bluuebunny/crossref_metadata_2025 huggingface dataset:
The Influence of Premotor lnterneuron Populations on the Frequency of the Spinal Pattern Generator for Swimming in Xenopus Embryos: A Simulation Study
Authors: Wolf E. , Roberts A.
Published: April 2006

Our aim was to test the hypothesis that the frequency of neuronal rhythm‐generating networks is partly controlled by the size of the active premotor interneuron population. We have tested possible mechanisms for frequency changes in a population model of the Xenopus laevis embryo spinal rhythm‐generating networks for swimming. After initiation by a brief sensory excitation, the frequency of swimming activity decreases to a steady level determined by the properties of the 24 interneurons and their connections. The initial frequency decrease was dependent on the time‐course of initiating sensory synaptic excitation. When some premotor excitatory interneurons were given weaker synaptic connections to reflect the variability in the spinal cord, they could drop out and stop firing during the initial frequency decrease while swimming activity continued. If the synaptic input of such weak excitatory interneurons was graded finely, they could drop out consecutively. This led to further decreases in the level of tonic excitation and in network frequency which depended on the number, type and distribution of excitatory interneurons that stopped firing. Silent weak excitatory interneurons could be recruited by a second sensory excitation and cause an increase in tonic depolarization and frequency which outlasted the sensory input. Such recruitment could occur on both sides after local sensory stimulation to only one region or one side of the model. We conclude that these computer simulations support the hypothesis that premotor interneuron drop‐out and recruitment is one mechanism which can control frequency in a locomotor central pattern generator.
Ontogeny of the D1 Dopamine Receptor in the Rat Striatonigral System: an lmmunohistochemical Study
Authors: Caille I. , Dumartin B. , Le Moine C. , Begueret J. , Bloch B.
Published: April 2006

Antibodies were raised against a recombinant protein to analyse the pre‐ and postnatal ontogeny of the neurons expressing the D1 dopamine receptor in the striatum by immunohistochemistry. We report that D1 immunoreactivity is detectable from gestational day (G) 15 and is distributed homogeneously throughout the striatum from G15 to G18. From G19–20 to postnatal day (P) 3, D1 immunoreactivity becomes heterogeneous and predominates in cell bodies of the patch Compartment while very limited immunoreactivity is detectable in the matricial compartment. The differential intensity between patches and matrix reaches its peak around PO. From P2, the pattern of D1 immunoreactivity progressively assumes the homogeneous distribution characteristic of the adult striatum. The expression of D1 mRNA in striatal neurons, as investigated by in situ hybridization, displays a similar pattern during this period. Substance P mRNA is also preferentially expressed in the patch compartment during the same period. D1 immunoreactivity appears at G17 in the substantia nigra as clusters of fibres and increases subsequently until reaching its adult form during the first postnatal week. These results demonstrate that the two compartments of the developing striatum display differential transcriptional and translational activity for the D1 gene and consequently two different and successive patterns of expression of D1 protein: patch neurons first express D1 receptor intensely while matrix neurons express it later and in smaller amounts so that D1 receptor appears transiently during the perinatal period as a marker of the patch compartment.
Axonal Target Choice and Dendritic Development of Ferret Beta Retinal Ganglion Cells
Authors: Wingate Richard J. T. , Thompson Ian D.
Published: April 2006

We have investigated the relationship between axon targeting and dendritic morphology in beta retinal ganglion cells in the postnatal ferret. Axonal projections were assessed by making separate injections of different fluorescent retrograde tracers into either the superior colliculus or lateral geniculate nucleus in viva The dendritic morphology of retrogradely labelled cells was revealed by the in vitro intracellular injection of Lucifer yellow in fixed retina. In this way, 405 retinal ganglion cells were triple‐ or double‐labelled and characterized by their dendritic branching styles. Both the distinct dendritic morphology of beta cells and the characteristic restriction of their adult axonal terminals to the lateral geniculate nucleus emerge postnatally. Beta cell dendritic morphology is established between postnatal days 5 and 9. As in the cat (Ramoa et al, 1989), beta cells extend and then retract a projection to the superior colliculus as part of their normal development. Transient beta axonal collaterals to the superior colliculus persist beyond the period of cell death, but nearly all are withdrawn by postnatal day 15. No dendritically distinct beta cell projects to the superior colliculus alone, at any age.

Heterochronic injections of different colours of retrograde tracer into the superior colliculus were used to study changes in the complement of the retinocollicular projection over time. A significant proportion of cells (58%) labelled at postnatal day O from the superior colliculus, which subsequently survived the period of cell death, were found to be beta cells that could no longer be demonstrated to have a retinocollicular axon. Neonatal decortication, which reduced the volume of the adult lateral geniculate nucleus by 36–86%, resulted in the limited stabilization of this normally transient beta cell projection to the superior colliculus. The fundamental dendritic branching style of beta ganglion cells is unchanged in decorticate ferrets, suggesting that it develops independently of their ultimate axonal target choice.
Development and Pharmacological Suppression of Secondary Afterdischarges in the Hippocampus of Amygdala‐kindled Rats
Authors: Ebert Ulrich , Rundfeldt Chris , Löscher Wolfgang
Published: April 2006

