bluuebunny/crossref_metadata_2025 huggingface dataset:
Embryonic Optic Nerve Tissue Fails to Support Neurite Outgrowth by Central and Peripheral Neurons In Vitro
Authors: Shewan Derryck , Berry Martin , Bedi Kuldip , Cohen James
Published: April 2006

The failure of axon regeneration in the injured mammalian central nervous system has been ascribed, in part, to the inhibitory effects of myelin proteins. To investigate the influence of myelination on neurite growth and regeneration by both central nervous system and peripheral nervous system neurons, isolated rat neonatal retinal ganglion cells and adult and neonatal dorsal root ganglion neurons were cultured on cryostat sections of both immature unmyelinated and mature fully myelinated adult rat optic nerve. In agreement with earlier studies using neonatal peripheral neurons, the adult optic nerve failed to support neurite outgrowth from any of the neurons tested. A new finding was that tissue sections from unmyelinated optic nerve (aged embryonic days 18 and 20, and postnatal days 1–3), also failed to support the growth of neurites from neonatal retinal ganglion cells and both neonatal and adult dorsal root ganglion neurons. Neonatal retinal ganglion cells also failed to extend neurites on sections of pre‐degenerated sciatic nerve, a tissue shown in our previous work to be a good substratum for supporting neurite growth for both neonatal and adult DRG neurons. These results suggest that cells in the immature optic nerve either express widely acting axon growth inhibitory molecules unrelated to previously described myelin proteins, or do not synthesize appropriate axon growth promoting molecules. They also reveal that, for axon regeneration, central nervous system and peripheral sensory neurons require distinct substratum interactions.
Monocularly Induced 2‐Deoxyglucose Patterns in the Visual Cortex and Lateral Geniculate Nucleus of the Cat: II. Awake Animals and Strabismic Animals
Authors: Löwel Siegrid , Singer Wolf
Published: April 2006

In the course of experiments studying the organization of ocular dominance columns in the visual cortex of cats, we noticed that—contrary to common belief—labelling with 2‐deoxyglucose after monocular stimulation failed to induce a pattern of ocular dominance columns but resulted in a rather homogeneous 2‐deoxyglucose uptake throughout area 17 in anaesthetized and paralysed animals. We wondered whether 2‐deoxyglucose columns could be obtained in awake animals and/or in strabismic animals, which have a more pronounced segregation of ocular dominance columns. To this end, we investigated 2‐deoxyglucose patterns after monocular stimulation in three groups of animals: (i) in awake normally reared cats, (ii) in awake strabismic cats and (iii) in anaesthetized and paralysed strabismic cats. Additionally, we labelled ocular dominance columns with intraocular [3H]proline injections. In all cats, monocular stimulation induced 2‐deoxyglucose patterns that were in precise register with the proline‐labelled ocular dominance columns in layer IV. Regions of increased 2‐deoxyglucose uptake extended in a columnar fashion through all cortical layers. In contrast to normally reared animals, in strabismic cats, the expression of 2‐deoxyglucose labelled ocular dominance columns was not abolished by anaesthesia or paralysis. However, there was a difference between the 2‐deoxyglucose patterns in the awake normally reared cats and the strabismic animals. In the former, the patches of 2‐deoxyglucose labelling were smaller and occupied less territory than the afferents of the stimulated eye in layer IV. Together with the results of the previous study, these data indicate that in awake normally reared and in awake and anaesthetized strabismic cats, but not in anaesthetized and paralysed normally reared animals, monocular stimulation activates selectively neurons in columns that are in register with the termination sites of afferents from the stimulated eye. This suggests the existence of a mechanism in normally reared animals which restricts cortical activation after monocular stimulation to territories that are in register with the afferents from the stimulated eye. This mechanism appears to be effective only when the animals are awake and actively exploring their environment. This and the fact that the active columns were narrower than the terminal fields of the stimulated eye suggest an active inhibitory process, perhaps related to mechanisms of selective attention. The observation that ocular dominance columns persist in strabismic cats even under anaesthesia can be accounted for by the lack of binocular convergence in these animals.
Visually Triggered Neuronal Oscillations in the Pigeon: An Autocorrelation Study of Tectal Activity
Authors: Neuenschwander Sergio , Varela Francisco J.
Published: April 2006

In this study we describe visually triggered gamma oscillations in the optic tectum of awake pigeons. This study was motivated by the potential relevance of synchronous oscillatory responses in perceptual binding in a laminated structure other than the mammalian neocortex. Tectal responses were recorded as local field potential and multiunit activity by differential filtering. The local field potential was analysed by computing its autocorrelation function and spectral power with a moving window applied to single response sweeps. The temporal structure of the spike trains was evaluated by computing averaged autocorrelograms. A damped sine wave function was fitted to the autocorrelograms in order to quantify the degree of oscillation of both signals. Epochs of significant oscillatory activity were observed in the local field potential in 60% of the trials (n= 39). In all trials, significant oscillations occupied ∼ 10% of the time the stimulus was present. The oscillatory events in both the local field potential and the multiunit activity had frequencies in the range of 20 – 50 Hz. It is important to emphasize the great variability in the frequency and in the probability of occurrence of the oscillatory responses from trial to trial, which makes the oscillatory behaviour of the tectal activity highly non‐stationary. The oscillatory activity we describe in the avian tectum has characteristics similar to those reported in the mammalian neocortex. These findings from a fully awake animal strengthen the universality of oscillations as a possible carrier for synchronization of activity in the constitution of neuronal assemblies.
The Isolated and Perfused Brain of the Guinea‐pig In Vitro
Authors: Mühlethaler M. , de Curtis M. , Walton K. , Llinás R.
Published: April 2006

