bluuebunny/crossref_metadata_2025 huggingface dataset:
Relationship Between Intracellular Free Calcium Concentration and NMDA‐induced Cerebellar Granule Cell Survival In Vitro
Authors: Pearson Heather , Graham Margaret E. , Burgoyne Robert D.
Published: April 2006

The survival of cerebellar granule cells in culture is stimulated by activation of the N‐methyl‐d‐aspartate (NMDA) class of glutamate receptors. Activation of these receptors at the key period for cell survival in vitro (3 days; 3DIV) resulted in a sustained elevation of intracellular free calcium concentration [Ca2+]i over the same concentration range of NMDA that led to granule cell survival. Agents that release Ca2+ from intracellular stores led to only small, transient elevations of [Ca2+]i and were unable to stimulate granule cell survival. Addition of the Ca2+ ionophore ionomycin to granule cell cultures at 3DIV resulted in increased granule cell number at 7DIV. The ability of ionomycin to stimulate granule cell survival was related to the [Ca2+]i elicited, indicating that a rise in [Ca2+]i is sufficient to activate the processes leading to granule cell survival and that the extent of the elevation in [Ca2+]i is crucially important in determining granule cell fate.
Synaptic Currents in Thalamo‐cortical Neurons of the Rat Lateral Geniculate Nucleus
Authors: Leresche N.
Published: April 2006

Thalamo‐cortical neurons were identified in slices of the rat dorsal lateral geniculate nucleus and whole‐cell currents were recorded using the patch‐clamp technique. Postsynaptic currents occurring spontaneously, or elicited by extracellular stimulation in the vicinity of the recorded neuron, were analysed. Spontaneous postsynaptic currents were observed in every recorded neuron. At a holding potential of –60 mV, and with a high internal Cl−, the currents were inward and had amplitudes ranging from <10 to 425 pA. All the spontaneous currents were blocked by 10 μM bicuculline, indicating that they were due to the activation of postsynaptic α‐aminobutyric acid (GABAA) receptors. The 10–90% rise time of these spontaneous GABAergic currents was 0.86 ± 0.19 ms. Their time course of decay could be fitted to an exponential function with one time constant of 18.19 ± 3.02 ms (mean ± SD), or two time constants of 4.47 ± 0.77 and 33.27 ± 3.74 ms. This activity was frequently organized in bursts. Stimulus‐evoked postsynaptic currents were recorded and shown to be due to the activation of glutamatergic receptors. Under similar experimental conditions a bicuculline‐sensitive component was also recorded. These stimulus‐evoked GABAergic currents had a 10–90% rise time of 1.93 ± 0.54 ms. Their time course of decay could also be fitted to an exponential function with one time constant of 24.42 ms or two time constants of 10.26 ± 2.46 and 49.30 ± 10.98 ms. The difference in the time course between spontaneous and evoked GABAergic currents suggests that these responses may arise from synapses having different locations.
An Organotypic Co‐culture of Embryonic Rat Brainstem and Tongue or Skeletal Muscle
Authors: Gueritaud J. P. , Seyfritz N.
Published: April 2006

Transverse sections of rat embryo brainstem (embryonic age 18–19 days) containing the brainstem motor nuclei were explanted, together with a small piece of tongue or skeletal muscle, in a plasma clot and maintained in culture for 2–3 weeks using the roller tube technique. The results show that brainstem motoneurons survived, differentiated and innervated newly formed multinucleated myotubes which displayed large synchronized contractions after 1 week in culture. Muscle fibre contractions could be induced by excitatory amino acid applications and suppressed by curarization. Muscle fibres differentiated normally. During the first week they showed diffuse acetylcholinesterase positivity and multi‐innervation. During the second and third weeks the number of motor end‐plates was greatly reduced and transverse striation was observed. In the presence of muscle fibres, the brainstem thinned out and spread, becoming one or two cell layers thick, and the motoneurons tended to migrate towards the muscle fibres, becoming clearly observable in the living culture. When the muscle explant was not present, the brainstem explant did not spread and remained several cell layers thick, while acetylcholinesterase‐positive cells, presumed to be motoneurons, tended to disappear. The preparation described is well suited for electrophysiological studies of differentiating motoneurons and offers direct access to their dendritic tree, a most desirable feature for patch‐clamp or multisite optical recording.
The L2/HNK‐1 Carbohydrate Epitope is Involved in the Preferential Outgrowth of Motor Neurons on Ventral Roots and Motor Nerves
Authors: Martini R. , Xin Y. , Schmitz B. , Schachner M.
Published: April 2006