The development and spread of afterdischarges in the ipsilateral limbic system during amygdala kindling, a model of complex partial seizures, was studied in male and female rats. Kindling stimulation was performed in the basolateral amygdala, and afterdischarges were recorded from the stimulation electrode and electrodes in the nucleus accumbens, the posterior piriform cortex and the ventral hippocampus, all implanted on the right side of the brain. All structures showed primary afterdischarges already after the first stimulation, indicating a close anatomical and physiological connection to the epileptogenic focus. The development of robust secondary afterdischarges, which occurred after the end of the primary afterdischarges in the amygdala and which always originated in the hippocampus but also spread to one or more of the other recording sites, is described. The secondary afterdischarges initially occurred after about nine kindling stimulations in both male and female rats, and were associated with an increase in primary afterdischarge duration and a progression from focal to motor seizures. In order to test the effect of common antiepileptic drugs on the secondary afterdischarges, a group of female rats were treated with valproate, carbamazepine or phenytoin. All drugs suppressed the secondary afterdischarges, although they had a different anticonvulsant efficacy on motor seizures and afterdischarge duration after amygdala stimulation. While valproate and carbamazepine dose‐dependently reduced all parameters of the kindled seizure, including the secondary afterdischarges in the hippocampus, phenytoin suppressed the secondary afterdischarges also in the absence of any anticonvulsant effect, suggesting that recurrent hippocampal activation is not crucial for the kindled state. Recording of secondary afterdischarges in the hippocampus may offer the possibility of studying the conditions for development and pharmacological suppression of recurrent hippocampal activation in amygdala‐kindled rats.
Expression of the Homeobox‐containing Gene En‐2 During the Development of the Chick Central Nervous System
Authors: Millet Sandrine , Alvarado‐Mallart Rosa‐Magda
Published: April 2006

The expression of the homeobox‐containing gene En‐2 was analysed with the monoclonal antibody 4D9 in the chick central nervous system throughout embryogenesis. Confirming previous studies, early expression of the En‐2 protein [beginning at stage 9 of Hamburger and Hamilton (HH9)] is restricted to a portion of the neural tube containing the primordia of the cerebellum, the isthmic region and the mesencephalic grisea, and forms a double gradient decreasing both caudally and rostrally from a high point located around the midbrain‐hindbrain constriction. This mes‐isthmo‐cerebellar region contains all the En‐2‐positive germinative cells and the great majority of the En‐2‐positive postmitotic neurons throughout embryogenesis. Nevertheless, as the postmitotic neurons appear, En‐2 expression also occurs outside this region: in two columns of non‐motoneuron cells in rhombomeres two to four (between HH20 and HH30) and, from HH24 onwards, throughout the grey matter of the lumbar and thoracic spinal cord, with the exception of the ventral motoneuron columns. Here, a detailed description of En‐2 expression is provided for the mes‐isthmo‐cerebellar region at stages HH30–32 [embryonic day (E) 71, HH37 (E11) and HH46 (E21, hatching). This allows the visualization of cellular groups with heterogeneous patterns of En‐2 expression, which are specific for each group in the intensity of En‐2 expression, the distribution of the labelled cells and the temporal regulation of the gene. The use of tyrosine hydroxylase antiserum shows coexpression of the tyrosine hydroxylase enzyme and En‐2 protein in the caudal part of the nuclei tegmenti pedunculo‐pontinus, the area ventralis of Tsai and the substantia grisea centralis, but not in the locus coeruleus. In the cerebellum, the first expression, which is located in the deep nuclei and parasagittal bands of Purkinje cells, is down‐regulated when the molecular layer interneurons and the granular cells begin to express the gene, at the end of embryogenesis. Finally, at hatching, En‐2 expression permits the visualization in the cerebellum of a population of small En‐2‐negative cells located around the Purkinje cells that may correspond to those described in chick/quail chimaeras as having an origin different from that of the bulk of granular neurons.
Antibodies to Postsynaptic PKC Substrate Neurogranin Prevent Long‐term Potentiation in Hippocampal CA1 Neurons
Authors: Fedorov N. B. , Pasinelli P. , Oestreicher A. B. , DeGraan P. N. E. , Reymann K. G.
Published: April 2006

Both pre‐ and postsynaptic protein kinase C have been implicated in long‐term potentiation. Neurogranin (also known as BICKs and RC3) is a neuronal postsynaptic protein kinase C substrate. In the present study we injected monoclonal IgGs that recognize the protein kinase C phosphorylation site in neurogranin and B‐50 (GAP‐43), and that have been shown to inhibit protein kinase C‐mediated B‐50 phosphorylation, through a whole‐cell clamp pipette into CA1 pyramidal neurons in rat hippocampal slices. Injection of neurogranin IgGs, but not of control IgGs, prevented the induction of tetanus‐induced long‐term potentiation without affecting posttetanic potentiation. Our results suggest that neurogranin is involved in mechanisms of activity‐dependent synaptic plasticity.
Laufband Therapy Based on‘Rules of Spinal Locomotion’is Effective in Spinal Cord Injured Persons
Authors: Wernig A. , Müller S. , Nanassy A. , Cagol E.
Published: April 2006