We describe here an isolated and perfused in vitro adult guinea‐pig whole brain preparation which is an extension of the previously described in vitro brainstem–cerebellum preparation, Viability was tested by the analysis of trans‐synaptic responses along the visual pathways following the electrical stimulation of the optic nerve or the optic radiations. The evoked field potentials were recorded in the dorsal lateral geniculate, the superior colliculus and the visual cortex. The distribution of extracellular currents was studied using current source density analysis, in order to determine the amplitude, time course and spatial organization of the synaptic activity at these sites. The study indicates that field potentials were very similar to those described in vivo. These data demonstrate the survival of a complex adult sensory system in vitro and suggest that this preparation can be used for the analysis of multisynaptic circuits in the mammalian brain.
FGF‐2 Modulates Dopamine and Dopamine‐related Striatal Transmitter Systems in the Intact and MPTP‐lesioned Mouse
Authors: Otto Dörte , Unsicker Klaus
Published: April 2006

Following a previous study in which we showed ameliorative effects of basic fibroblast growth factor (FGF‐2) locally applied to the nigrostriatal system in 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP)‐lesioned mice, we investigated FGF‐2 actions at different time intervals after the lesion and effects on non‐dopaminergic striatal transmitter systems. A triple intraperitoneal injection of 30 mg/kg MPTP at 24 h intervals caused a reduction of striatal dopamine to 23% of control levels that lasted for at least 4 weeks. Four μg FGF‐2 soaked into gel foam and placed onto the right striatum partially and bilaterally restored dopamine levels and tyrosine hydroxylase activity after 2 weeks, when the treatment started simultaneously or 1 day after the toxin lesion. FGF‐2 was ineffective, if administration commenced with a delay of 7 days. Striatal neurotransmitters that are known to be linked to the dopaminergic system were also altered by the MPTP treatment. GABA was significantly increased, while somatostatin levels were reduced. Upon FGF‐2 administration both GABA and somatostatin levels were partially normalized. Our data are consistent with the notion that FGF‐2 protects and rescues acutely and subacutely MPTP‐lesioned nigrostriatal neurons and that its effects must be mainly indirect. Likewise, positive effects of FGF‐2 on non‐dopaminergic neurons may be due to the partial restoration of striatal dopamine.
Neuroprotective Effect of the χ‐Agonist Enadoline (CI‐977) in Rat Models of Focal Cerebral Ischaemia
Authors: Hayward Neil James , McKnight Alexander Thorburn , Woodruff Geoffrey Neil
Published: April 2006

The neuroprotective efficacy of the χ‐opioid agonist enadoline (CI‐977) was examined in two acute rat models of focal cerebral ischaemia [non‐recovery (4 h) and recovery (24 h)]. In the non‐recovery model, Sprague ‐ Dawley rats were anaesthetized throughout the study period. Focal ischaemia was produced by the permanent occlusion of the left middle cerebral artery (MCA). The amount of early ischaemic damage was assessed in coronal sections at nine pre‐determined stereotaxic planes. Enadoline at doses of 0.1, 0.3 and 1.0 mg/kg (n= 8), administered s.c. 30 min prior to ischaemia, produced dose‐dependent amelioration of cortical damage. Importantly, enadoline had no significant effect on any of the physiological parameters monitored (blood pressure, blood gases, glucose, pH). In the recovery model the left MCA was permanently occluded under isoflurane anaesthesia. Animals were allowed to recover and were killed 24 h later. The amount of ischaemic brain damage and swelling was assessed histologically. In this model pretreatment with enadoline at either 0.1, 0.3, or 1.0 mg/kg s.c. was followed by continuous s.c. infusion at 0.017, 0.05 or 0.17 mg/kg/h respectively (n= 8–17). Enadoline produced dose‐dependent reductions in the volumes of infarction and brain swelling; the greatest reductions were seen at 1.0 mg/kg plus 0.17 mg/kg/h in both infarction (reduced by 37.4% from controls) and swelling (reduced by 47.8%). Therefore the χ‐opioid agonist enadoline affords dose‐dependent neuroprotection in both the non‐recovery and recovery models of focal cerebral ischaemia in the rat. In the recovery model enadoline was effective in reducing infarction and oedema; the reduction in brain swelling occurs in parallel with the reduction in ischaemic damage.
Trophic Actions of Ciliary Neurotrophic Factor on Murine Embryonic Carcinoma Cells
Authors: Gupta S. K. , Haggarty A. J. , Carbonetto S. , Riopelle R. J. , Richardson P. M. , Dunn R. J.
Published: April 2006

Recombinant rat CNTF (ciliary neurotrophic factor) at picomolar concentration prevents the death of P19 murine embryonic carcinoma cells that usually follows upon withdrawal of serum from the culture medium. For prolonged survival of P19 cells in serum‐free medium, insulin must also be present. In the presence or absence of serum, CNTF stimulates the differentiation of P19 cells, inducing the formation of neurites and synthesis of neurofilament. The results of radioautographic studies with radioiodinated CNTF indicate the presence of high‐affinity binding sites on P19 cells. Equilibration of P19 cells with [125I]CNTF followed by incubation with cross‐linking reagents reveals evidence for at least two putative receptors of ∼78 and ∼167 kDa.
Voltage‐dependent Currents in Microvillar Receptor Cells of the Frog Vomeronasal Organ
Authors: Trotier Didier , Døving Kjell B. , Rosin Jean‐François
Published: April 2006