Based on the observation that in adult mice the carbohydrate epitope L2/HNK‐1 is detectable on Schwann cells in ventral spinal roots, but only scarcely in dorsal roots (Martini et al., Dev. Biol., 129, 330–338, 1988), the possibility was investigated that the carbohydrate is involved in the outgrowth of regenerating motor neuron axons on peripheral nerve substrates expressing the epitope. To monitor whether the L2 carbohydrate remains present during the time periods in which regenerating axons penetrate the denervated distal nerve stumps, the expression of L2 in motor and sensory branches of the femoral nerve was investigated in normal animals and after a crush lesion. During the first two postoperative weeks, L2 immunoreactivity remained high in the myelinating Schwann cells of the motor branch, whereas L2 immunoreactivity was virtually absent in the sensory branch. In a first experimental approach, cryosections of ventral and dorsal spinal roots and of motor and sensory nerves of adult rats and mice were used as substrates for neurite outgrowth. Neurites of motor neurons from chicken embryos were ∼35% longer after 30 h of maintenance on ventral roots than on dorsal roots. Neurites from sensory neurons had the same length on dorsal as on ventral motors and were as long as neurites from motor neurons grown on dorsal roots. L2 antibodies reduced neurite outgrowth of motor neurons on ventral roots but not on dorsal roots. Neurite outgrowth of sensory neurons on both roots was not altered by the antibodies. Neurite outgrowth of motor neurons on a mixture of the extracellular matrix glycoprotein laminin and the L2 carbohydrate‐carrying glycolipid was significantly higher than on the laminin substrate mixture with GD1b ganglioside or sulphatide. L2 antibodies reduced neurite outgrowth of motor neurons by 50% on the L2 glycolipid, but not on GD1b or sulphatide. These observations indicate that the L2 carbohydrate promotes neurite outgrowth of motor neurons in vitro and may thus contribute to the preferential reinnervation of motor nerves by regenerating motor axons in vivo.
Intrastriatal Dopamine‐rich Implants Reverse the Increase of Dopamine D2 Receptor mRNA Levels Caused by Lesion of the Nigrostriatal Pathway: A Quantitative In Situ Hybridization Study
Authors: Chritin M. , Savasta M. , Mennicken F. , Bal A. , Abrous D. N. , Le Moal M. , Feuerstein C. , Herman J. P.
Published: April 2006

Changes in striatal dopamine D2 receptor mRNA levels provoked by unilateral 6‐hydroxydopamine‐induced lesion of the nigrostriatal dopamine pathway were studied by in situ hybridization. The influence of embryonic dopaminergic neurons implanted into the dopamine‐depleted striatum on the lesion‐induced changes was also examined. Changes in D2 mRNA levels were compared with changes in D2 receptor densities measured in the same animals by receptor autoradiography using [3H]spiperone or [3H]SDZ 205–501 as ligands. The distribution of D2 mRNA in the striatum of control animals closely paralleled that of the D2 receptor itself, as assessed by autoradiography, and the highest density of D2 mRNA occurred in the lateral part of the striatum. One month after lesion, levels of D2 mRNA were 34% higher in the dorsolateral part of the dopamine‐depleted striatum than in the corresponding region of the contralateral control striatum. D2 receptor density in this region was increased by 40% relative to the control level. No significant increases could be measured in the medial part of the striatum. The increases in the lateral part were similar at 7 months post‐lesion; however, at this time the increase in both D2 mRNA and receptor levels had spread to the medial part of the striatum as well. In the graft‐bearing striatum levels of both D2 mRNA and D2 receptors reverted to control levels. This study shows that the post‐lesion increase in striatal dopamine receptor and mRNA level is a biphasic phenomenon with a late‐occurring component in the medial striatum. It also shows that once the increase in striatal D2 receptor gene expression is accomplished, it is maintained unchanged for long periods, similar to that of D2 receptor levels themselves. Moreover, grafts of embryonic dopaminergic neurons are able to modulate the expression of the dopamine D2 receptor gene.
The Relative Contribution of NMDA Receptor Channels in the Expression of Long‐term Potentiation in the Hippocampal CA1 Region
Authors: Asztely Fredrik , Wigström Holger , Gustafsson Bengt
Published: April 2006

Long‐term potentiation (LTP) was studied in the hippocampal CA1 region of guinea‐pigs using a solution containing 0.1 mM magnesium and 10 μM of the non‐N‐methyl‐d‐aspartate (non‐NMDA) antagonist 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (CNQX), leaving an NMDA‐mediated field excitatory postsynaptic potential (EPSP). Brief high‐frequency afferent tetanization induced a substantial synapse‐specific potentiation of the NMDA EPSP with a time course closely resembling that previously described for LTP of the non‐NMDA‐mediated EPSP. This NMDA EPSP potentiation was occluded by prior induction of LTP in normal solution. Using a solution containing 0.1 mM magnesium and 1 μM CNQX, the EPSP was composed of both a non‐NMDA‐ and an NMDA‐mediated component which could be measured separately and in parallel. Manipulations that cause increased transmitter release, such as phorbol ester application and changes in stimulation frequency, enhanced the two measures nearly equally. Afferent tetanization induced an increase of both EPSP components, with a similar time course, the NMDA component showing a relative increase of about one‐third of that of the non‐NMDA one. These results suggest that, to the extent that LTP is based on an increased release of transmitter, the mechanism exhibits features distinct from those underlying other forms of enhanced release.
Adult Retinofugal Axons Regenerating Through Peripheral Nerve Grafts Can Restore the Light‐induced Pupilloconstriction Reflex
Authors: Thanos Solon
Published: April 2006

To investigate the ability of mature regenerating retinal axons to form functional connections within central targets, severed axons were guided into the primary visual centres which subserve the pupillary constriction reflex in response to light. The ocular stump of the transected optic nerve of adult rats was connected by means of an autologous peripheral nerve graft with the pretectal region which contains the relay nucleus of pupillary constriction, the olivary pretectal nucleus. This nucleus is efferently connected with preganglionic neurons in the oculomotor nuclear complex which innervates parasympathetically the muscle constrictors of the iris. Six to sixteen weeks after optic nerve transection and peripheral nerve transplantation, brisk responses were observed in the pupils upon illumination of the transplanted eye. Recovery of the pupil responses indicated that retinal neurons used the peripheral nerve ‘bridge’ to access the pretectum, in which they established synaptic contacts in sufficient density and with appropriate specificity to reconstitute the function of the traumatically interrupted neuronal circuitry.
Regulation of Afferent Connectivity in the Adult Spinal Cord by Nerve Growth Factor
Authors: Lewin Gary R. , Winter Janet , McMahon Stephen B.
Published: April 2006