Rehabilitation of locomotion in spinal cord (s.c.) injured patients is unsatisfactory. Here we report the effects of a novel ‘Laufband (LB; treadmill) therapy’ based on ‘rules of spinal locomotion’ derived from lower vertebrates. Eighty‐nine incompletely paralysed (44 chronic and 45 acute) para‐ and tetraplegics underwent this therapy, then were compared with 64 patients (24 chronic and 40 acute) treated conventionally. The programme consisted of daily upright walking on a motor driven LB initially with body weight support (BWS) provided by a harness and assisted limb movements by the therapists when necessary. Forty‐four chronic patients with different degrees of paralysis undertook the programme for 3‐20 weeks (median = 10.5), 0.5–18 years after S.C. damage. At the onset of LB therapy 33/44 patients were wheelchair‐bound (no standing and/or walking without help by others) compared with 25 at completion of LB therapy, i.e. 76% had learned to walk independently, 7 patients with help. Only 1 subject did not improve. It was striking that voluntary muscle activity in the resting position was still low in several patients who had gained walking capability. Eleven patients who could already walk before LB therapy improved in speed and endurance. Of the 44 patients, six were capable of staircase walking before LB therapy compared with 34 afterwards. In order to validate the apparent superiority of LB therapy two types of comparisons were performed. In a ‘temporal’ control 12 spastic paretic patients, still wheelchair‐bound after the period of postacute conventional therapy, performed LB immediately thereafter. After completion of LB therapy nine of these patients had learned to walk without help from others. In another control, two groups of chronic patients, matched in type of injury, time after injury (1 or more years) and history of previous rehabilitation (one or several periods of conventional therapy), were compared. One group underwent LB therapy (n= 29), the other conventional therapy (n= 24) for similar periods of time. From 18 wheelchair‐bound patients, 14 became independent walkers after LB, but only 1/14 after conventional therapy. From 45 acute patients 92% (33/36) of those wheelchair‐bound at the onset of LB therapy became independent, but only 50% (12/24) after conventional therapy (n= 40). It is noteworthy that voluntary muscle activity at rest was similar in both groups. The presumed underlying mechanisms are discussed in the light of the absence of any apparent effects in seven functionally completely paralysed paraplegic persons.
Electron Microscopic Observation of Substance P‐like Immunoreactive Nerve Fibres and Innervation in the Anterior Pituitary of Macaques
Authors: Liu Shaojun
Published: April 2006

The anterior pituitary has recently been confirmed to be innervated by substance P‐immunoreactive nerve fibres in the monkey and human. The present study investigated the morphology of the nerve fibres and their relationship with anterior pituitary gland cells in the macaque by means of immunoelectron microscopy. Many substance P‐immunoreactive and unlabelled nerve fibres were distributed among the gland cells or formed nerve fibre bundles. These nerve fibres were unmyelinated and varicose, and contained a great many clear and/or large dense‐cored vesicles. Substance P‐immunoreactive cells were seen in the pars distalis of the anterior pituitary. Direct contacts between these nerve fibres and every cell type of the gland could be ascertained, including substance P‐immunoreactive cells. Synaptoid contacts were seen among the substance P‐immunoreactive varicosities and somatotropes, opiocortico‐melanotropes and folliculostellate cells. Typical synapses were identified in somatotropes and opiocortico‐melanotropes.
Vulnerability of Medium Spiny Striatal Neurons to Glutamate: Role of Na+/K+ ATPase
Authors: Calabresi Paolo , De Murtas Marco , Pisani Antonio , Stefani Alessandro , Sancesario Giuseppe , Mercuri Nicola B. , Bernardi Giorgio
Published: April 2006

In Huntington's disease neuronal degeneration mainly involves medium‐sized spiny neurons. It has been postulated that both excitotoxic mechanisms and energy metabolism failure are implicated in the neuronal degeneration observed in Huntington's disease. In central neurons, >40% of the energy released by respiration is used by Na+/K+ ATPase to maintain ionic gradients. Considering that impairment of Na+/K+ ATPase activity might alter postsynaptic responsivity to excitatory amino acids (EAAs), we investigated the effects of the Na+/K+ ATPase inhibitors, ouabain and strophanthidin, on the responses to different agonists of EAA receptors in identified medium‐sized spiny neurons electrophysiologically recorded in the current‐ and voltage‐clamp modes. In most of the cells both ouabain and strophanthidin (1–3 μM) did not cause significant change in the membrane properties of the recorded neurons. Higher doses of either ouabain (30 μM) or strophanthidin (30 μM) induced, per se, an irreversible inward current coupled to an increase in conductance, leading to cell deterioration. Moreover, both ouabain (1–10 μM) and strophanthidin (1–10 μM) dramatically increased the membrane depolarization and the inward current produced by subcritical concentrations of glutamate, AMPA and NMDA. These concentrations of Na+/K+ ATPase inhibitors also increased the membrane responses induced by repetitive cortical activation. In addition, since it had previously been proposed that dopamine mimics the effects of Na+/K+ ATPase inhibitors and that dopamine agonists differentially regulate the postsynaptic responses to EAAs, we tested the possible modulation of EAA‐induced membrane depolarization and inward current by dopamine agonists. Neither dopamine nor selective dopamine agonists or antagonists affected the postsynaptic responses to EAAs. Our experiments show that impairment of the activity of Na+/K+ ATPase may render striatal neurons more sensitive to the action of glutamate, lowering the threshold for the excitotoxic events. Our data support neither the role of dopamine as an ouabain‐like agent nor the differential modulatory action of dopamine receptors on the EAA‐induced responses in the striatum.
Properties of the Ryanodine‐sensitive Release Channels that Underlie Caffeine‐induced Ca2+Mobilization from Intracellular Stores in Mammalian Sympathetic Neurons
Authors: Hernández‐Cruz A. , Díaz‐Muñoz M. , Gómez‐Chavarín M. , Canñedo‐Merino R. , Protti D. A. , Escobar A. L. , Sierralta J. , Suárez‐Isla B. A.
Published: April 2006