Vomeronasal receptor cells are differentiated bipolar neurons with a long dendrite bearing numerous microvilli. Isolated cells (with a mean dendritic length of 65 μm) and cells in mucosal slices were studied using whole‐cell and Nystatin‐perforated patch‐clamp recordings. At rest, the membrane potential was −61 ± 13 mV (mean ± SD; n= 61). Sixty‐four per cent of the cells had a resting potential in the range of –60 to –86 mV, with almost no spontaneous action potential. The input resistance was in the GΩ range and overshooting repetitive action potentials were elicited by injecting depolarizing current pulses in the range of 2 – 10 pA. Voltage‐dependent currents were characterized under voltage‐clamp conditions. A transient fast inward current activating near –45 mV was blocked by tetrodotoxin. In isolated cells, it was half‐deactivated at a membrane potential near –75 mV. An outward K+ current was blocked by internal Cs+ ions or by external tetraethylammonium or Ba2+ ions. A calcium‐activated voltage‐dependent potassium current was blocked by external Cd2+ ions. A voltage‐dependent Ca2+ current was observed in an iso‐osmotic BaCl2 solution. Finally, a hyperpolarization‐activated inward current was recorded. Voltage‐dependent currents in these microvillar olfactory receptor neurons appear qualitatively similar to those already described in ciliated olfactory receptor cells located in the principal olfactory epithelium.
Processing of the FMRFamide Precursor Protein in the Snail Lymnaea stagnalis: Characterization and Neuronal Localization of a Novel Peptide, ‘SEEPLY’
Authors: Santama Niovi , Wan Li Ka , Bright Kerris E. , Yeoman Mark , Geraerts Wijnand P. M. , Benjamin Paul R. , Burke Julian F.
Published: April 2006

In the pulmonate snail Lymnaea stagnalis, FMRFamide‐like neuropeptides are encoded by a multi‐exon genomic locus which is subject to regulation at the level of mRNA splicing. We aim to understand the post‐translational processing of one resulting protein precursor encoding the tetrapeptide FMRFamide and a number of other putative peptides, and determine the distribution of the final peptide products in the central nervous system (CNS) and periphery of Lymnaea. We focused on two previously unknown peptide sequences predicted by molecular cloning to be encoded in the tetrapeptide protein precursor consecutively, separated by the tetrabasic cleavage site RKRR. Here we report the isolation and structural characterization of a novel non‐FMRFamide‐like peptide, the 22 amino acid peptide SEQPDVDDYLRDWLQSEEPLY. The novel peptide is colocalized with FMRFamide in the CNS in a number of identified neuronal systems and their peripheral motor targets, as determined by in situ hybridization and immunocytochemistry. Its detection in heart excitatory motoneurons and in nerve fibres of the heart indicated that the novel peptide may play a role, together with FMRFamide, in heart regulation in the snail. The second predicted peptide, STEAGGQSEEMTHRTA (16 amino acids), was at very low abundance in the CNS and was only occasionally detected. Our current findings, suggestive of a distinct pattern of post‐translational processing, allowed the reassessment of a previously proposed hypothesis that the two equivalent sequences in the Aplysia FMRFamide gene constitute a molluscan homologue of vertebrate corticotrophin releasing factor‐like peptides.
Detection of a Purine Analogue‐sensitive Kinase in Frog Sciatic Nerves—Possible Involvement in Nerve Regeneration
Authors: Svensson Bodil , Biessels Geert‐Jan , Per Ekström
Published: April 2006

This study reports the existence of a purine analogue‐sensitive protein kinase in adult frog sciatic nerves. Cell‐free supernatants of homogenized regenerating sclatic nerves were found to contain a phosphoprotein (MW 90 kDa, referred to as PP90), that was phosphorylated to a much higher degree than in normal, uninjured nerves. The spatial and temporal characteristics of PP90 phosphorylation suggested a relationship with the injury‐induced proliferation of support cells of the regenerating nerve, i.e. its appearance and increment over time correlated with that of [3H]thymidine incorporation in the nerve. PP90 was phosphorylated under conditions that excluded enzyme activities due to Ca2+/calmodulin kinases, cyclic nucleotide‐dependent kinases or protein kinase C. On the other hand, the phosphorylation could be selectively inhibited by the purine analogues adenosine, 2‐aminopurine and 6‐thioguanine (6‐TG). The latter was the most potent and gave complete inhibition at 50 μM. Addition of histone H1 to the cell‐free assay stimulated the phosphorylation of several proteins in both normal and regenerating nerves. The stimulation could be blocked by 6‐TG, indicating the presence of a purine‐sensitive kinase also in uninjured nerves. Separate experiments showed that in vitro regeneration of the frog sciatic sensory axons, as well as the proliferation of the support cells, was inhibited by 100 μM 6‐TG.
Functional Heterogeneity of Hippocampal GABAA Receptors
Authors: Schönrock Beate , Bormann Joachim
Published: April 2006

γ‐Aminobutyric acid type A (GABAA) receptors were studied in cultured neurons taken from rat hippocampus at early postnatal stages. GABA‐induced whole‐cell currents showed a broad range of peak amplitudes and time‐courses of desensitization. Dose – response curves of rapidly and slowly desensitizing cells revealed EC50 values of 8.5 and 37.3 μM GABA, respectively, with the Hill coefficient being greater than unity. The main‐state conductance of GABAA receptor channels was 28 – 31 pS in all cells. GABA responses of low‐affinity cells were more strongly affected by benzodiazepine receptor agonists (e.g. flunitrazepam, clonazepam) and inverse agonists (e.g. methyl‐6,7‐dimethoxy‐4‐ethyl‐β‐carboline‐3‐carboxylate), as compared to cells exhibiting high‐affinity GABA responses. Currents were also potentiated by zolpidem, but were little affected by Ro 15‐4513 and Zn2+. These data suggest the presence of physiologically and pharmacologically distinct GABAA receptor isoforms in neurons of the early postnatal hippocampus, which may subserve different inhibitory control mechanisms in this brain region.
Enrichment of Glutamate‐like Immunoreactivity in Primary Afferent Terminals Throughout the Spinal Cord Dorsal Horn
Authors: Broman Jonas , Anderson Sonya , Ottersen Ole P.
Published: April 2006