During development, nerve growth factor (NGF) regulates the density and character of peripheral target innervation (Barde, Neuron, 2, 1525–1534, 1989; Ritter et al., Soc. Neurosci. Abstr., 17, 546.2, 1991); its role in adult animals is less well defined. Here we have asked if the availability of growth factors such as NGF in peripheral tissues can influence the pattern of primary afferent connections in the CNS. Using osmotic minipumps, we raised the levels of NGF in rat skeletal muscle in vivo, a tissue where the levels of this factor are normally very low (Korsching and Thoenen, Proc. Natl. Acad. Sci. USA, 80, 3513–3516, 1983; Shelton and Reichardt, Proc. Natl. Acad. Sci. USA, 81, 7951–7955, 1984; Goedert et al., Mol. Brain Res., 1, 85–92, 1986). After 2 weeks of treatment we asked if the sensory neurons innervating this tissue showed an altered strength and distribution of connections with dorsal horn neurons. The contralateral (vehicle‐treated) muscle, and totally untreated animals, served as controls. In normal and vehicle‐treated animals, electrical stimulation of muscle afferents excited relatively few neurons in the dorsal horn, and these generally showed only weak responses. In contrast, on the NGF‐treated side many more dorsal horn neurons in the lumbar enlargement of the spinal cord were excited by muscle afferents. The increased responsiveness could not be explained by a generalized increase in dorsal horn excitability, since spontaneous activity was not enhanced, nor by a change in A‐fibre‐mediated inhibitions from the treated afferents. Thus, these afferents appeared to establish new synaptic connections or strengthened previously weak ones as a result of increased neurotrophic factor availability. The data suggest that, in the adult rat, the levels of growth factors in peripheral targets may be used to regulate an appropriate degree of afferent connectivity within the central nervous system.
Identification of the Serum Complex Which Induces Cerebellar Granule Cell In Vitro Differentiation and Resistance to Excitatory Amino Acids
Authors: Mercanti D. , Galli C. , Liguori M. , Ciotti M. T. , Gullà P. , Calissano P.
Published: April 2006

The protein complex promoting in vitro terminal differentiation of cerebellar granule cells has been isolated from rabbit serum. We designate the complex the neurite outgrowth and adhesion complex (NOAC). The apparent molecular weight, evaluated by gel filtration, is 80–100 kDa. Rat cerebellar granule cells cultured in NOAC exhibit much lower glial cell contamination and survive, in their differentiated state, much longer than in 10% foetal calf serum. While they bind tetanus toxin, express specific antigens such as synapsin l, synaptophisin and A2B5, and release [3H]d‐aspartate in a fashion similar to that shown by cells cultured in foetal calf serum, they show a 60% reduction in the total number of kainate binding sites. Excitatory amino acid (EAA)‐triggered and depolarization‐stimulated calcium influx, measured in the presence of different agonists, is 50–80% lower in NOAC‐cultured cells. NOAC cells are resistant to excitotoxic stimuli carried by EAAs or by depolarizing treatments with 50 mM KCI or 6 μM veratridine. The marked resistance of NOAC‐cultured neurons to EAAs can be attributed to decreased calcium entry through EAA‐coupled and voltage‐gated calcium channels and possibly to other, as yet unidentified, phenotypic properties of these cells. These findings demonstrate that rabbit serum contains one or more polypeptide(s) endowed with the properties of promoting in vitro survival and differentiation of rat cerebellar granule cells and of conferring an EAA‐resistant phenotype.
Activation of Protein Kinase C Modulates Light Responses in Horizontal Cells of the Turtle Retina
Authors: Akopian Abram , McReynolds John , Weiler Reto
Published: April 2006

The effect of phorbol esters on the light‐evoked responses of horizontal cells were studied in the turtle eyecup preparation. Phorbol esters caused a reduction in receptive field size and a significant decrease in the amplitude of responses to annular and full‐field illumination; however, they caused only minor changes in responses to small spots in the receptive field centre. The dark membrane potential was not affected. The results suggest that phorbol esters may affect both coupling resistance and membrane resistance in horizontal cells. The effects of phorbol esters were blocked by the protein kinase C inhibitor staurosporine, and inactive phorbol ester had no effect, making it very likely that the phorbol ester effects were mediated through activation of protein kinase C. The above effects of the phorbol esters were considerably reduced by the dopamine antagonists haloperidol and fluphenazine, suggesting that they were in part mediated by release of dopamine.
Calcitonin Gene‐related Peptide Causes Intraspinal Spreading of Substance P Released by Peripheral Stimulation
Authors: Schaible H.‐G. , Hope P. J. , Lang C. W. , Duggan A. W.
Published: April 2006