The most compelling evidence for a functional role of caffeine‐sensitive intracellular Ca2+reservoirs in nerve cells derives from experiments on peripheral neurons. However, the properties of their ryanodine receptor calcium release channels have not been studied. This work combines single‐cell fura‐2 microfluorometry, [3H]ryanodine binding and recording of Ca2+release channels to examine calcium release from these intracellular stores in rat sympathetic neurons from the superior cervical ganglion. Intracellular Ca2+measurements showed that these cells possess caffeine‐sensitive intracellular Ca2+stores capable of releasing the equivalent of 40% of the calcium that enters through voltage‐gated calcium channels. The efficiency of caffeine in releasing Ca2+showed a complex dependence on [Ca2+]i. Transient elevations of [Ca2+]iby 50–500 nM were facilitatory, but they became less facilitatory or depressing when [Ca2+]ireached higher levels. The caffeine‐induced Ca2+release and its dependence on [Ca2+]iwas further examined by [3H]ryanodine binding to ganglionic microsomal membranes. These membranes showed a high‐affinity binding site for ryanodine with a dissociation constant (KD= 10 nM) similar to that previously reported for brain microsomes. However, the density of [3H]ryanodine binding sites (Bmax= 2.06 pmol/mg protein) was at least three‐fold larger than the highest reported for brain tissue. [3H]Ryanodine binding showed a sigmoidal dependence on [Ca2+] in the range 0.1–10 μM that was further increased by caffeine. Caffeine‐dependent enhancement of [3H]ryanodine binding increased and then decreased as [Ca2+] rose, with an optimum at [Ca2+] between 100 and 500 nM and a 50% decrease between 1 and 10 μM. At 100 μM [Ca2+], caffeine and ATP enhanced [3H]ryanodine binding by 35 and 170% respectively, while binding was reduced by >90% with ruthenium red and MgCl2. High‐conductance (240 pS) Ca2+release channels present in ganglionic microsomal membranes were incorporated into planar phospholipid bilayers. These channels were activated by caffeine and by micromolar concentrations of Ca2+from the cytosolic side, and were blocked by Mg2+and ruthenium red. Ryanodine (2 μM) slowed channel gating and elicited a long‐lasting subconductance state while 10 mM ryanodine closed the channel with infrequent opening to the subconductance level. These results show that the properties of the ryanodine receptor/Ca2+release channels present in mammalian peripheral neurons can account for the properties of caffeine‐induced Ca2+release. Our data also suggest that the release of Ca2+by caffeine has a bell‐shaped dependence on Ca2+in the physiological range of cytoplasmic [Ca2+].
Agonist‐induced Down‐regulation of NMDA Receptors in Cerebellar Granule Cells in Culture
Authors: Resink A. , Villa M. , Boer G. J. , Möhler H. , Balázs R.
Published: April 2006

In contrast to the acute toxic effect of NMDA on mature cerebellar granule cells, chronic treatment with NMDA (140 μM from 1 to 9 days in vitro) did not compromise cell survival. Such treatment markedly suppressed NMDA receptor activity: at 8 days in vitro NMDA‐induced 45Ca2+ influx was reduced by ‐60% and acute exposure to NMDA (highest concentration tested, 1 mM) at 9 days in vitro did not cause detectable toxicity. The reduction in NMDA receptor activity was accompanied by a significant decrease (±80% at 9 days in vitro) in the level of the NR1 and the NR2A NMDA receptor subunit protein, detected using the selective photoaffinity ligand [125I]CGP55802A. It seems, therefore, that the agonist‐induced decrease in NMDA receptor activity is due to receptor down‐regulation. In contrast to the marked influence of chronic NMDA exposure on the cellular content of the NMDA receptor subunit proteins, mRNA levels of the different subunits (NR1, NR2A, NR2B and NR2C) were not significantly affected. It seems, therefore, that agonist‐induced down‐regulation of the NMDA receptor involves critically mRNA translation and/or post‐translational regulation.
Fibroblast Growth Factors Regulate Calcitonin Gene‐related Peptide mRNA Expression in Rat Motoneurons after Lesion and in Culture
Authors: Piehl Fredrik , Ru‐Rong Ji , Cullheim Staffan , Hökfelt Tomas , Lindholm Dan , Hughes Richard A.
Published: April 2006

In this study, we have investigated the effect of fibroblast growth factors (bFGF and FGF‐5) and brain derived neurotrophic factor (BDNF) on the expression of calcitonin gene‐related peptide (CGRP) in rat motoneurons in vivo and in vitro. Following sciatic nerve transection in adult rats, the levels of α‐CGRP and β‐CGRP mRNA were up‐ and down‐regulated respectively in axotomized motoneurons, revealed by in situ hybridization histochemistry. Local administration of 1 μg bFGF was able to entirely abolish the up‐regulation of α‐CGRP mRNA, and to further down‐regulate β‐CGRP. These effects, albeit less pronounced, were still evident with 0.2 μg bFGF. In contrast, bFGF did not attenuate the lesion‐induced decrease of choline acetyltransferase (ChAT) mRNA. Administration of BDNF did not significantly alter the expression of CGRP or ChAT mRNA in axotomized motoneurons. Both α‐ and β‐CGRP mRNAs could be detected by PCR in enriched motoneuron cultures prepared from rat embryos at embryonic day 14–15. Comparing the amplification of α‐ and β‐CGRP mRNAs with that of mRNA encoding glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) in parallel samples, we found that cultures treated with FGF‐5 had a lower ratio of α‐ and β‐CGRP mRNA to GAPDH mRNA, than did control or BDNF‐treated cultures. BDNF, on the other hand increased α‐CGRP and decreased β‐CGRP mRNA levels, though these effects were moderate compared with the effects of FGF‐5. The results obtained in this study suggest that members of the FGF family of growth factors influence the expression of CGRP in rat motoneurons, and that the increase of this neuropeptide induced by axotomy may, at least in part, be due to deprivation of these target‐derived factors.
All‐or‐none Excitatory Postsynaptic Potentials in the Rat Visual Cortex
Authors: Volgushev Maxim , Voronin Leon L. , Chistiakova Marina , Artola Alain , Singer Wolf
Published: April 2006