Although several lines of evidence indicate that glutamate is a neurotransmitter in primary afferent terminals, controversies exist on the proportion and types of such terminals that release glutamate. In the present study quantitative analysis of immunogold labelling was used to assess the presence of glutamate‐like immunoreactivity in primary afferent terminals in laminae I – V of the rat spinal cord dorsal horn. Anterograde transport of choleragenoid – horseradish peroxidase from a spinal ganglion and tetramethyl benzidine histochemistry were used to identify primary afferent terminals in laminae I and III – V. Presumed C‐fibre terminals in lamina II were identified on morphological criteria (dense sinusoid axon terminals). Primary afferent terminals in all dorsal horn laminae displayed significantly higher levels of glutamate‐like immunoreactivity than pleomorphic vesicle‐containing profiles in laminae III – IV and large neuronal cell bodies in laminae III – V. The density of gold particles over primary afferent terminals also significantly exceeded the average density of gold particles over laminae II and III – IV. The highest densities of gold particles were present over dense sinusoid axon terminals in lamina II. These findings suggest that glutamate, alone or in combination with other neuroactive compounds, is involved in the transfer of all sensory modalities from primary afferent fibres to dorsal horn neurons.
Muscle Afferents Innervating Skin Form Somatotopically Appropriate Connections in the Adult Rat Dorsal Horn
Authors: Lewin Gary R. , McMahon Stephen B.
Published: April 2006

We have studied the somatotopic reorganization in dorsal horn neurons after a disruption in the normal spatial arrangement of primary sensory axons in adult rats. Muscle afferents were redirected to skin by cutting and cross‐anastomosing the hindlimb gastrocnemius nerve (GN) and sural nerve (SN). It has previously been shown that after 10 – 12 weeks GN afferents innervate the hairy skin of the lateral ankle and calf (previously innervated by SN afferents) and become potentially capable of relaying information on the location and intensity of stimuli applied to the skin. We determined the receptive field and response properties of dorsal horn neurons in the lumbar spinal cord, in regions where the lower hindlimb is normally represented. In control animals (with intact or self‐anastomosed sural nerves) very few neurons (<8%) received any synaptic input from the GN as assessed by electrical stimulation of the nerve. In contrast, when this nerve innervated skin, many cells responded to GN stimulation, and these nearly all had receptive field components in the former SN territory. Moreover, in animals with cross‐anastomosed nerves, cells without GN inputs all had receptive fields outside the former SN skin territory. We have shown that in all likelihood GN afferents substituted for SN afferents in subserving the low and high threshold receptive fields of dorsal horn neurons. Furthermore, for many neurons, receptive fields were formed from inappropriately regrown GN afferents and adjacent intact cutaneous afferents (in the tibial or common peroneal nerves). Therefore, when GN afferents innervate skin in adult animals, they alter their central connectivity in an appropriate manner for their new peripheral terminations, so that an orderly somatotopic representation of the hind limb skin is maintained. We suggest that this plasticity of dorsal horn somatotopy is driven in part by activity‐dependent mechanisms.
Functional Topography of the Myelin‐associated Glycoprotein. I. Mapping of Domains by Electron Microscopy
Authors: Fahrig Thomas , Probstmeier Rainer , Spiess Eberhard , Meyer‐Franke Anke , Kirchhoff Frank , Drescher Bernd , Schachner Melitta
Published: April 2006

The functional topography of the myelin‐associated glycoprotein (MAG) was investigated by electron microscopic analysis of rotary‐shadowed molecules of a MAG fragment (MAG 90) comprising the five immunoglobulin‐like domains of the extracellular part of the molecule. MAG 90 molecules appeared as rod‐like structures (18.5±1.2 nm long and 4.0±0.8 nm wide) with a globular domain at one end. Antibodies directed against the amino‐ and carboxy‐terminus of MAG 90 interacted with the non‐globular terminal region, indicating that the molecule is bent in the globular region with the amino‐ and carboxy‐terminal arms in close apposition to each other. An antibody which interferes with the binding of MAG to neurons interacted predominantly with the globular domain of MAG 90. The fibril‐forming collagen types I, III and V bound mainly to the non‐globular terminal region of MAG 90, whereas the majority of heparin molecules interacted with the globular region of the molecule. The L2/HNK‐1 carbohydrate structure was localized at the non‐globular region in the protein fragment comprising the fourth and fifth immunoglobulin‐like domains.
Persistence of Axonal Transport in Isolated Axons of the Mouse
Authors: Smith R. S. , Bisby M. A.
Published: April 2006