Experiments were performed in barbiturate‐anaesthetized, spinalized cats to investigate the effect of calcitonin gene‐related peptide (CGRP) on the spatial distribution of immunoreactive substance P (ir‐SP) in the spinal cord released by electrical nerve stimulation and noxious mechanical stimuli. The presence of ir‐SP was assessed with microprobes bearing C‐terminus‐directed antibodies to SP. CGRP was microinjected into the grey matter of the spinal cord near microprobe insertion sites at depths of 2500, 2000, 1500 and 1000 μm using minute amounts (in total 0.2–0.5 μl) of Ringer solution containing CGRP at a concentration of 10−5 or 10−3 M. In the untreated cord electrical stimulation of the tibial nerve (suprathreshold for all C fibres) elicited release of ir‐SP which was centred in and around the lamina II. After microinjection of CGRP, stimulation‐associated ir‐SP was detected in a region extending from the cord surface down to the ventral horn. This pattern was similar to that observed after the microinjection of synthetic peptidase inhibitors (Duggan et al., Brain Res., 579, 261–269, 1992). The large expansion of sites accessed by ir‐SP was time‐dependent, reaching a maximal effect within 10–40 min after microinjection of CGRP, and reversal was observed in subsequent probes. A similar expansion of the regions accessed by ir‐SP after microinjection of CGRP was also observed when release of ir‐SP was evoked by noxious mechanical stimulation of the toes. These results indicate that one important function of CGRP in the spinal cord may be the control of the intraspinal sites and neuronal circuits accessed by released substance P, possibly by inhibition of endopeptidases responsible for peptide degradation.
Cycloheximide Reduces the Effects of Anoxic Insult In Vivo and In Vitro
Authors: Papas S. , Crépel V. , Hasboun D. , Jorquera I. , Chinestra P. , Ben‐Ari Y.
Published: April 2006

In vivo and in vitro techniques were utilized to examine the influence of a protein synthesis blocker, cycloheximide (CHX), on the damaging effects of anoxia in the rat. CHX administered 1 h before transient (30 min) forebrain ischaemia increased the survival of animals, decreased body weight loss and reduced the occurrence of delayed degeneration in the CA1 pyramidal region. The same dose of CHX injected 1 h after ischaemia induced status epilepticus, a decrease in survival rate, and did not reduce weight loss or CA1 damage in any of the surviving rats. Electrophysiological techniques were then used to determine the effects of various periods of anoxia and aglycaemia (AA) on CA1 field excitatory postsynaptic potentials (EPSPs) in hippocampal slices incubated in the presence or absence of CHX. In CHX‐treated slices, recuperation of EPSP amplitude (45±16%) was significantly greater than in control slices (9±9%) following an AA episode of 3 min 45 s. No difference was seen in the percent recuperation of EPSPs in the control and CHX‐treated slices after shorter or longer episodes of AA. From these studies, it appears that CHX protects against the damaging effect of ischaemia in vivo or AA in vitro.
On the Distribution of GAP‐43 and its Relation to Serotonin in Adult Monkey and Cat Spinal Cord and Lower Brainstem
Authors: Arvidsson Ulf , Risling Mårten , Cullheim Staffan , Dagerlind Åke , Lindå Hans , Shupliakov Oleg , Ulfhake Brun , Hökfelt Tomas
Published: April 2006

By use of a monoclonal antibody, the distribution of growth‐associated protein (GAP)‐43‐like immunoreactivity (LI) has been studied in the spinal cord of adult grey monkeys (Macaca fascicularis) and adult cats by use of immunofluorescence and peroxidase‐antiperoxidase techniques. The brainstem was also studied with in situ hybridization histochemistry. In both monkeys and cats, a dense innervation of GAP‐43‐immunoreactive (IR) fibres was seen in close apposition to large cell bodies and their processes in the motor nucleus of the ventral horn. Double‐labelling experiments revealed a high degree of coexistence between GAP‐43‐ and 5‐hydroxytryptamine (5‐HT, serotonin)‐LI in the monkey motor nucleus, while in the cat no such colocalization could be verified. At the electron microscopic level, GAP‐43 labelling was seen as a coating of vesicles and axolemma inside the terminals. In both monkey and cat, cell bodies expressing mRNA encoding GAP‐43 were demonstrated in the medullary midline raphe nuclei. A similar location was also encountered for mRNA for aromatic l‐amino acid decarboxylase, an enzyme found in both catecholamine‐ and serotonin‐containing neurons. The present results suggest that GAP‐43 is present in the 5‐HT bulbospinal pathway of the monkey. In the cat, GAP‐43 mRNA‐expressing cell bodies were demonstrated in areas where descending 5‐HT neurons are located, but no convincing colocalization of 5‐HT‐ and GAP‐43‐LI was found at spinal cord levels, despite the existence of extensive fibre networks containing either of the two compounds. Possible explanations for this species discrepancy are discussed. The function of GAP‐43 in nerve terminals impinging on the motoneurons is unknown. However, it may play a role in transmitter release and/or plasticity, since such roles have been proposed for this protein in other systems.
Protein Kinase C Facilitation of Acetylcholine Release at the Rat Neuromuscular Junction
Authors: D'Angelo Egidio , Rossi Paola , Tanzi Franco , Taglietti Vanni
Published: April 2006