Intracellular recordings were obtained from supragranular neurons in slices of the rat visual cortex. In ∼25% of the cells large (0.5–1.6 mV) excitatory postsynaptic potentials (EPSPs) of constant amplitude were observed after minimal, presumably single‐fibre stimulation. The amplitude variance of these large EPSPs was surprisingly small and within the range of the variance of the noise. These EPSPs could be reduced in amplitude by paired‐pulse and low‐frequency stimulation or by raising extracellular Mg2+ concentration. Reduced EPSPs could either continue to behave as all‐or‐none responses, or they could fluctuate between several amplitude levels. Conversely, responses where the amplitude fluctuated from trial to trial under control conditions could be converted into large all‐or‐none responses by paired‐pulse facilitation. This indicates that the large all‐or‐none EPSPs were composed of several subunits, probably reflecting the action of several different release sites. It is concluded that these release sites are either independent and operate with a probability close to 1 or, if operating with a lower probability, are coordinated by a mechanism which synchronizes release. Several observations suggest that release probabilities can switch from values close to 1 to 0 with repetitive stimulation or high Mg2+ concentration. Thus, a substantial fraction of single‐fibre inputs to supragranular cells possess synapses which operate with high synaptic efficiency and extremely low variance under control conditions but can undergo drastic changes in efficacy when release probabilities are interfered with. Such modifications of release probability could serve as an effective mechanism to regulate the gain of synaptic transmission.
TGF‐βs and cAMP Regulate GAP‐43 Expression in Schwann Cells and Reveal the Association of this Protein with the Trans‐Golgi Network
Authors: Stewart Helen J. S. , Curtis Rory , Jessen Kristjan R. , Mirsky Rhona
Published: April 2006

We have shown previously that growth‐associated protein 43 (GAP‐43) is expressed by rat Schwann cells and is restricted to non‐myelin‐forming Schwann cells in vivo. Here we examined the regulation of GAP‐43 using agents that are known to control Schwann cell differentiation in vitro. GAP‐43 protein and mRNA levels are decreased by forskolin and other agents that elevate intracellular cAMP (and promote expression of the myelinating Schwann cell phenotype). We also found that expression of GAP‐43 protein but not mRNA is down‐regulated by transforming growth factor betas (TGF‐βs). Moreover, TGF‐β treatment of Schwann cells results in cell clumping, process retraction and disappearance of GAP‐43 from the plasma membrane, revealing that GAP‐43 is associated with the Golgi apparatus. This association was confirmed by partial overlap of GAP‐43 with the trans‐Golgi network marker (23c) and the disruption of the Golgi with brefeldin A or monensin leading to altered GAP‐43 distribution. Golgi‐associated GAP‐43 appeared to have the same molecular weight as the plasma membrane‐associated GAP‐43. Thus these results show that GAP‐43 expression in Schwann cells is subject to regulation by both extracellular and intracellular signalling molecules and that Schwann cell GAP‐43 is often associated with the Golgi apparatus.
Stimulation of Ca2+‐Activated Non‐specific Cationic Channels by Phospholipase C‐linked Glutamate Receptors in Synaptoneurosomes?
Authors: Vignes Michel , Blanc Emmanuelle , Récasens Max
Published: April 2006

The regulation of intracellular Ca2+ concentration ([Ca2+]i) by glutamate metabotropic receptors (mGluR) was studied in 8‐day‐old rat forebrain synaptoneurosomes using spectrofluorimetric methods. Here we demonstrate that metabotropic glutamate agonists induce in rat brain synaptoneurosomes a Ca2+ influx largely dependent upon the presence of Ca2+ in the external medium. The pharmacological profile of this influx is strongly correlated with the pharmacological profile of the activation of phosphoinositide hydrolysis, i.e. quisqualic acid ∼ 1S,3R‐amino‐1‐dicarboxylate‐1,3 cyclopentane ≅ glutamate. This metabotropic glutamate receptor‐induced Ca2+ influx is insensitive to voltage‐dependent Ca2+ channel antagonists and occurs through a Mn2+ impermeant pathway. The study of the rapid kinetics shows that this influx is triggered after a 300 ms delay compared with that elicited by depolarizing agents and Ca2+ ionophore A23187. In order to assess further if mGluR stimulate this influx through the recruitment of inositol triphosphate (IP3)‐sensitive intracellular Ca2+ stores, we have tested the effect of thapsigargin on membrane potential and intracellular Ca2+ simultaneously. Thapsigargin induces a depolarization of the synaptoneurosomal membrane followed by a massive Ca2+ influx, occurring via a Mn2+ nonpermeant route. This depolarizing effect is sensitive to the presence of the intracellular Ca2+ chelator [1,2‐bis(2‐aminophenoxy)ethane‐N, N, N', N'‐tetraacetoxymethyl ester], and partially sensitive to extracellular Na+, but insensitive to the presence of extracellular Ca2+. Taken together, our data suggest that mGluR stimulate self‐maintained increases of [Ca2+]i in rat forebrain synaptoneurosomes via the activation of a multistep mechanism, sequenced in the following steps: (i) mGluR‐induced IP3 synthesis; (ii) IP3‐stimulated intracellular Ca2+ release; (iii) Ca2+‐activated non‐specific cation channel, leading to local depolarization and a Ca2+ influx; and (iv) activation of Ca2+‐sensitive phospholipase C.
Focal Adhesion Kinase in Rat Central Nervous System
Authors: Burgaya Ferran , Menegon Andrea , Menegoz Mathias , Valtorta Flavia , Girault Jean‐Antoine
Published: April 2006