We have examined the hypothesis, for the case of mouse axons, that isolating an axon from its cell body will lead to a rapid failure of fast axonal transport as anterogradely moving organelles vacate the axon in a proximo‐distal direction, and retrogradely moving organelles vacate it in the opposite direction. We used CD1 and BALB/c mice and the Wallerian degeneration‐resistant mutant C57BL/Ola. Sciatic nerves were cut high in the thigh; at various times up to 8 days later nerves were removed from the animal and individual myelinated axons from the segment distal to the cut were examined by video light microscopy to detect rapid organelle transport. Bidirectional fast organelle transport did decrease in amount with time but not nearly as rapidly as predicted, and anterograde and retrograde organelle velocities remained normal through time. In the C57BL/Ola mouse some structurally preserved axons contained organelles that transported at normal velocities in the anterograde and retrograde directions for as long as 8 days after axotomy. To test one of the possible origins of transported organelles in long‐surviving axons we examined organelle transport very close to narrow lesions in axons bathed in a medium compatible with intracellular function. No organelles crossed the lesion but bidirectional organelle transport took place proximal and distal to the lesion; the amounts were compatible with the interpretation that ∼30% of organelles reversed transport direction on either side of the lesion. We propose that at least some of the organelles that undergo persistent transport in axons isolated from their cell bodies shuttle back and forth between the ends of the isolated segment.
Tracing Neuroepithelial Cells of the Mesencephalic and Metencephalic Alar Plates During Cerebellar Ontogeny in Quail – chick Chimaeras
Authors: Hallonet Marc E. R. , Le Douarin Nicole M.
Published: April 2006

The quail – chick chimaera system was used to investigate the origin of the various neuronal cell types of the cerebellum from the mesencephalic and metencephalic brain vesicles at the 11‐ to 14‐somite stage in the avian embryo. We have already demonstrated that the cerebellum is derived from both the mesencephalic and metencephalic brain vesicles. The mesencephalic contribution to the cerebellum is restricted to a mediodorsal territory inserted as a V‐shaped area into the primitive metencephalic vesicle through complex morphogenetic movements taking place from day 2 to day 4 of embryonic development. Here we report that the cerebellar presumptive territory extends along the anteroposterior axis, over the caudal half of the mesencephalon and the rostral half of the metencephalon. Along the dorsoventral axis, the cerebellar anlage is located in the alar plates at the exclusion of the roof and basal plates, i.e. lies in the lateral walls of the neuroepithelium in the area included between ∼25 and 120° with respect to the sagittal plane. We also report that the neuroepithelium corresponding to the cerebellar presumptive territory also yields other brain structures (e.g. part of the optic tectum in the mesencephalon). The external granular layer (EGL) arises only from the rostral metencephalon, undergoing extensive tangential movements which we have analysed in detail: the more ventral the position of cells in the metencephalic alar plates the more rostral and lateral is their position in the EGL. Finally, we discuss the fact that the cerebellar cortex, an integrative structure of the brain, arises from the alar plates of the neural tube. This is consistent with the general spatial organization of the neural anlage of the vertebrate embryo, in which this part of the neuroepithelium is devoted to the production of interneurons, whereas the basal plate and the neural folds yield motor structures and primary sensory neurons respectively.
In Vitro Enfolding of Olfactory Neurites by p75 NGF Receptor Positive Ensheathing Cells from Adult Rat Olfactory Bulb
Authors: Ramón‐Cueto Almudena , Pérez Julio , Nieto‐Sampedro Manuel
Published: April 2006

Secondary cultures of adult rat olfactory bulb (OB) contained three different types of cell: (i) process‐bearing cells; (ii) macrophage‐like cells and (iii) fusiform cells. The immunohistochemical properties of process‐bearing cells closely corresponded to those described for ensheathing glia in vivo. The most distinctive feature of these cells was their immunoreactivity for low affinity nerve growth factor receptor (NGFR). Process‐bearing cells also shared the ultrastructural properties of ensheathing glia in vivo, as well as the ability to ensheath olfactory axons. In contrast, macrophage‐like cells had the immunostaining properties of microglia, and fusiform cells were likely capillary endothelial cells.

Neurites outgrowing from olfactory epithelium explants, when co‐cultured with adult OB cells, grew preferentially over NGFR positive cells. Olfactory neurites exhibited NGFR immunoreactivity and were enfolded by NGFR positive cells. After ensheathment, this immunoreactivity decreased from the neurite and disappeared from the glial membrane in contact with the neurite. However, NGFR immunoreactivity was maintained in the portion of the glial membrane not involved in ensheathing. In summary, ensheathing cells in vitro retained both the ultrastructure shown in vivo and the ability to ensheath olfactory neurites. The Schwann cell‐like properties of ensheathing glia, could partially explain the permissibility of adult OB to axonal growth.
Carbachol Potentiates Q Current and Activates a Calcium‐dependent Non‐specific Conductance in Rat Hippocampus In Vitro
Authors: Colino A. , Halliwell J. V.
Published: April 2006

