nitric oxide pathway is involved in some, but not all of the central effects of Δ9‐THC.
Enhanced CREB phosphorylation in immature dentate gyrus granule cells precedes neurotrophin expression and indicates a specific role of CREB in granule cell differentiation
Authors: Bender R. A. , Lauterborn J. C. , Gall C. M. , Cariaga W. , Baram T. Z.
Published: March 2003

Differentiation and maturation of dentate gyrus granule cells requires coordinated interactions of numerous processes. These must be regulated by protein factors capable of integrating signals mediated through diverse signalling pathways. Such integrators of inter and intracellular physiological stimuli include the cAMP‐response element binding protein (CREB), a leucine‐zipper class transcription factor that is activated through phosphorylation. Neuronal activity and neurotrophic factors, known to be involved in granule cell differentiation, are major physiologic regulators of CREB function. To examine whether CREB may play a role in governing coordinated gene transcription during granule cell differentiation, we determined the spatial and temporal profiles of phosphorylated (activated) CREB throughout postnatal development in immature rat hippocampus. We demonstrate that CREB activation is confined to discrete, early stages of granule cell differentiation. In addition, CREB phosphorylation occurs prior to expression of the neurotrophins BDNF and NT‐3. These data indicate that in a signal transduction cascade connecting CREB and neurotrophins in the process of granule cell maturation, CREB is located upstream of neurotrophins. Importantly, CREB may be a critical component of the machinery regulating the coordinated transcription of genes contributing to the differentiation of granule cells and their integration into the dentate gyrus network.
Influence of glucocorticoids on dopaminergic transmission in the rat dorsolateral striatum
Authors: Barrot Michel , Abrous Djoher Nora , Marinelli Michela , Rougé‐Pont Françoise , Le Moal Michel , Piazza Pier Vincenzo
Published: March 2003

Glucocorticoid hormones exert strong influences on central neurotransmitter systems. In the present work, we examined the functional consequences of corticosterone suppression on the dopaminergic transmission in the dorsolateral striatum by studying the expression of Fos‐like proteins and extracellular dopamine levels. Glucocorticoid hormones were suppressed by adrenalectomy, and the specificity of the effects assessed by restoring physiological plasmatic corticosterone concentrations. We show that, in the dorsolateral striatum, glucocorticoids modify postsynaptic dopaminergic transmission. Suppression of glucocorticoids decreased the induction of Fos proteins in response to a direct agonist of dopamine D1 receptors (SKF 82958, 1.5 mg/kg, i.p.), but not the release of dopamine induced by morphine (2 mg/kg, s.c.) or the density of the limiting enzyme of dopamine synthesis, tyrosine hydroxylase. In contrast to the dopaminergic response to morphine, the response to cocaine (15 mg/kg, i.p.) was modified by the suppression of corticosterone. In this case, adrenalectomy increased cocaine‐induced changes in extracellular dopamine but did not modify the expression of Fos‐like proteins. This absence of changes in cocaine‐induced Fos‐like proteins might result from a compensatory mechanism between the increase in the dopaminergic response and the decrease in the functional activity of dopamine D1 receptors. The increased dopaminergic response to cocaine also contrasts with the decreased response previously observed in the shell of the nucleus accumbens [Barrot et al. (2000) Eur. J. Neurosci., 12, 973–979]. The present data highlight the profound heterogeneous influence of glucocorticoids within dopaminergic projections.
Immunohistochemical localization of voltage‐gated calcium channels in substantia nigra dopamine neurons
Authors: Takada M. , Kang Y. , Imanishi M.
Published: March 2003

The rhythmic firing of dopaminergic (DA) neurons in the substantia nigra pars compacta (SNc) is thought to be mediated by nifedipine‐sensitive Ca2+channels, although an involvement of ω‐conotoxin‐sensitive Ca2+channels is also suggested. In an attempt to localize such Ca2+channels at both the regional and cellular levels, their expression and distribution patterns were immunohistochemically investigated in the rat SNc. The three distinct subtypes of voltage‐gated Ca2+channels were tested: the class B N‐type α1 subunit (CNB1), the class C L‐type α1 subunit (CNC1) and the class D L‐type α1 subunit (CND1). A large number of SNc neurons showed intense immunoreactivity against CND1 and they were distributed throughout the entire extent. By contrast, many fewer neurons displayed less intense CNC1 immunoreactivity and many of them were located in the lateral aspect of the SNc. No immunoreactivity against CNB1 was detected in the SNc. Moreover, double immunofluorescence analysis in combination with tyrosine hydroxylase staining revealed that virtually all DA neurons were CND1‐immunoreactive whereas many DA neurons especially in the medial SNc exhibited only faint or no immunoreactivity against CNC1. Both CNC1 and CND1 were expressed in cell bodies and proximal dendrites of SNc DA neurons, whilst their distal dendrites that penetrated into the substantia nigra pars reticulata expressed CND1 alone. Thus, the ubiquitously and intensely expressed class D α1 subunit of L‐type Ca2+channels that is sensitive to both nifedipine and ω‐conotoxin may be responsible for the pacemaker activity of SNc DA neurons.
Bcl‐2 overexpression per se does not promote regeneration of neonatal crushed optic fibers
Authors: Lodovichi Claudia , Di Cristo Graziella , Cenni Maria Cristina , Maffei Lamberto
Published: March 2003

We have explored whether overexpression of the bcl‐2gene ‘per se’ can promote regeneration of retinal ganglion cells (RGCs) after optic nerve axotomy in developing transgenic mice. We have used newborn mice (postnatal day 5) because at this age the central nervous system environment is more permissive for regeneration than in adults, thus, maximizing the probability to detect a regeneration‐promoting role of bcl‐2. Thirty days postsurgery we found that in mice overexpressing bcl‐2, a high proportion of retinal ganglion cells survived and also that some fibers in the proximal stump of the optic nerve were preserved. However, the optic nerve of transgenic mice does not show signs of regeneration. On the contrary, in the presence of Schwann cell transplants, there are signs of fiber regrowth. Indeed, many axonal terminals cross the crush site and reach the chiasm in both wild type and transgenic mice nerves. These results suggest that bcl‐2 overexpression is not sufficient ‘per se’ to increase the regenerative potentiality of axotomized RGCs.