Protein kinase C (PKC) is a Ca2+‐dependent enzyme involved in synaptic transmission, which can be experimentally activated by the phorbol ester, phorbol 12‐myristate‐13‐acetate (TPA). We studied the effects of TPA application on acetylcholine (ACh) release at the rat neuromuscular junction by means of the focal recording technique; possible effects of TPA at the postsynaptic site had been ruled out in preliminary studies. In extracellular solutions containing 2 mM Ca2+ and at the stimulation frequency of 0.1 Hz, TPA increased endplate current (EPC) amplitude. In non‐stimulated preparations spontaneous current frequency was increased at a similar rate. The similar time course of TPA action on evoked and spontaneous currents suggests that an increased presynaptic Ca2+ efficacy can be considered to be the probable mechanism of action. The interactions of PKC with ACh release were further investigated. In 0.1 mM Ca2+ extracellular solutions, TPA enhanced evoked currents only at stimulation frequencies (e.g. 40 Hz) that were themselves capable of inducing facilitation. This facilitation is classically associated with presynaptic Ca2+ accumulation, indicating that PKC interacts synergistically with Ca2+ to facilitate ACh release. In particular, since mean quantum size and release probability remained almost unchanged during TPA facilitation, it was concluded that PKC acted by enlarging the immediately available store. Interestingly, TPA also increased the presynaptic currents that were observed to be largely brought about by Ca2+‐dependent K+ currents: evidence was obtained to suggest that increases in these currents provide negative feedback against excess release activation rather than being an expression of enhanced Ca2+ influx.
Survival of Purkinje Cells in Cerebellar Cultures is Increased by Insulin‐like Growth Factor I
Authors: Torres‐Aleman I. , Pons S. , Santos‐Benito F. F.
Published: April 2006

Insulin‐like growth factor I (IGF‐I) is a trophic factor for both neurons and glia. Its presence in the developing and adult cerebellum suggests a role for this growth factor in this area of the brain. Recently, we have described the existence of an IGF‐I‐containing pathway in afferents of Purkinje neurons arising from the inferior olive. In addition, IGF‐I receptors are present in the molecular layer of the cerebellar cortex. These observations prompted us to investigate whether the Purkinje cell is a target for IGF‐I. Addition of IGF‐I to rat cerebellar cultures produced a 7‐fold increase in the number of Purkinje cells (calbindin‐positive) together with an increase in the calbindin content of the cultures. IGF‐I also doubled the number of surviving neurons and produced a moderate, non‐significant increase in [3H]thymidine incorporation by the cultures. On the other hand, basic fibroblast growth factor (bFGF), which is also present in the cerebellum, produced a dramatic increase in both the proportion of astrocytes and in the mitotic activity of the cultures, without affecting neuron survival. We conclude that IGF‐I is a specific promoter of Purkinje cell survival and that its effects differ from those produced by bFGF in fetal cerebellar cultures. These findings reinforce our hypothesis that the Purkinje cell is a target neuron for IGF‐I action in the developing cerebellum.
Diversity of mRNAs in the Axonal Compartment of Peptidergic Neurons in the Rat
Authors: Mohr Evita , Richter Dietmar
Published: April 2006

Vasopressin and oxytocin mRNAs, which are normally translated in the perikarya of magnocellular neurons, have recently been demonstrated to be also present in axons and nerve terminals which are located in the posterior pituitary. The physiological significance of this observation has not yet been resolved. In order to gain further insight into the function and plasticity of the peptidergic neuron the question was addressed whether axonal localization is a unique feature of the above‐mentioned transcripts. Biochemical evidence is presented that magnocellular axons and nerve terminals also contain mRNA species encoding a member of the neurofilament protein family and the prodynorphin precursor. These data imply that axons may harbour a variety of additional protein‐encoding transcripts. Furthermore, it is shown that in the mutant (Brattleboro) rat, which lacks detectable levels of vasopressin but which still transcribes the corresponding gene, axonal vasopressin but not oxytocin mRNA contents are dramatically reduced. Most likely, vasopressin transcripts are absent from the nerve terminals as a consequence of the impaired precursor biosynthesis in the cytoplasm of the mutant rat.
Endothelins Inhibit Junctional Permeability in Cultured Mouse Astrocytes
Authors: Giaume Christian , Cordier Jocelyne , Glowinski Jacques
Published: April 2006

Endothelins, a family of potent vasoconstrictor peptides initially characterized in peripheral tissues, have also been reported to be synthesized in the brain. In this structure several cell types, including astrocytes, endothelial cells and certain neurons, are potential targets for these peptides. In astrocytes, endothelins induce changes in the concentration of several second messengers (calcium, diacylglycerol, arachidonic acid, cAMP) known to be involved in the regulation of gap junction channels. Using the scrape loading/dye transfer technique we have observed that two isoforms of endothelin, endothelin‐1 and endothelin‐3, strongly inhibit the extent of dye‐coupling between confluent astrocytes, suggesting that gap junction permeability was reduced. This inhibitory effect on dye coupling was reproduced by the snake venom sarafotoxin. When used at 10−7 M, these three compounds had inhibitory effects on gap junction channels which were comparable to those induced by the well known uncoupling agents octanol and halothane. In the absence of extracellular calcium, the effects of endothelins were largely prevented, suggesting that second messengers linked to the activation of phospholipases C and/or A2, which both are dependent on external calcium, could be involved in the uncoupling mechanism.
Cell Counts of Purkinje and Inferior Olivary Neurons in the ‘Hyperspiny Purkinje Cells’ Mutant Mouse
Authors: Frederic F. , Hainaut F. , Thomasset M. , Guenet J. L. , Delhaye‐Bouchaud N. , Mariani J.
Published: April 2006