Intracellular recordings were made from CA1 neurons in rat hippocampal slices maintained in vitro. When Na+ currents were blocked with tetrodotoxin and K+ conductances known to be sensitive to suppression by muscarinic agonists were blocked by 2 mM Ba2+, CA1 cells were depolarized by carbachol (3 – 10 μM) with an attendant conductance increase, whereas prior to Ba2+ the agonist produced a decrease or no change in conductance. Under voltage clamp at ∼–60 mV and in the presence of tetrodotoxin and Ba2+, carbachol (3 – 10 μM) induced a variable‐latency biphasic inward current of up to 380 pA associated with a conductance increase of ∼50%. The first phase was associated with an increase (more than 2‐fold) of the Cs+‐sensitive, hyperpolarization‐activated cationic current, IQ. Carbachol also accelerated the kinetics of IQ at – 100 mV with an average 24% reduction in its activation time constant. The second phase reflected an additional inward current that was Cs+‐resistant, displayed little apparent voltage sensitivity and had a mean extrapolated reversal potential, determined in the presence of external Cs+ (>5 mM), of ∼–20 mV. In a small proportion of cells the second phase of inward current was followed (or overlapped) by an outward current, also associated with a conductance increase, which reversed at ∼–70 mV. These carbachol actions were prevented by extracellular 300 μM Cd2+ and 2 mM Mn2+, by high levels (>5 mM) of extracellular Mg2+ or Ca2+, and by omission of Ca2+ or reduction of extracellular Na+ to 25 mM by substitution of NaCl with Tris or N‐methyl‐d‐glucamine. Carbachol action was not mimicked by oxotremorine (≤60 μM), but was irreversibly blocked by this drug. Likewise, atropine (100 nM) irreversibly and gallamine (10 μM) reversibly antagonized carbachol's action. The action of carbachol was blocked shortly after prior exposure of slices to 2 – 5 mM caffeine. Chronic or acute incubation of slices with 2 mM Li+ potentiated (between 1‐ and 2‐fold) carbachol responses. The data indicate that muscarinic activation increases cationic flux by a calcium‐dependent potentiation of IQ and activation of a non‐selective conductance. The probability that inositol phospholipid metabolism is involved in triggering these events is discussed.
Induction of Fictive Locomotion by Sulphur‐containing Amino Acids in an In Vitro Newborn Rat Preparation
Authors: Houssaini Youssef Sqalli , Cazalets Jean‐René , Martini Fabienne , Clarac François
Published: April 2006

The role of the sulphur‐containing amino acids (SAAs) in the initiation of fictive locomotion was tested in an isolated spinal cord preparation from newborn rats. These substances were bath‐applied and the fictive locomotion was recorded in the lumbar ventral roots. It emerged from this study that all the compounds tested could trigger an organized pattern (alternating left and right bursts of activity) with a dose‐dependent response. However, specific frequency and concentration ranges were observed with each of these SAAs. Moreover, a clear‐cut difference between d and l isomers in the ability of the SAAs to induce this activity was observed: the SAAs of the d‐forms were found to be generally more potent than those of the l‐forms. The effects of the SAAs were found to be mediated by both NMDA and non‐NMDA receptors, since they were blocked in a dose‐dependent manner by the specific antagonists of these receptors. Moreover, it was observed that β‐p‐chlorophenylglutamic acid, an uptake inhibitor of homocysteic acid (HCA), potentiated the effect of exogenously applied HCA, which supports the idea that HCA may act as a transmitter. The sulphuric and non‐sulphuric amino acids were also classified in their order of potency. The most potent compound turned out to be d‐homocysteine sulphinic acid, while d‐cysteine sulphinic acid was the least potent. It also emerged that the maximal frequencies obtained with SAAs and excitatory amino acids were in the same range, which might correspond to the maximal limits of this system.
Axotomy‐induced Changes in Ca2+ Homeostasis in Rat Sympathetic Ganglion Cells
Authors: Sánchez‐Vives Maria V. , Valdeolmillos Miguel , Martínez Salvador , Gallego Roberto
Published: April 2006

Some of the marked biochemical and electrophysiological changes provoked by section of the axon in mature neurons suggest that the intracellular calcium concentration ([Ca2+]i) may be increased. We have measured the [Ca2+]i using the fluorescent indicator Indo‐1 microinjected into rat superior cervical ganglion neurons. No differences in resting [Ca2+]i levels were found between control neurons and cells which had been axotomized 7–10 days before. However, the rise in [Ca2+]i evoked by orthodromic or antidromic stimulation and the recovery after the stimulating train were considerably slower in axotomized neurons than in control cells. We also found that the number of calbindin‐D28k‐immunopositive cells in the ganglion increases after axotomy, which could be related to the observed differences in calcium homeostasis.
Differentiation of Transmitter Phenotypes in Rat Cerebral Cortex
Authors: Götz Magdalena , Bolz Jürgen
Published: April 2006

Cortical neurons differ in their neurochemical properties. Projection neurons use excitatory amino acids as transmitters, most local interneurons contain the inhibitory transmitter GABA, and specific subtypes of local circuit neurons express distinct neuropeptides. How this cellular diversity is generated during development is not known. We have been studying the transmitter differentiation of cortical neurons in different in vitro systems using immunohistochemical techniques. Transmitter phenotypes of cortical neurons were examined in slice cultures, i.e. in the absence of extrinsic cortical connections, and in dissociated cortical cell cultures, i.e. in the absence of extrinsic and intrinsic cortical connections. The expression of vasoactive intestinal polypeptide in cortical interneurons occurred normally in slice cultures prepared from neonatal rats between birth and 2 days of age, but was strongly impaired in dissociated cell cultures prepared at the same time. These results suggest that the intact cortical environment present in the slice cultures exerts crucial influences for neuropeptide differentiation. In contrast, the transmitters glutamate and GABA were expressed normally in the appropriate cell types and similar in proportions in dissociated cell cultures prepared from cortices at embryonic day 19. Only cells dissociated during S‐phase failed to express glutamate and GABA in vitro. When cells were kept for 24 h after mitosis in a cortical slice preparation in vitro, however, they later expressed their appropriate transmitter phenotypes. Thus, signals from the local cortical environment that act early in the cell cycle are required for the specification of transmitter phenotypes of cortical neurons.
Purinergic Modulation of the Evoked Release of [3H]Acetylcholine from the Hippocampus and Cerebral Cortex of the Rat: Role of the Ectonucleotidases
Authors: Cunha Rodrigo A. , Ribeiro J. A. , Sebastião A. M.
Published: April 2006