Focal adhesion kinase (pp125FAK, FAK) is a 125 kDa non‐receptor tyrosine kinase enriched in focal adhesions of various cell types, where it is thought to transduce signals triggered by contact with the extracellular matrix. We have studied the expression and localization of FAK in rat CNS. Immunoblotting, immunohistochemistry and in situ hybridization revealed the presence of FAK in all regions of the adult brain and demonstrated its enrichment in specific neuronal populations of the cerebral and cerebellar cortex, as well as in the hippocampus. During development, FAK protein levels were highest around birth in cerebral cortex and caudate putamen and decreased in the adult. In situ hybridization revealed enrichment of FAK mRNA in the ventricular germinative and external layers during the last period of embryonic growth. In primary cultures FAK immunoreactivity was localized in focal adhesions in astrocytes, whereas in developing neurons the highest levels were found in growth cones and perikarya. In the growth cone, FAK immunoreactivity colocalized with actin filaments. In mature neurons FAK appeared to be distributed in the whole cytoplasm, with no enrichment in any cellular compartment. Our results demonstrate the presence of high levels of FAK in rat CNS, maximal during development but persistent in the adult. Its enrichment in growth cones suggests that it may play a role in neurite outgrowth, as well as in plasticity in the adult.
G‐Protein Modulation of Glycine‐resistant NMDA Receptor Desensitization in Rat Cultured Hippocampal Neurons
Authors: TureCek Rostislav , Vyklicky Ladislav , Vlachova Viktorie , Vyklicky Ladislav
Published: April 2006

Activation of N‐methyl‐D‐aspartate (NMDA) subtype glutamate receptors increases the excitability of most neurons within the CNS. A common feature of ionotropic glutamate receptors is their ability to undergo desensitization. In the present experiments we have examined the role of guanine nucleotide‐binding proteins(G‐proteins) in the regulation of NMDA receptor desensitization. Repeated NMDA receptor activation with 2 mM extracellular Ca2+ increased the degree of glycine‐resistant NMDA receptor desensitization of subsequent responses to NMDA recorded in the presence of 0.2 mM Ca2+. The recovery of glycine‐resistant NMDA receptor desensitization after repeated NMDA receptor activation in the presence of 2 mM Ca2+ was significantly reduced in neurons intracellularly dialysed with guanosine‐5'‐0‐(3‐thiotriphosphate), guanosine‐5'‐triphosphate or AIC13 and CsF, compounds known to activate G‐proteins. lntracellular dialysis with guanosine‐5'‐0(2thiodiphosphate), adenosine triphosphate, or adenosine‐5'‐G(3‐thiotriphosphate) was ineffective. The calcium permeability of NMDA receptor‐channels was not altered by intracellular dialysis with GTPyS. This suggests that modulation of NMDA receptor desensitization by G‐proteins represents a novel mechanism forregulation of glutamate‐gated ion channel activity.
Anti‐nociception Induced by Systemic or PAG‐microinjected Lysine‐acetylsalicylate in Rats. Effects on Tail‐flick Related Activity of Medullary Off‐ and On‐cells
Authors: Tortorici Victor , Vanegas Horacio
Published: April 2006

Previous experiments using metamizol have shown that this non‐steroidal anti‐inflammatory drug (NSAID) produces a central anti‐nociceptive effect probably through neural substrates that also support the analgesic effects of opiates, such as the periaqueductal grey matter (PAG) and the off‐ and on‐cells of the rostral ventromedial medulla (RVM). Off‐ and on‐cells have been postulated to respectively inhibit and facilitate nociceptive transmission, since the heat‐elicited tail flick reflex (TF) occurs only after off‐cells have decreased (pause), and on‐cells have increased (burst) their activity. The aim of the present study was to examine whether the effect of metamizol upon TF and off‐ and on‐cells responses could be generalized to other NSAIDs such as, in this case, lysine‐acetylsalicylate (LASA). Fifty‐nine off‐ and on‐cells of the RVM were recorded in lightly anaesthetized rats. Systemic administration (200 and 300 mg/kg) or PAG microinjection (30, 50 and 100 μg) of LASA caused retardation of the heat‐elicited off‐cell pause, on‐cell burst and the corresponding TF. Neuronal responses and TF retained their mutual time relationship but shifted simultaneously toward longer latencies. This anti‐nociceptive effect of LASA was dose‐dependent, present 5 min after administration and reached a maximum in 30 min for both administration methods. These data confirm that analgesics typically defined as peripherally‐acting, such as metamizol and LASA in this study, may also have an anti‐nociceptive effect by acting directly upon PAG, and suggest that this central effect involves the RVM off‐ and on‐cells.
Visual Learning for an Auditory Secondary Reinforcer by Macaques is Intact after Uncinate Fascicle Section: Indirect Evidence for the Involvement of the Corpus Striatum
Authors: Gaffan D. , Eacott M. J.
Published: April 2006

Three cynomolgus monkeys (Macaca fascicularis) were trained preoperatively in visual discrimination learning for an auditory secondary reinforcer. Each new discrimination problem was solved on the basis of the secondary reinforcer, and primary reinforcement (food reward) was given only after a new problem had been solved. The animals learned 50 new problems in each daily session and it was therefore possible to assess accurately their average rate of learning new discrimination problems in this procedure. After the learning rate had stabilized preoperatively the animals were operated upon to transect the uncinate fascicle, the cortico‐cortical pathway from visual association cortex in the temporal lobe to prefrontal cortex. The animals' learning rate was unchanged after uncinate fascicle section. A previous experiment had shown that visual learning for an auditory secondary reinforcer is unaffected by disconnection of visual association cortex from the amygdala and the fornix. Taken together, this negative evidence points strongly to the conclusion that visual learning for an auditory secondary reinforcer depends upon interaction of temporal lobe visual association cortex with the corpus striatum, since other possibilities have been excluded.
Extracellular Activation of Unitary Excitatory Synapses Between Hippocampal CA3 and CA1 Pyramidal Cells
Authors: Raastad Morten
Published: April 2006