The mutant mouse ‘hyperspiny Purkinje cells’(hpc) has morphologically abnormal Purkinje cells and below normal intracerebellar calbindin‐D28k, a calcium‐binding protein that, in the cerebellum, is found only in the Purkinje cells. We counted the Purkinje cells on serial sections stained with thionin or labelled with anti‐calbindin‐D28k antibodies to investigate whether the depletion of the cerebellar content of calbindin‐D28k was correlated with a reduced number of Purkinje cells. We also counted the inferior olivary neurons, as they are one of the major afferents of the Purkinje cells and also contain calbindin‐D28k. The hpc mutant mice had 27% fewer cerebellar Purkinje cells and 12% fewer inferior olivary neurons than did controls. Their Purkinje cells were evenly immunostained but slightly atrophic. These data suggest that the depleted cerebellar calbindin‐D28k content could be explained both by the loss of some Purkinje cells and the reduced size of the remaining ones.
Ca2+‐Mediated Plateau Potentials in a Subpopulation of Interneurons in the Ventral Horn of the Turtle Spinal Cord
Authors: Hounsgaard J. , Kjaerulff O.
Published: April 2006

The response properties of interneurons in the ventral horn were studied in transverse slices of segments D8 to S2 from the turtle spinal cord, using the current clamp technique. In about half of the neurons the response properties were dominated by their ability to generate plateau potentials. In these cells the plateau potential could account for delayed onset of spiking and a phase of increasing spike frequency during depolarizing current pulses and for a depolarizing afterpotential following the stimulus. The cells usually received monosynaptic and polysynaptic input from the ipsilateral dorsal root and occasionally from the contralateral root. The plateau potential was insensitive to tetrodotoxin but blocked by nifedipine and by replacing Ca2+ with Co2+ in the medium. It is concluded that the response properties of neurons in the ventral horn outside the motor nucleus have differentiated response properties that may well contribute to spinal motor function.
Adult Rat Dorsal Root Ganglion Neurons Extend Neurites on Predegenerated But Not on Normal Peripheral Nerves In Vitro
Authors: Bedi Kuldip S. , Winter Janet , Berry Martin , Cohen James
Published: April 2006

The abilities of embryonic and adult rat sensory neurons to regenerate were compared when cultured on cryostat sections of normal and lesioned sciatic nerve tissues. Differences in neurite growth, visualized by GAP‐43 immunolabelling, were most pronounced on substrata consisting of longitudinal sections of normal versus predegenerated sciatic nerve. Adult dorsal root ganglion (DRG) neurons grew only on the lesioned nerves. Neurites extended along these sections in a characteristically longitudinal orientation, and this growth was not dependent on nerve growth factor. Embryonic DRG neurons extended neurites on sections from both types of nerves. These results highlight important differences in the regenerative abilities of embryonic and adult DRG neurons when grown on physiologically appropriate substrata.
L1 Cell Adhesion Molecule is Expressed by Noradrenergic but not Adrenergic Chromaffin Cells: A Possible Major Role for L1 in Adrenal Medullary Design
Authors: Léon Catherine , Grant Nancy , Aunis Dominique , Langley Keith
Published: April 2006

The adrenal medulla of higher animals is constituted of homotypic groups of chromaffin cells secreting either adrenalin or noradrenalin. Since not all chromaffin cells are individually innervated by fibres of the splanchnic nerve, this tissue characteristic is crucial to the physiological function of the gland. In an attempt to analyse differences between these chromaffin cell types which might underlie the establishment of this tissue pattern, we examined the expression of the adhesion molecule L1 in this gland by immunocytochemistry at the optical and ultrastructural levels in rats. L1, an adhesion molecule abundant in the central nervous system, was found to be present in the adrenal medulla of adults; it was strongly expressed on innervating axons and their surrounding Schwann cells and also on a subpopulation of chromaffin cells. The nature of these chromaffin cells was examined by immunocytochemistry using antibodies against the catecholamine‐synthesizing enzyme phenylethanolamine N‐methyltransferase (PNMT), which are capable of distinguishing between adrenergic and noradrenergic cells. Immunofluorescence labelling of sequential frozen sections demonstrated that chromaffin cells which express L1 do not express PNMT; conversely, L1 was not detected in any chromaffin cells expressing PNMT. Ultrastructural immunocytochemistry confirmed the existence of two non‐overlapping populations of chromaffin cells. It is concluded that, in the adrenal medulla, noradrenergic but not adrenergic chromaffin cells express this adhesion molecule. These data, together with our previous observations that all chromaffin cells express the neural cell adhesion molecule, NCAM, suggest that L1, in cooperation with NCAM, could be responsible for the association of noradrenergic cells in the form of homotypic aggregates segregated from groups of adrenergic cells within the adrenal medulla.
A Hidden Retinal Regenerative Capacity from the C Ciliary Margin is Reactivated In Vitro, that is Accompanied by Down‐regulation of Butyrylcholinesterase
Authors: Willbold Elmar , Layer Paul G.
Published: April 2006

The chicken retina has a capacity to regenerate in vivo, which is restricted up to embryonic day 4 (E4). Here we test the proliferative patterns of dissociated chicken cells from the centre retina or the ciliary margin, including pigmented cells, after their transfer into rotation culture. For central cells in culture, the uptake of [3H]thymidine after a short initial rise decreases similarly to their in ovo counterparts. In contrast, marginal cells that have been shown to regenerate up to E9 into retinotypic stratospheroids re‐enter a novel and long‐lasting phase of in vitro cell division. We have shown previously that cell types of all nuclear layers are produced. Both observations taken together indicate a pronounced self‐renewal of multipotent stem cells. Molecularly, the enzyme butyrylcholinesterase, which in other systems has been shown to mark transitory neuronal cells between proliferation and differentiation, is strongly expressed at the ciliary margin over most of the embryonic period. After these cells are transferred into rotation culture, butyrylcholinesterase is down‐regulated. Concomitantly, the neuronal differentiation marker acetylcholinesterase increases. We conclude that the regenerative capacity of the chick retina is not lost at E4, but rather remains hidden in the chicken ciliary margin, since it can be reactivated In vitro at least up to E9. We suggest that butyrylcholinesterase may be linked to the regulation of stem cell activity.
Squint‐induced Modification of Visual Receptive Field in the Lateral Suprasylvian Cortex of the Cat: Binocul Interaction, Vertical Effect and Anomalous Correspondence
Authors: Sireteanu Ruxandra , Best Johannes
Published: April 2006