Modulation by exogenous and endogenous adenine nucleotides and adenosine of [3H]acetylcholine release evoked by veratridine (10 μM) was compared in synaptosomal fractions from the hippocampus and the cerebral cortex of the rat. In both brain areas, exogenously added ATP or adenosine (10–100 μM) inhibited the evoked tritium release. In the hippocampus, ATPμS, an ATP analogue more resistant to catabolism than ATP, was virtually devoid of effect on tritium release, and the effect of ATP was prevented by the ecto‐5′‐nucleotidase inhibitor α,β‐methylene ADP (100 μM), by adenosine deaminase (2 U/ml) and by the A1 adenosine receptor antagonist 1,3‐dipropyl‐8‐cyclopentylxanthine (DPCPX, 20 nM). In contrast, in the cerebral cortex, the effect of ATP on tritium release was not prevented by either α,β‐methylene ADP (100 μM) or adenosine deaminase (2 U/ml), and several ATP analogues (30 μM) inhibited release. The order of intensity of the inhibitory effects of the ATP analogues was: ATPγS > ATP > ñ,γ‐imido ATP > β,γ‐methylene ATP > 2‐methyl‐S‐ATP, α,β‐methylene ATP. The effect of ATPγS in the cerebral cortex was prevented by DPCPX (20 nM) and was not affected by the P2 purinoceptor antagonist suramin (100 μM). In the hippocampus, α,β‐methylene ADP (100 μM) increased the evoked release of tritium, and adenosine deaminase (2 U/ml) produced an even greater increase; when adenosine deaminase was added in the presence of α,β‐methylene ADP, adenosine deaminase still increased the evoked release of tritium. In the cerebral cortex, DPCPX (20 nM) and adenosine deaminase (2 U/ml) increased the evoked tritium release by a similar magnitude, but the effect of adenosine deaminase was smaller than in the hippocampus. It is concluded that in the cerebral cortex ATP as such presynaptically inhibits acetylcholine release, whereas in the hippocampus the role of adenine nucleotides is as a source of endogenous extracellular adenosine that tonically inhibits acetylcholine release. The results also show that besides formation of adenosine from adenine nucleotides, released adenosine as such contributes in nearly equal amounts to the pool of endogenous adenosine that presynaptically inhibits acetylcholine release in the hippocampus.
Differential Localization and pH Dependency of Phosphoinositide 1,4,5‐IP3, 1,3,4,5‐IP4 and IP6 Receptors in Rat and Human Brains
Authors: Parent Angèle , Quirion Rémi
Published: April 2006

It is well established that the inositol lipids mediate signal transduction in several cellular populations. Many neurotransmitters, hormones and growth factors act at plasma membrane receptors to induce the hydrolysis of phosphatidylinositols and hence the generation of various inositol phosphates (IP). The best known member of this family is 1,4,5‐IP3, which is associated with the release of Ca2+ from intracellular pools. It has also been proposed that two others inositides, 1,3,4,5‐IP4 and IP6, may be involved in Ca2+ homeostasis. In order to study the possible relevance of these various inositides in neuronal tissues, we have localized the respective receptors in rat and human brain under both acidic and basic pH conditions. In the hippocampal formation, [3H]1,3,4,5‐IP4 binding sites are concentrated in the hilus and the molecular layer while a clearly different pattern of distribution is seen for [3H]1,4,5‐IP3, its highest concentration of labelling being concentrated in the oriens and radiatum laminae. This contrasting profile of distribution is also observed in other brain areas such as the caudate‐putamen, the septo‐hippocampal area, and the molecular and granular layers of the cerebellum. Moreover, while highest amounts of specific [3H]1,4,5‐IP3 binding are obtained at pH 8.5, the opposite is found for [3H]1,3,4,5‐IP4, with high binding levels seen under acidic conditions. [3H]IP6 binding sites are broadly distributed with specific labelling concentrated in areas enriched with neuronal perikarya such as the granular cell layer of the dentate gyrus, the pyramidal cell layers of the hippocampus and the granular cell layer of the cerebellum. [3H]IP6 binding is particularly abundant at basic pH in a few regions such as the accessory olfactory bulb, the piriform cortex and the septo‐hippocampal nucleus. The pH dependency of each inositide binding may be correlated to changes in intracellular pH that are observed following the activation of phospholipase C. The apparent distribution of specific 1,4,5‐IP3, 1,3,4,5‐IP4 and IP6 binding sites is rather similar in rat and human brains, suggesting their preserved functions across these two species.
Neonatal Sciatic Nerve Section Results in a Rearrangement of the Central Terminals of Saphenous and Axotomized Sciatic Nerve Afferents in the Dorsal Horn of the Spinal Cord of the Adult Rat
Authors: Shortland Peter , Fitzgerald Maria
Published: April 2006