The possibility of regular activation of unitary excitatory synapses on hippocampal CA1 cells by electrical stimulation of Schaffer collaterals was explored in the rat. The amplitude of the excitatory postsynaptic currents (EPSCs) and failures in response to a range of stimulation intensities around the threshold for the smallest detectable EPSC were analysed. After an abrupt appearance of EPSCs in response to increasing stimulation strength, both EPSC amplitude and failure rate could reach a plateau where increasing stimulation intensity did not cause additional responses. This was interpreted as a regular activation of mainly a single axon. Statistical methods showed, however, that only 12 out of ∼50 experiments using threshold stimulation were without significant contamination from additional fibres. In this subset of experiments, upper limits for contamination from other fibres were estimated by using bootstrapping methods. More than 90% of the responses were probably due to faithful activation of a single axon, assuming that the density of axons connecting to one target cell is relatively homogeneous. This result makes the described method suitable for examining some aspects of the transmission between individual hippocampal cells.
Transmitter Release Associated with Long‐term Synaptic Depression in Rat Corticostriatal Slices
Authors: Calabresi Paolo , Fedele Emesto , Pisani Antonio , Fontana Giovanni , Mercuri Nicola B. , Bernardi Giorgio , Raiteri Maurizio
Published: April 2006

Using a corticostriatal slice preparation, we have recently shown that tetanic stimulation of the corticostriatal pathway produces long‐term depression (LTD) of striatal excitatory synaptic transmission. In the present study we have analysed the relationship between LTD and the striatal release of different endogenous transmitters. Samples of perfusate were collected via a small cannula placed just above the surface of the striatal slice close to the recording electrode, and were analysed by HPLC. The high‐frequency stimulation (100 Hz, three trains, 3 s duration, 20 s intervals) used to induce LTD caused a significant but transient increase in the release of both excitatory (aspartate and glutamate) and inhibitory (glycine and GABA) amino acid transmitters. Tetanic stimulation also produced a significant, but transient increase in the release of endogenous dopamine. We conclude that the tetanic stimulation of the corticostriatal pathway is able to induce a large but transient release of excitatory amino acids and of dopamine, whose participation in the induction of striatal LTD has been demonstrated previously. Moreover, the maintenance of this form of synaptic plasticity does not seem to require a sustained change in transmitter release.
Retraction of Spinule‐type Neurites from Carp Retinal Horizontal Cell Dendrites during Dark Adaptation Involves the Activation of Ca2+/Calmodulin‐dependent Protein Kinase II
Authors: Weiler Reto , Schultz Konrad , Janssen‐Bienhold Ulrike
Published: April 2006

The formation of spinules at the terminal dendrites of retinal horizontal cells with the onset of light and their subsequent retraction during darkness is a remarkable example of synaptic plasticity where sensory experience modifies reversibly, and on a time scale of minutes the ultrastructure of synaptic connectivity. The signals and the subsequent intracellular cascades underlying the prominent morphological alterations are only partially understood. We show here that lowering the external calcium concentration did prevent dark‐ and AMPA‐induced retraction of spinules in an eyecup preparation. Furthermore, spinule retraction was prevented in vivo by the injection of calmidazolium, an inhibitor of calmodulin, into the eyeball, and also by the injection of KN‐62, an inhibitor of Ca2+/calmodulin‐dependent protein kinase II (CaMkll). We conclude that local Ca2+ influx through AMPA‐gated channels followed by activation of CaMkll is an important step for spinule retraction during dark adaptation. The phosphorylation patterns of phosphoproteins derived from purified horizontal cells was affected by the inhibitors of camodulin and CaMkll respectively. Some of the affected phosphoproteins appeared to be cytoskeleton‐associated proteins, including GAP‐43. Based on these observations, a putative scenario for the retraction of spinules is proposed.
Differential Formation of Topographic Maps in the Cerebral Cortex and Superior Colliculus of the Mouse by Temporally Correlated Tactile‐Tactile and Tactile‐Visual Inputs
Authors: Benedetti Fabrizio
Published: April 2006

Coincident electrical activity of nerve fibres seems to play a fundamental role in the development of ordered connections in the CNS. To test this hypothesis on the formation of topographic maps we connected two cutaneous regions of the body of newborn mice by implanting an artificial bridge of pig hair. Through this procedure we produced the mechanical fusion of the ear with either the shoulder or the nose. In these conditions the ear not only was connected with shoulder or nose, but was also in relation with the nasal or the inferior portion of visual space. Therefore the probability of temporally correlated tactile‐tactile inputs (earshoulder or ear‐nose) as well as tactile‐visual inputs (ear‐inferior or ear‐nasal visual space) increased. By recording from the primary somatosensory cortex and superior colliculus, we found that the formation of topographic maps was based on different principles. The somatosensory cortex developed in terms of tactiletactile correlated inputs, showing somatosensory neurons with receptive fields extending through the fused parts of the body. Conversely, the superior colliculus processed tactile‐visual correlated inputs; we found somatosensory‐visual bimodal neurons with visual receptive fields extending into the portion of visual space where the artificial bridge was directed. These results suggest that the fusion of two body parts is represented in terms of cutaneous coordinates in the cortex and external world (visual) coordinates in the superior colliculus. Therefore the differential tactile‐tactile and tactile‐visual coincident activity seems to be correlated to the different meaning of information processing of these two brain regions.
Guidance of Thalamocortical Axons by Growth‐promoting Molecules in Developing Rat Cerebral Cortex
Authors: Hübener Mark , Götz Magdalena , Klostermann Stefan , Bolz Jürgen
Published: April 2006