In this study, we investigated the receptive field properties of single cells in the lateral suprasylvian area of strabismic cats. More than three times as many cells could be driven by the non‐operated than by the operated eye. Many more cells could be activated by the contralateral than by the ipsilateral eye. In both area 17 and the lateral suprasylvian area, more cells preferred horizontal rather than vertical or oblique orientations (‘vertical effect’). In addition, a small percentage of cells in the lateral suprasylvian area showed an adaptive shift of spatial coordinates; this shift could provide the neural basis of anomalous retinal correspondence.
Baclofen and the Release of Neuropeptides in the Cat Spinal Cord
Authors: Morton C. R. , Hutchison W. D. , Lacey G.
Published: April 2006

The antibody microprobe technique was used to study the effect of baclofen on the release of immunoreactive substance P and immunoreactive calcitonin gene‐related peptide within the lower lumbar spinal cord of pentobarbitone‐anaesthetized spinalized cats. Both peptides were released in the region of the substantia gelatinosa during ipsilateral noxious cutaneous stimulation or high‐intensity electrical stimulation of a hind limb nerve. Intravenous administration of baclofen suppressed the excitation of lumbar dorsal horn neurons, but did not produce detectable alterations of the evoked release of immunoreactive substance P or immunoreactive calcitonin gene‐related peptide in the superficial grey matter dorsal to these neurons. The results suggest that the antinociceptive action of baclofen does not involve a reduction of the intraspinal release of substance P or calcitonin gene‐related peptide from the central terminals of nociceptive sensory fibres.
Immunoreactivity for Taurine Characterizes Subsets of Glia, GABAergic and non‐GABAergic Neurons in the Neo‐ and Archicortex of the Rat, Cat and Rhesus Monkey: Comparison with Immunoreactivity for Homocysteic Acid
Authors: Kritzer M. F. , Cowey A. , Ottersen O. P. , Streit P. , Somogyi P.
Published: April 2006

The cerebral cortex is an area rich in taurine (2‐aminoethanesulphonic acid), but only limited information exists regarding its cellular distribution. We therefore examined taurine‐like immunoreactivity in the cerebral cortex of the rat, cat and macaque monkey using antiserum directed against glutaraldehyde‐conjugated taurine. Immunostaining was assessed at the light and electron microscopic level, and patterns obtained in light microscopic studies were compared to those produced with antiserum to γ‐aminobutyric acid (GABA) and homocysteic acid (HCA). In all three species, strong taurine‐like immunoreactive perivascular endothelial cells, pericytes and oligodendrocytes were found. These cells were located throughout the neuropil, which itself showed a low level of immunoreactivity. In rats and cats, a small number of weakly taurine‐enriched neurons were observed, particularly in superficial layers. In all cortical areas of the macaque, however, glial staining was matched by strong, selective staining of subpopulations of cortical neurons which were distributed in a bilaminar pattern involving layers II/III and VI. In addition, in primary visual cortex, area 17, immunopositive neurons were also present in sublayer IVCβ, while in the hippocampus strongly taurine‐positive neurons were most conspicuous in the granule cell layer of the dentate gyrus. In all regions, strongly taurine‐positive neurons constituted only a subpopulation of the neurons occupying a given layer. Examination of adjacent sections for GABA immunoreactivity showed that the most strongly taurine‐positive neurons in layers II/III were immunoreactive for GABA. The cells located in layers IVCβ and VI, and the granule cells of the dentate gyrus, however, were GABA‐negative. The morphological features of these latter groups suggested that the antiserum to taurine identifies subsets of spiny stellate, small pyramidal and dentate granule cells. None of these neurons showed immunoreactivity with antiserum to HCA in the primate; HCA‐positive glia were found along the pial and white matter boundaries of the cortex, and showed no overlap with strongly taurine‐positive glial elements. Although a transmitter role for taurine may be unlikely, particularly in view of its enrichment in subpopulations of both inhibitory and excitatory cells, the capacity of taurine to influence membrane‐associated functions in excitable tissues, and its selective distribution demonstrated here, provides the potential for a contribution to communication between cortical cells.
Differential Localization of Two Alternatively Spliced GABAA Receptor γ2‐Subunit mRNAs in the Chick Brain
Authors: Glencorse Thora A. , Bateson Alan N. , Darlison Mark G.
Published: April 2006