Previous studies have shown that following neonatal peripheral nerve injury, adjacent intact myelinated and unmyelinated primary afferents sprout into the central denervated terminal area. The present study investigates this in more detail and goes further, to study the fate of the central terminals of the surviving axotomized primary afferent neurons. Bulk labelling of the sciatic and saphenous nerves with horseradish peroxidase conjugated to choleragenoid (B‐HRP), to label the A fibres, or wheatgerm agglutinin (WGA‐HRP), to label C fibres were employed to investigate the central consequences of sciatic nerve section and ligation on the day of birth, in adult rats. Bulk labelling of the axotomized sciatic or intact saphenous nerve with either tracer and comparison with contralateral controls revealed alterations to the terminal field. The intact saphenous nerve terminal field expanded caudally from mid L4 to the L4‐L5 boundary when labelled with WGA‐HRP and to the sacral cord when labelled with B‐HRP. Labelling the axotomized sciatic nerve with either tracer revealed little change in the overall somatotopic organization of central terminals, although labelling was less intense compared to control nerves and more variable with WGA‐HRP. Invasion of the substantia gelatinosa (SG) by axotomized A fibres was observed in segments L3‐5, into the area occupied by axotomized C fibres. This area was also invaded by intact saphenous A fibres in the L4–5 segments. These results demonstrate that following neonatal nerve section: (i) axotomized primary afferents are able to retain a ‘normal’ somatotopic map in the rostrocaudal plane; (ii) both A and C fibres from adjacent intact nerves sprout into the denervated territory, but A fibres sprout further caudally; (iii) axotomized A fibres and invading intact A fibres both sprout dorsally into denervated SG. As a result, there is considerable overlap between nerve territories in denervated spinal cord, suggesting that competition for laminar termination sites exists between A and C fibres and also between axotomized and intact primary afferents.
Localization of Oxytocin Binding Sites in the Thoracic and Upper Lumbar Spinal Cord of the Adult and Postnatal Rat: A Histoautoradiographic Study
Authors: Reiter M. K. , Kremarik P. , Freund‐Mercier M. J. , Stoeckel M. E. , Desaulles E. , Feltz P.
Published: April 2006

Oxytocin binding sites were detected by autoradiography on films and emulsion‐coated sections in the spinal cord of adult and postnatal rats from C8 to L2, using a highly selective 125l‐labelled oxytocin antagonist. Oxytocin binding sites were detected on all transverse sections in the dorsal horn, where labelling was scattered over laminae I and II. The autonomic areas, i.e. the intermediolateral cell column, the central grey (lamina X) and the nucleus intercalatus were labelled. Binding in the intermediolateral cell column was most frequently observed on sections from T9 to T11 in adult and T7 to T8 in postnatal rats. In this location, oxytocin binding sites were highly concentrated on cell bodies of putative sympathetic preganglionic neurons; however, not all of these cells were labelled. Diffuse labelling occurred on the dorsal part of the central grey, mainly between T8 and L2. Isolated labelled cells belonging to the nucleus intercalatus were scattered between the central canal and the intermediolateral cell column. In addition, oxytocin binding sites were found on some motoneurons of the lateral group of T12‐T13, but only in postnatal rats. The distribution of oxytocin binding sites in the rat spinal cord coincides with that of the oxytocin innervation and strongly suggests a modulatory role of this peptide in sensory and autonomic functions.
Rapid Desensitization of the Metabotropic Glutamate Receptor that Facilitates Glutamate Release in Rat Cerebrocortical Nerve Terminals
Authors: Herrero I. , Miras‐Portugal M. T. , Sánchez‐Prieto J.
Published: April 2006

The metabotropic autoreceptor of glutamatergic nerve terminals from the cerebral cortex of adult rats has been characterized. Receptor activation involves a rapid and transient increase in diacylglycerol, which is sensitive to l‐2‐amino‐3‐phosphonopropionate (l‐AP3) and l‐2‐amino‐4‐phosphonobutanoic acid (l‐AP4) and is partially blocked by pertussis toxin. Protein kinase C (PKC) has a negative feedback control in this transduction pathway because the activation of the kinase, either by phorbol esters or by the endogenous diacylglycerol produced by the receptor, results in a reversible receptor desensitization, with loss of the ability to further facilitate glutamate release. It is concluded that the facilitatory metabotropic receptor located at the glutamatergic nerve endings belongs to the subclass coupled to phosphoinositide hydrolysis and that the rapid and use‐dependent desensitization of the facilitatory pathway may underlie a mechanism to prevent its permanent activation and thereby to avoid neurotoxicity.
Embryonic Purkinje Cells Grafted on the Surface of the Adult Uninjured Rat Cerebellum Migrate in the Host Parenchyma and Induce Sprouting of Intact Climbing Fibres
Authors: Rossi Ferdinando , Borsello Tiziana , Strata Piergiorgio
Published: April 2006

By grafting solid pieces of cerebellar anlage onto the surface of the adult rat cerebellum, we have investigated the problem of the interactions between embryonic and adult neurons in an intact brain. A few days after grafting, embryonic astrocytic processes crossed the graft‐host interface and radiated into the recipient molecular layer. Several grafted Purkinje cells also migrated into the host brain along such processes as well as adult Bergmann glia. Adult climbing fibres, labelled by means of Phaseolus vulgaris leucoagglutinin (PHA‐L), sprouted new collateral branches which terminated on embryonic Purkinje cells at both extra‐ and intraparenchymal levels. No sign of activation of host astroglia or microglia was evident in the host cerebellum in relation to these processes. Embryonic Purkinje cells which migrated into the host cerebellum developed an adult‐like morphology. Intraparenchymal grafts of neocortical embryonic tissue induced conspicuous growth of host olivary axons, characterized by a pattern which was different from that observed following cerebellar grafts. By contrast, when neocortical tissue was placed onto the surface of the recipient cerebellum, graft‐host interactions were limited and climbing fibre sprouting was rarely seen. These results show that (i) supernumerary Purkinje cells can penetrate and settle in the adult intact cerebellar cortex, (ii) adult climbing fibres are able to innervate these new targets in the absence of any injury or activation of non‐neuronal cells of the adult brain, and (iii) in the absence of damage to the adult brain, the plasticity of adult olivary axons is specifically elicited and controlled by embryonic Purkinje cells.