Substrate‐bound guidance cues play an important role during the development of thalamocortical projections. We used time‐lapse video microscopy to study the growth behaviour of thalamic axons on different substrates. On embryonic cortical membranes and on a pure laminin substrate, thalamic fibres advanced relatively slowly (∼15 μm/h) and on average their growth cones retracted transiently every ∼5 h. In contrast, on membranes prepared from early postnatal cortex, thalamic fibres grew twice as fast and spontaneous growth cone collapse occurred ∼8 times less often. Experiments in which we used the sugar‐binding lectin peanut agglutinin or heat inactivation to change the membrane properties indicated that these differences are due to growth‐supporting molecules on postnatal cortical membranes. When offered a choice between embryonic and postnatal cortical membranes, thalamic axons preferred the postnatal membrane substrate. Time‐lapse imaging revealed that borders between these two substrates effectively guided thalamic fibres, and in most cases axons changed their direction without collapse of the growth cone. Our results suggest that thalamic axons can be guided by the spatial distribution of growth‐promoting molecules in the developing cortex.
The Relationship Between Glycine and Gephyrin in Synapses of the Rat Spinal Cord
Authors: Todd A. J. , Spike R. C. , Chong D. , Neilson M.
Published: April 2006

In order to examine the relationship between gephyrin (the peripheral membrane protein associated with glycine receptors) and glycinergic boutons, we have carried out a post‐embedding immunogold study of glycine‐like immunoreactivity on sections of rat lumbar spinal cord which had previously been reacted with monoclonal antibody to gephyrin. In all three areas examined (laminae I and II, lamina III and lamina IX) the majority of profiles which were presynaptic at gephyrin‐immunoreactive synapses were enriched with glycine‐like immunoreactivity. It was estimated that at least 83% of profiles presynaptic to gephyrin‐immunoreactive synapses in the superficial dorsal horn (laminae I and II) were glycine‐immunoreactive, while for lamina III and the ventral horn (lamina IX) the proportions were at least 91% and 98% respectively. This provides strong evidence that glycine is a transmitter at those synapses where gephyrin‐ and glycine‐like immunoreactivities are both present, but suggests that gephyrin may sometimes be expressed at non‐glycinergic synapses and indicates the need for caution in using gephyrin‐immunoreactivity as a marker for glycinergic synapses within the spinal cord. By reacting serial sections of dorsal horn with antisera to glycine and GABA, we have shown that many boutons in laminae I‐III of the dorsal horn show both types of immunoreactivity and are therefore likely to use both amino acids as inhibitory transmitters. Many of the boutons which were presynaptic at axoaxonic synapses in the ventral part of lamina II and in lamina III were glycine‐ and GABA‐immunoreactive and in many cases the postsynaptic element was the central axon of a type II synaptic glomerulus. Taken together with pharmacological evidence, this suggests that inhibitory intemeurons in the dorsal horn which use both GABA and glycine may be important in controlling the flow of information from hair follicle afferents to other spinal neurons.
Morphine Selectively Depresses the Slowest, NMDA‐independent Component of C‐fibre‐evoked Synaptic Activity in the Rat Spinal Cord In Vitro
Authors: Sivilotti L G. , Gerber G. , Rawat B. , Woolf C. J.
Published: April 2006

The effects of morphine on the depolarizing synaptic responses produced in motoneurons by electrical stimulation of primary sensory neurones have been recorded in hemisected spinal cord preparations (8‐ to 12‐day‐old rat pups). Morphine at concentrations of 0.1‐20 μM reduced a slow, long‐lasting (latency greater than 1 s, duration up to 10 s) component of the ventral root potential (VRP) evoked by C‐fibre strength stimulation of the dorsal root. At 2μM the reduction in area of this slow synaptic potential was 71.7 ± 0.9% of control values (n= 15). The earliest components of the C‐fibre strength VRP (the first 100 ms) and the responses to Aβ strength stimuli were unaffected by the opioid even at 10‐20μM. The intermediate, NMDA receptor antagonist (D‐AP5, 40μM)‐sensitive component (which lasts 100‐1000 ms) was reduced by 34 ± 2.2% of control (n= 15), which was significantly less than the reduction of the later NMDA‐independent component (P < 0.001). Morphine (0.1‐20 μM) also depressed the cumulative depolarization generated by the temporal summation of synaptic responses evoked by brief trains of C‐fibre strength stimuli (1 or 10 Hz). A significantly greater reduction at the lower frequency of stimulation (56.3 ± 2.0%) than at the higher (20.3 ± 1.69%, n = 10, measured at 2 μM morphine) was found (P < 0.005). The effects of morphine were reversible upon wash‐out or superfusion with the opioid receptor antagonist naloxone (2 μM). The depression in the C‐fibre‐evoked VRPs produced by the NMDA receptor antagonist D‐AP5 (10‐40 μM) was different from that produced by morphine, as D‐AP5 only reduced a shorter latency component of the VRP (100 ms‐1 s) and was equally effective in decreasing the cumulative depolarizations evoked by 1 and 10 Hz trains. The preferential effect of morphine on the longest latency and longest lasting components of the high‐threshold VRP are discussed in relation to its possible site and mechanism of analgesic action.