Two variants of the GABAA receptor γ2 subunit are known to exist, which differ by the presence (γ2L) or absence (γ2S) of eight amino acids in the presumed intracellular loop between the third and fourth membrane‐spanning domains. These variants have been shown to be generated by alternative splicing of the γ2‐subunit primary gene transcript in mouse (Kofuji et at., J. Neurochem., 56, 713–715, 1991), and in bovine and human (Whiting et al., Proc. Natl. Acad. Sci. USA, 87, 9966–9970, 1990) brain. We describe here the cloning, from chick (Gallus domesticus) brain, of cDNAs that encode the γ2L and γ2S subunits, and report on the regional and cellular localization of the corresponding mRNAs as revealed by in situ hybridization histochemistry with transcript‐specific oligonucleotide probes. While the two transcripts are found to be colocalized throughout the chick neuroaxis, certain nuclei (for example, the nucleus isthmi, pars magnocellularis, the nucleus isthmi, pars parvocellularis, the nucleus solitarius and the paleostriatum primitivum) are found to contain predominantly either the γ2S‐ or the γ2L‐subunit mRNA. We conclude that receptors that contain either the γ2S or the γ2L subunit occur, and that these probably have functionally different roles in the modulation of neurotransmission in the central nervous system. In addition, our data indicate that certain cells may produce both transcripts. Consequently, these will have either a single receptor subtype that contains both a γ2S and a γ2L subunit, or two receptor subtypes, one of which contains a γ2S subunit and the other a γ2L subunit.
Distribution of Cutaneous Nociceptive and Tactile Climbing Fibre Input to Sagittal Zones in Cat Cerebellar Anterior Lobe
Authors: Garwicz M. , Ekerot C.‐F. , Schouenborg J.
Published: April 2006

Climbing fibres projecting to the cerebellar C3 zone (and the related C1 and Y zones) receive spatially well organized tactile and nociceptive inputs from the skin. In the present study, cutaneous tactile and nociceptive input to climbing fibres projecting to the X, B, C2 and D1 zones in lobule V were investigated in pentobarbitone‐anaesthetized cats. From the present results and previous studies, it is concluded that the X, C1, CX, C3 and Y zones receive cutaneous nociceptive climbing fibre input. By contrast, climbing fibres to the B, C2 and D1 zones lack cutaneous nociceptive input. Tactile input was found in all zones. The spatial organization of receptive fields of climbing fibres projecting to the X and D1 zones was similar to that in the C3 zone. They were located on the ipsilateral forelimb, mainly its lateral and distal parts, and their proximal borders were located close to joints. In the B zone, more than half of the receptive fields of climbing fibres were confined to the ipsilateral hind‐ or forelimb. However, frequently more than one limb and parts of the trunk were included. In the C2 zone, the majority of climbing fibres had distal ipsi‐ or bilateral receptive fields on the forelimbs, often also including the head/face. Some of the bilateral forelimb receptive fields additionally included the hindlimbs ipsi‐ or bilaterally. The results indicate that each zone has a characteristic set of climbing fibre receptive fields, which is probably related to its efferent control functions.
Somatosensory Trigeminal Projections to the Inferior Olive, Cerebellum and other Precerebellar Nuclei in Rabbits
Authors: Van Ham Jacqueline J. , Yeo Christopher H.
Published: April 2006

We have analysed the pathways through which somatosensory information from the face reaches the inferior olive and the cerebellum in rabbits. We used wheatgerm agglutinin ‐ horseradish peroxidase (WGA‐HRP) to trace projections from all parts of the somatosensory trigeminal system to the olive, cerebellar cortex, the cerebellar deep nuclei and the pontine nuclei. Projections to the cerebellar cortex and inferior olive were verified using retrograde transport of WGA‐HRP. Two regions of the inferior olive–the medial dorsal accessory olive and the ventral leaf of the principal olive–receive inputs from pars interpolaris (Vi) and rostral pars caudalis (Vc) of the spinal trigeminal nucleus and from the principal trigeminal nucleus (Vp). Another area in the caudal medial accessory olive receives inputs from rostral Vo (pars oralis of the spinal trigeminal nucleus), caudal Vi and Vc. There are trigemino‐olivo‐cortical inputs to lobule HVI via all these olivary areas and to the paramedian lobe via the principal olive only. Cerebellar cortex–lobules HVI, crus I and II, paramedian lobe and IX–receives direct mossy fibre inputs from Vp, Vo and rostral Vi. The pontine nuclei receive an input only from rostral Vi. We saw no trigeminal projections to other precerebellar nuclei or to the deep cerebellar nuclei. The concentration of face somatosensory cortical inputs, via several pathways, upon lobule HVI may underlie its important role in the regulation of learned and unlearned eyeblinks.
High Degree of NCAM Diversity Generated by Alternative RNA Splicing in Brain and Muscle
Authors: Barthels Dagmar , Vopper Gaby , Boned Annie , Cremer Harold , Wille Wolfgang
Published: April 2006

The neural cell adhesion molecules (NCAMs) are cell surface glycoproteins involved in vertebrate cell contact formation. Several NCAM mRNA types are generated from a single primary transcript by alternative splicing and differential polyadenylation. In this presentation we analyse sequence heterogeneities within NCAM transcripts detected in the junctions of exons 7/8, 12/13 and 13/14. The highest degree of evolutionary conservation is observed in the 30‐nucleotide insertion (π) between exons 7 and 8, coding for an identical peptide sequence in the mouse, rat and chicken. The most complex splicing pattern is found between exons 12 and 13, called splice site a. Three alternative exons of 15, 48 and 42 nucleotides can be inserted in various combinations, which may also contain the additional trinucleotide AAG. In mouse muscle cell lines, differential ‘extra exon’ and AAG usage in splice site α creates up to 16 NCAM diversity forms, some (if not all) of which are also expressed in mouse brain. Additional microdiversity is generated by the insertion of an alternative AAG trinucleotide in exon junction 13/14. If all combinations of splicing patterns identified so far were to occur and to be translated, there could be up to 192 different NCAM proteins.